WO2022006278A1 - Réduction de la fibrose et traitement de maladies, de troubles et d'états associés - Google Patents

Réduction de la fibrose et traitement de maladies, de troubles et d'états associés Download PDF

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Publication number
WO2022006278A1
WO2022006278A1 PCT/US2021/039899 US2021039899W WO2022006278A1 WO 2022006278 A1 WO2022006278 A1 WO 2022006278A1 US 2021039899 W US2021039899 W US 2021039899W WO 2022006278 A1 WO2022006278 A1 WO 2022006278A1
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compound
patient
mean
population
disease
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PCT/US2021/039899
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English (en)
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Shakil ASLAM
III John Francis NEYLAN
Teresa M. ALVAREZ-DIEZ
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Angion Biomedica Corp.
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4965—Non-condensed pyrazines
    • A61K31/497—Non-condensed pyrazines containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys

Definitions

  • Fibrotic diseases contribute to as much as 45% of deaths in the industrialized world (Wynn, T. A. J. Pathol. 2008;214:199-210). Fibrosis, i.e., excessive tissue scarring, is a common feature of many chronic diseases, as well as acute injuries, where accumulation of scar tissue can lead to loss of organ function and, ultimately, organ failure (Friedman, S. U, et al. Sci. Transl. Med. 2019 Jan 9;5(167):167srl).
  • the present disclosure provides certain technologies for reducing fibrosis.
  • the present disclosure provides certain technologies for reducing fibrosis of, e.g., gastrointestinal tract, heart, kidney, lung, liver, muscle, pancreas, and/or skin.
  • the present disclosure provides methods for treating (e.g., lessening the severity of, such as by delaying onset and/or reducing degree and/or frequency of one or more features of) a disease, disorder, or condition associated with fibrosis.
  • such methods comprise, for example, administering an antifibrotic agent (e.g., a small molecule, such as Compound 1 described herein).
  • an antifibrotic agent e.g., a small molecule, such as Compound 1 described herein.
  • the present disclosure also provides methods of administering an antifibrotic agent (e.g., Compound 1 described herein) to a subject or a population of subjects in need thereof.
  • an antifibrotic agent e.g., Compound 1 described herein
  • the present disclosure encompasses the recognition that particular modes of administering an antifibrotic agent (e.g., Compound 1 described herein) achieve one or more particular effects, e.g., in a population of healthy subjects or in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria.
  • the present disclosure demonstrates that administration of an antifibrotic agent (e.g., Compound 1 described herein) achieves different pharmacokinetic properties (e.g., higher mean Cmax and/or mean AUCo-iast) than what was expected based on standard animal models.
  • the present disclosure demonstrates that an antifibrotic agent (e.g., Compound 1 described herein) is suitable for administration once daily and/or twice daily. In some embodiments, the present disclosure demonstrates that an antifibrotic agent (e.g., Compound 1 described herein) is well-tolerated, e.g., in healthy subjects, e.g., at doses suitable to achieve one or more particular effects (e.g., certain PK parameters).
  • an antifibrotic agent e.g., Compound 1 described herein
  • the present disclosure demonstrates that administration of an antifibrotic agent (e.g., Compound 1 described herein) achieves one or more particular effects (e.g., higher mean Cmax and/or comparable mean AUCo-iast) when administered to subjects in one fed/fasted state, compared to subjects in another fed/fasted state.
  • administration of an antifibrotic agent e.g., Compound 1 described herein
  • achieves different effects e.g., outcomes indicative of reduced fibrosis
  • the present disclosure encompasses the recognition that one or more properties of an antifibrotic agent (e.g., Compound 1 described herein) make it particularly suitable for treating subjects with certain fibrotic diseases or disorders.
  • an antifibrotic agent e.g., Compound 1 described herein
  • the present disclosure demonstrates that Compound 1 may be suitable to treat such subjects, e.g., as indicated by one or more properties described herein.
  • FIG. 1 is a graph of mean Compound 1 blood concentration over time from SAD cohorts in a fasted state.
  • FIG. 2 is a graph of mean Compound 1 plasma concentration over time in SAD food effect cohorts receiving 200 mg Compound 1.
  • FIG. 3 depicts a Western analysis demonstrating inhibition of phosphorylation of PDGFRp at tyrosine 751 by Compound 1 in serum-starved hepatic stellate cells.
  • FIG. 4 depicts a Western analysis demonstrating inhibition of phosphorylation of VEGFR2 at tyrosine 1175 by Compound 1 in serum-starved HUVEC cells.
  • FIG. 5 depicts results of a KINOMEscan profile of Compound 1 in small airway epithelial cells and lung fibroblasts (SAEMyF); lung fibroblasts only (MyoF); and renal proximal tubular epithelial cells and lung fibroblasts (REMyoF).
  • SAEMyF small airway epithelial cells and lung fibroblasts
  • MyoF lung fibroblasts only
  • REMyoF renal proximal tubular epithelial cells and lung fibroblasts
  • FIG. 6 is a graph showing soluble collagen concentrations in normal human lung fibroblasts (NHLF) treated with TGFpi and Compound 1.
  • FIG. 7A is a graph showing lung-to-body weight ratio of mice treated with Compound 1 in a bleomycin-induced injury model of idiopathic pulmonary fibrosis (IPF).
  • FIG. 7B is a graph showing hydroxyproline levels in mice treated with Compound 1 in a bleomycin- induced injury model of IPF.
  • FIG. 7C is a graph showing histopathological damage in lung tissue sections using the Ashcroft scale from mice treated with Compound 1 in a bleomycin- induced injury model of IPF.
  • FIG. 7D is a graph showing extent of picrosirius red staining in lung tissue sections from mice treated with Compound 1 in a bleomycin-induced injury model of IPF.
  • FIG. 7E is a graph showing extent of immunohistological staining for TGFpi in lung tissue sections from mice treated with Compound 1 in a bleomycin-induced injury model of IPF.
  • FIG. 8A is a graph showing lung fibrosis score using Ashcroft scale based on H&E staining in lung tissue sections from mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 8B is a graph hydroxyproline levels in mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 8C is a graph showing extent of picrosirius red staining in lung tissue sections from mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 8A is a graph showing lung fibrosis score using Ashcroft scale based on H&E staining in lung tissue sections from mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 8B is a graph hydroxyproline levels in mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 8C is
  • FIG. 8D is a graph showing extent of aSMA staining in lung tissue sections from mice treated with Compound 1 in an inducible TGFpi mouse model of lung fibrosis.
  • FIG. 9A is a graph showing dermal thickness, measured as the distance between epidermal-dermal junction and dermal-subcutaneous fat junction in H&E-stained slides prepared from a skin biopsy from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9B is a graph showing skin fibrotic score, judged based on H&E-stained slides prepared from a skin biopsy from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9A is a graph showing dermal thickness, measured as the distance between epidermal-dermal junction and dermal-subcutaneous fat junction in H&E-stained slides prepared from a skin biopsy from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9B is
  • FIG. 9C is a graph showing lung hydroxyproline levels from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9D is a graph showing lung fibrosis score using Ashcroft scale based on H&E staining in lung tissue sections from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9E is a graph showing kidney hydroxyproline levels from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 9F is a graph showing extent of picrosirius red staining in kidney tissue sections from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 10A is a graph showing urine protein levels in rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 10B is a graph showing intraperitoneal fluid volume in rats treated with Compound 1 in a FSGS-relevant model of PAN- induced proteinuria.
  • FIG. IOC is a graph showing GFR, measured using FITC-sinistrin decay kinetics, in rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 10D is a graph showing glomerular diameter, measured using histopathological analysis of P AS-stained renal coronal sections from rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 10E is a graph showing collagen-3 expression, determined in glomeruli using quantitative immunohistochemistry, in rats treated with Compound 1 in a FSGS- relevant model of PAN-induced proteinuria.
  • FIG. 11A is a graph showing urine protein levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 1 IB is a graph showing urine albumin levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. llC is a graph showing urine albumin to creatinine ratio in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 1 ID is a graph showing urine kidney injury molecule-1 (KIM1) levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • KIM1 urine kidney injury molecule-1
  • FIG. 11E is a graph showing renal hydroxyproline levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 1 IF is a graph showing renal damage scores, based on H&E stained slides of kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 11G is a graph showing extent of picrosirius red staining from slides of kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 11H is a graph showing extent of aSMA staining in kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • EIUO unilateral ureteral obstruction
  • FIG. 13A is a graph showing serum creatinine (SCr) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13B is a graph showing BUN levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13C is a graph showing kidney weight at sacrifice in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13D is a graph showing kidney weight as a percentage of body weight at sacrifice rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13E is a graph showing kidney hydroxyproline levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13F is a graph showing cystic index, quantified in H&E stained kidney tissue sections using digital planimetry, in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13G is a graph showing 24-hour urine volume in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13H is a graph showing urine protein levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 131 is a graph showing urine albumin levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13J is a graph showing urine neutrophil gelatinase-associated lipocalin (NGAL) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13K is a graph showing urine KIM1 levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13L is a graph showing urine interleukin 18 (IL18) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 13M is a graph showing urine cystatin C levels in in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 14A is a graph showing body weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14B is a graph showing colon weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14C is a graph showing colon length after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14D is a graph showing colon damage score in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14A is a graph showing body weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14B is a graph showing colon weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14C is a graph showing
  • FIG. 14E is a graph showing colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis. Each of (a) colon architecture, (b) degree of inflammatory cell infiltration, (c) muscle thickening, and (d) crypt damage and goblet cells loss was assessed.
  • FIG. 14F is a graph showing composite colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14E is a graph showing colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14G is a graph showing extent of Alcian blue staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 14H is a graph showing extent of myeloperoxidase staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 141 is a graph showing extent of F4/80 staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 15A is a graph showing body weight in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15B is a graph showing colon weight in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15C is a graph showing colon length in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15D is a graph showing gross morphological colon damage score in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15E is a graph showing colon length in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15F is a graph showing gross morphological colon damage score in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15G is a graph showing histopathological colon damage score in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 15H is a graph showing extent of Alcian blue staining in slides of colon tissue sections from mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 16A is a graph showing colon length in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 16B is a graph showing macroscopic colon damage score in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 16C is a graph showing colon hydroxyproline levels in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 16D is a graph showing histopathological injury score from H&E-stained slides of colon tissue section from mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 17A is a graph showing lung hydroxyproline levels in mice treated with Compound 1 or pirfenidone (PFD) in a bleomycin-induce lung fibrosis model.
  • FIG. 17B is a graph showing Ashcroft lung histopathological score in lung tissue sections from mice treated with Compound 1 or pirfenidone (PFD) in a bleomycin-induce lung fibrosis model.
  • FIG. 17C is a graph showing extent of picrosirius red staining in lung tissue sections from mice treated with Compound 1 or pirfenidone (PFD) in a bleomycin-induce lung fibrosis model.
  • FIG. 17D is a graph showing extent of TGFpi staining in lung tissue sections from mice treated with Compound 1 or pirfenidone (PFD) in a bleomycin-induce lung fibrosis model.
  • FIG. 18 provides a cartoon schematic of varying relationships between genomic responses and kidney disease outcomes.
  • FIG. 19 provides proteinuria of PAN treated model rats treated with Compound 1.
  • FIG. 20 provides fold change in expression for COL1 A1 mRNA rat PAN model with treatment of Compound 1.
  • FIG. 21 provides fold change in expression for COL3 A1 mRNA rat PAN model with treatment of Compound 1.
  • FIG. 22 depicts a Compound 1 interactome built using in vitro activity data.
  • FIG. 23 provides urine protein levels in PAN treated model rats.
  • FIG. 24A provides COL3 A1 mRNA levels in PAN-treated model rats.
  • FIG. 24B provides an association of COL3 A1 mRNA levels and urine protein values.
  • FIG. 25A depicts a glomerular COL3 A1 interactome.
  • FIG. 25B depicts a tubular COL3A1 interactome.
  • FIG. 26A is a graph showing that, prior to randomization, proteinuria of Alport mice is elevated compared to wild-type mice.
  • FIG. 26B is a Kaplan Myer survival graph of Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26C and FIG. 26D are graphs of body weight and kidney weight, respectively, for mice in an Alport model study.
  • FIG. 26E is a graph of proteinuria in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26F is a graph of protein-to-creatinine ratio in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • 26G is a graph of serum BUN levels in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26H is a graph of serum creatinine levels in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 261 is a graph of kidney hydroxyproline levels in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26 J is a graph of kidney injury scores based on H&E stained kidney samples in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26K is a graph of renal fibrosis scores based on Masson’s Trichome- stained kidney samples in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • 26L is a graph of IHC staining for TGFpi in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26M is a graph of IHC staining for aSMA in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26N is a graph of IHC staining for Collagen- 1 in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 260 is a graph of picrosirius red staining for collagen in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 26P depicts a Western blot analysis for collagen 1, aSMA, TGFpi, and PDGFR in kidney tissue samples from sacrificed Alport mice.
  • FIG. 26Q is a graph of densitometric analysis of a Western blot for collagen 1, aSMA, TGFpi, and PDGFR in Alport mice treated with Compound 1 vs. Alport mice treated with vehicle.
  • FIG. 27A is a Kaplan Meier curve showing survival analysis of rats in a Passive Heymann nephritis model.
  • FIG. 27B is a graph of body mass of rats in a Passive Heymann nephritis model.
  • FIG. 27C is a graph of kidney mass of rats in a Passive Heymann nephritis model.
  • FIG. 27D is a graph of kidney mass to body mass ratio of rats in a Passive Heymann nephritis model.
  • FIG. 27E is a graph of protein-to-creatinine ratio (PCR) of rats in a Passive Heymann nephritis model.
  • FIG. 27F is a graph plotting PCR of rats in a Passive Heymann nephritis model from Day 7 until end of study.
  • FIG. 27G is a graph of serum cholesterol of rats in a Passive Heymann nephritis model.
  • FIG. 27H is a graph of serum triglycerides of rats in a Passive Heymann nephritis model.
  • FIG. 271 is a graph of serum creatinine of rats in a Passive Heymann nephritis model.
  • FIG. 27J is a graph of BUN of rats in a Passive Heymann nephritis model.
  • FIG. 27K is a graph of hydroxyproline content per kidney in rats in a Passive Heymann nephritis model.
  • FIG. 27L depicts representative Trichrome-stained kidney tissue samples from rats in a Passive Heymann nephritis model.
  • FIG. 27M depicts representative PAS-stained kidney tissue samples from rats in a Passive Heymann nephritis model.
  • FIG. 27N is a graph of glomerular damage score in rats in a Passive Heymann nephritis model.
  • FIG. 270 is a Western blot analysis on total kidney lysates from rats in a Passive Heymann nephritis model.
  • FIG. 27P is a graph of PDGFRP levels, normalized to GAPDH, from rats in a Passive Heymann nephritis model.
  • FIG. 28 summarizes pre-screening and screening processes for a Phase 2
  • FIG. 29 is a graph of mean oral blood PK profile of Compound 1 over time from SAD cohorts in a fasted state.
  • FIG. 30A is a graph of mean oral blood PK profile of Compound 1 over time from BID MAD cohorts in a fasted state at Day 1.
  • FIG. 30B is a graph of mean oral blood PK profile of Compound 1 over time from BID MAD cohorts in a fasted state at Day 14.
  • FIG. 31 is a graph of mean Compound 1 blood concentration vs. time profiles at steady state for QD MAD cohorts in the fed state at Day 14.
  • FIG. 32 is a graph of Compound 1 blood concentration vs. time in a food effect cross-over study with a 600 mg single dose.
  • administering typically refers to administration of a composition to a subject to achieve delivery of an active agent to a site of interest (e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.)
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • one or more particular routes of administration may be feasible and/or useful in the practice of the present disclosure.
  • administration may be parenteral
  • the term “comparable” refers to two or more agents, entities, situations, sets of conditions, circumstances, individuals, or populations, etc. that may not be identical to one another but that are sufficiently similar to permit comparison there between so that one skilled in the art will appreciate that conclusions may reasonably be drawn based on differences or similarities observed.
  • comparable agents, entities, situations, sets of conditions, circumstances, individuals, or populations are characterized by a plurality of substantially identical features and one or a small number of varied features.
  • dosage form may be used to refer to a physically discrete unit of an active agent (e.g., a therapeutic agent) for administration to a subject.
  • an active agent e.g., a therapeutic agent
  • each such unit contains a predetermined quantity of active agent.
  • such quantity is a unit dosage amount (or a whole fraction thereof) appropriate for administration in accordance with a dosing regimen that has been determined to correlate with a desired or beneficial outcome when administered to a relevant population (i.e., with a therapeutic dosing regimen).
  • the total amount of a therapeutic composition or agent administered to a particular subject is determined by one or more attending physicians and may involve administration of multiple dosage forms.
  • salt form refers to a form of a relevant compound as a salt appropriate for use in pharmaceutical contexts, i.e., salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and/or lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describes pharmaceutically acceptable salts in detail in ./. Pharmaceutical Sciences , 66: 1-19 (1977).
  • the term “reference” describes a standard or control relative to which a comparison is performed. For example, in some embodiments, an agent, individual, population, sample, sequence or value of interest is compared with a reference or control agent, individual, population, sample, sequence or value. In some embodiments, a reference or control is tested and/or determined substantially simultaneously with the testing or determination of interest. In some embodiments, a reference or control is a historical reference or control, optionally embodied in a tangible medium. Typically, as would be understood by those skilled in the art, a reference or control is determined or characterized under comparable conditions or circumstances to those under assessment. Those skilled in the art will appreciate when sufficient similarities are present to justify reliance on and/or comparison to a particular possible reference or control.
  • risk of a disease, disorder, and/or condition refers to a likelihood that a particular individual will develop the disease, disorder, and/or condition. In some embodiments, risk is expressed as a percentage. In some embodiments, risk is from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed as a risk relative to a risk associated with a reference sample or group of reference samples. In some embodiments, a reference sample or group of reference samples have a known risk of a disease, disorder, condition and/or event. In some embodiments a reference sample or group of reference samples are from individuals comparable to a particular individual. In some embodiments, relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.
  • the term “subject” refers an organism, typically a mammal (e.g., a human). In some embodiments, a subject is suffering from a relevant disease, disorder or condition. In some embodiments, a human subject is an adult, adolescent, or pediatric subject.
  • a subject is at risk of (e.g., susceptible to), e.g., at elevated risk of relative to an appropriate control individual or population thereof, a disease, disorder, or condition.
  • a subject displays one or more symptoms or characteristics of a disease, disorder or condition.
  • a subject does not display any symptom or characteristic of a disease, disorder, or condition.
  • a subject is someone with one or more features characteristic of susceptibility to or risk of a disease, disorder, or condition.
  • a subject is an individual to whom diagnosis and/or therapy and/or prophylaxis is and/or has been administered.
  • the terms “subject” and “patient” are used interchangeably herein.
  • the term “treat” refers to any administration of a therapy that partially or completely alleviates, ameliorates, relieves, inhibits, delays onset of, reduces severity of, and/or reduces incidence of one or more symptoms, features, and/or causes of a particular disease, disorder, and/or condition.
  • such treatment may be of a subject who does not exhibit signs of the relevant disease, disorder and/or condition and/or of a subject who exhibits only early signs of the disease, disorder, and/or condition.
  • such treatment may be of a subject who exhibits one or more established signs of the relevant disease, disorder and/or condition.
  • treatment may be of a subject who has been diagnosed as suffering from the relevant disease, disorder, and/or condition.
  • Compound 1 is in a pharmacological class of tyrosine kinase inhibitors (TKI).
  • TKI tyrosine kinase inhibitors
  • Compound 1 is an orally bioavailable small molecule dual kinase inhibitor of platelet-derived growth factor receptors (PDGFR) and vascular endothelial growth factor receptors (VEGFR2).
  • PDGFR platelet-derived growth factor receptors
  • VEGFR2 vascular endothelial growth factor receptors
  • Compound 1 is provided and/or utilized (e.g., for inclusion in, e.g., during one or more steps of manufacturing of, a composition and/or for delivery to a subject) in accordance with the present disclosure in a form such as a salt form (e.g., a pharmaceutically acceptable salt form).
  • a salt form e.g., a pharmaceutically acceptable salt form
  • pharmaceutically acceptable salt forms are well known in the art.
  • Compound 1 is provided and/or utilized in a hydrochloride salt form, a maleate salt form, a mesylate salt form, or a tosylate salt form.
  • Compound 1 is provided and/or utilized (e.g., for inclusion in, e.g., during one or more steps of manufacturing of, a composition and/or for delivery to a subject) in accordance with the present disclosure in a form such as a solid form.
  • Compound 1 is provided and/or utilized in accordance with the present disclosure in an amorphous solid form, in a crystalline solid form, or in a mixture thereof.
  • a crystalline solid form may be or comprise a solvate, hydrate, or an unsolvated form. The use of any and all such forms are contemplated by the present disclosure.
  • Compound 1 is provided and/or utilized (e.g., for inclusion in, e.g., during one or more steps of manufacturing of, a composition and/or for delivery to a subject) as a hydrochloride salt form (e.g., a channel hydrate comprising up to about 4 equivalents of water, up to about 3 equivalents of water, up to about 2 equivalents of water, or up to about 1 equivalent of water).
  • a hydrochloride salt form of Compound 1 is characterized by one or more peaks in its XRPD selected from those at about one or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta.
  • such a hydrochloride salt form of Compound 1 is characterized by two or more peaks in its XRPD selected from those at about one or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta. In some embodiments, such a hydrochloride salt form of Compound 1 is characterized by one or more peaks in its XRPD selected from those at about three or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta.
  • such a hydrochloride salt form of Compound 1 is characterized by the following peaks in its XRPD pattern at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta. In some embodiments, such a hydrochloride salt form of Compound 1 is characterized by peaks in its XRPD pattern at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta, corresponding to d- spacing of about 16.74, 8.33, 7.67, 5.20, and 4.23 angstroms, respectively.
  • such a hydrochloride salt form of Compound 1 is characterized by substantially all of the peaks in its XRPD pattern selected from 5.28, 5.76, 10.63, 11.54, 12.73, 13.13, 14.08, 15.34, 15.64, 16.00, 16.55, 17.05, 17.78, 18.86, 19.08, 20.16, 20.68, 20.98, 21.62, 22.05, 22.82, 23.97, 24.94, 25.23, 25.61, 26.58, 27.01, 27.78, 29.89, 30.51, 30.91, 31.72, and 33.39 degrees 2- theta.
  • Compound 1 refers to Compound 1 (i.e., methyl (Z)-3-(((4-(N-methyl-2-(4-methylpiperazin-l- yl)acetamido)phenyl)amino)(phenyl)methylene)-2-oxo-2,3-dihydro-lH-pyrrolo[2,3-b]pyridine- 6-carboxylate) in any available form, such as, e.g., a salt form and/or solid form. It will be understood, therefore, that reference to an amount (e.g., in mg) of Compound 1 means the amount of Compound 1 in free base form.
  • Compound 1 may be provided and/or utilized as, e.g., a salt form of Compound 1 such that the amount of the salt (or other form) is an amount that corresponds to the “free base equivalent” of Compound 1.
  • “50 mg Compound 1” means, e.g., approx. 53.4 mg of Compound 1 Hydrochloride anhydrate, approx. 58.4 mg of Compound 1 Hydrochloride trihydrate, and approx. 58.9 mg of Compound 1 Mesylate anhydrate, etc.
  • compositions that delivers Compound 1 e.g., in some embodiments, a composition that is or comprises Compound 1, or a composition that otherwise delivers Compound 1 - e.g., that is or comprises a prodrug of Compound 1, a complex or other entity that releases Compound 1 upon administration, etc.
  • a composition that delivers Compound 1 e.g., in some embodiments, a composition that is or comprises Compound 1, or a composition that otherwise delivers Compound 1 - e.g., that is or comprises a prodrug of Compound 1, a complex or other entity that releases Compound 1 upon administration, etc.
  • the present disclosure provides methods of administering Compound 1 to a subject or population of subjects, according to a regimen established to achieve one or more particular effects.
  • a regimen has been established to achieve one or more particular effects, relative to that observed for a comparable reference population that has not received Compound 1 (e.g., that has received a reference composition which does not deliver Compound 1).
  • a regimen has been established to achieve one or more particular effects in a population of healthy subjects.
  • a regimen has been established to achieve one or more particular effects in a population of healthy subjects, relative to that observed for a comparable reference population that has not received Compound 1 (e.g., that has received a reference composition which does not deliver Compound 1).
  • certain parameters may be evaluated to determine if a particular effect is achieved. For example, in some embodiments, certain measures of, e.g., TEAEs, Cmax, Tmax, AUCo-iast, AUCo-inf, Vz/F, CL/F, and/or ti/2, can be obtained. Additionally, in some embodiments, certain measure of, e.g., Tiast, AUCo-12, AUCo-24, C12, C24, Ctrough, RaccCmax, RaccAUC, CLss/F, and/or Vz ss /F, etc. can be obtained.
  • certain measures of e.g., TEAEs, Cmax, Tmax, AUCo-iast, AUCo-inf, Vz/F, CL/F, and/or ti/2, can be obtained.
  • a reference population has not received a composition providing Compound 1.
  • a reference population has received an otherwise comparable reference composition that does not provide Compound 1 (e.g., a placebo and/or another therapy).
  • a placebo as used herein is a dosage form that matches that of an active study compound, but does not deliver the active study compound (e.g., Compound 1).
  • a placebo can be a capsule that visually matches an active study drug and is composed of the same capsule shell but is filled with a pharmaceutical excipient (and lacking the active study drug), e.g., silicified microcrystalline cellulose.
  • a reference composition may be or may have been administered at the same intervals and/or in the same amounts as a composition providing Compound 1.
  • composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, e.g., at a particular time point.
  • a composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, e.g., at a particular time point, relative to an appropriate reference as described herein (e.g., as is observed in a comparable population who has not received a composition providing Compound 1).
  • the present disclosure provides methods of administering to a subject or population of subjects in need thereof an amount of Compound 1, wherein said amount, when administered as a single oral dose to a population of healthy subjects (e.g., healthy subjects in a fed or fasted state), was established to achieve one or more particular effects.
  • a particular effect is or comprises a particular mean maximum concentration (Cmax).
  • Cmax mean maximum concentration
  • a mean Cmax is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a mean Cmax is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • QD once daily
  • BID twice daily
  • a particular effect is or comprises a mean Cmax of from about 30 ng/mL to about 800 ng/mL, from about 85 ng/mL to about 800 ng/mL, from about 230 ng/mL to about 800 ng/mL, from about 400 ng/mL to about 800 ng/mL, from about 30 ng/mL to about 400 ng/mL, or from about 85 ng/mL to about 400 ng/mL.
  • a particular effect is or comprises a mean Cmax of less than about 800 ng/mL, less than about 400 ng/mL, less than about 235 ng/mL, or less than about 90 ng/mL.
  • a particular effect is or comprises a mean Cmax of greater than about 30 ng/mL, greater than about 85 ng/mL, greater than about 230 ng/mL, or greater than about 400 ng/mL.
  • a particular effect is or comprises a particular median time to maximum concentration (Tmax).
  • Tmax is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a median Tmax is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • QD once daily
  • BID twice daily
  • a particular effect is or comprises a median Tmax of from about 1 hour to about 4 hours, from about 1 hour to about 2 hours, or from about 1 hour to about 1.5 hours. In some embodiments, a particular effect is or comprises a median Tmax of from about 0.5 hours to about 4 hours, from about 1 hour to about 4 hours, from about 1 hour to about 2 hours, or from about 1 hour to about 1.5 hours. In some embodiments, a particular effect is or comprises a median Tmax of less than about 4 hours, less than about 2 hours, or less than about 1.5 hours. In some embodiments, a particular effect is or comprises a median Tmax of greater than about 1 hour, greater than about 1.5 hours, or greater than about 2 hours.
  • a particular effect is or comprises a particular mean area under drug concentration-time curve (AUCo-iast).
  • AUCo-iast mean area under drug concentration-time curve
  • a mean AUCo-iast is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a particular effect is or comprises a mean AUCo-iast of from about 60 ng*h/mL to about 4000 ng*h/mL, from about 60 ng*h/mL to about 2200 ng*h/mL, from about 170 ng*h/mL to about 2200 ng*h/mL, from about 400 ng*h/mL to about 2200 ng*h/mL, from about 1260 ng*h/mL to about 2200 ng*h/mL, from about 60 ng*h/mL to about 1260 ng*h/mL, or from about 170 ng*h/mL to about 1260 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-iast of from about 100 ng*h/mL to about 4000 ng*h/mL, from about 1000 ng*h/mL to about 2400 ng*h/mL, from about 180 ng*h/mL to about 2400 ng*h/mL, from about 430 ng*h/mL to about 2400 ng*h/mL, from about 1260 ng*h/mL to about 2400 ng*h/mL, from about 100 ng*h/mL to about 1260 ng*h/mL, or from about 180 ng*h/mL to about 1260 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-iast of less than about 4000 ng*h/mL, less than about 2200 ng*h/mL, less than about 1270 ng*h/mL, less than about 420 ng*h/mL, or less than about 180 ng*h/mL. In some embodiments, a particular effect is or comprises a mean AUCo-iast of less than about 4000 ng*h/mL, less than about 2400 ng*h/mL, less than about 1270 ng*h/mL, less than about 440 ng*h/mL, or less than about 190 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-iast of greater than about 60 ng*h/mL, greater than about 170 ng*h/mL, greater than about 400 ng*h/mL, or greater than about 1260 ng*h/mL.
  • a particular effect is or comprises a particular mean area under drug concentration-time curve (AUCo-inf).
  • AUCo-inf mean area under drug concentration-time curve
  • a mean AUCo-inf is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a particular effect is or comprises a mean AUCo-inf of from about 60 ng*h/mL to about 4000 ng*h/mL, from about 60 ng*h/mL to about 2410 ng*h/mL, from about 190 ng*h/mL to about 2410 ng*h/mL, from about 490 ng*h/mL to about 2410 ng*h/mL, from about 1300 ng*h/mL to about 2410 ng*h/mL, or from about 190 ng*h/mL to about 1300 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-inf of from about 60 ng*h/mL to about 4000 ng*h/mL, from about 60 ng*h/mL to about 2580 ng*h/mL, from about 180 ng*h/mL to about 2580 ng*h/mL, from about 550 ng*h/mL to about 2580 ng*h/mL, from about 1300 ng*h/mL to about 2580 ng*h/mL, or from about 180 ng*h/mL to about 1300 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-inf of less than about 4000 ng*h/mL, less than about 2410 ng*h/mL, less than about 1300 ng*h/mL, less than about 500 ng*h/mL, or less than about 200 ng*h/mL. In some embodiments, a particular effect is or comprises a mean AUCo-inf of less than about 4000 ng*h/mL, less than about 2580 ng*h/mL, less than about 1310 ng*h/mL, less than about 560 ng*h/mL, or less than about 190 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-inf of greater than about 60 ng*h/mL, greater than about 190 ng*h/mL, greater than about 490 ng*h/mL, or greater than about 1300 ng*h/mL.
  • a particular effect is or comprises a particular mean area under drug concentration-time curve (AUCo-12).
  • AUCo-12 is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • QD once daily
  • BID twice daily
  • a particular effect is or comprises a mean AUCo-12 of from about 100 ng*h/mL to about 1500 ng*h/mL, from about 120 ng*h/mL to about 1340 ng*h/mL, from about 250 ng*h/mL to about 1340 ng*h/mL, from about 920 ng*h/mL to about 1340 ng*h/mL, from about 120 ng*h/mL to about 930 ng*h/mL, or from about 250 ng*h/mL to about 930 ng*h/mL.
  • a particular effect is or comprises a mean AUCo-12 of less than about 1500 ng*h/mL, less than about 1300 ng*h/mL, less than about 930 ng*h/mL, or less than about 260 ng*h/mL. In some embodiments, a particular effect is or comprises a mean AUCo-12 of greater than about 115 ng*h/mL, greater than about 250 ng*h/mL, or greater than about 920 ng*h/mL.
  • a particular effect is or comprises a particular mean apparent volume of distribution (Vz/F).
  • a mean Vz/F is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a particular effect is or comprises a mean Vz/F of from about 2000 L to about 5500 L, from about 2200 L to about 5500 L, or from about 2100 L to about 2300 L.
  • a particular effect is or comprises a mean Vz/F of from about 2000 L to about 7800 L, from about 2200 L to about 5500 L, or from about 5400 L to about 7800 L.
  • a particular effect is or comprises a mean Vz/F of less than about 5500 L, less than about 2300 L, or less than about 2200 L. In some embodiments, a particular effect is or comprises a mean Vz/F of less than about 7800L, less than about 5500 L, less than about 2300 L, or less than about 2200 L. In some embodiments, a particular effect is or comprises a mean Vz/F of greater than about 2100 L, greater than about 2200 L, or greater than about 5100 L.
  • a particular effect is or comprises a particular mean apparent total clearance (CL/F).
  • a mean CL/F is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a particular effect is or comprises a mean CL/F of from about 300 L/h to about 600 L/h, from about 300 L/h to about 450 L/h, or from about 400 L/h to about 600 L/h.
  • a particular effect is or comprises a mean CL/F of from about 270 L/h to about 590 L/h, from about 270 L/h to about 370 L/h, or from about 340 L/h to about 590 L/h. In some embodiments, a particular effect is or comprises a mean CL/F of less than about 600 L/h, less than about 450 L/h, or less than about 350 L/h. In some embodiments, a particular effect is or comprises a mean CL/F of greater than about 300 L/h, greater than about 400 L/h, or greater than about 575 L/h. In some embodiments, a particular effect is or comprises a mean CL/F of greater than about 250 L/h, greater than about 340 L/h, or greater than about 370 L/h.
  • a particular effect is or comprises a particular mean half-life (ti/2).
  • a mean ti/2 is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received a single oral dose of Compound 1.
  • a mean ti/2 is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • QD once daily
  • BID twice daily
  • a particular effect is or comprises a mean ti/2 of from about 2 hours to about 30 hours, from about 3 hours to about 30 hours, from about 10 hours to about 30 hours, from about 2 hours to about 11 hours, or from about 3 hours to about 11 hours.
  • a particular effect is or comprises a mean ti/2 of from about 12 hours to about 21 hours.
  • a particular effect is or comprises a mean ti/2 of less than about 30 hours, less than about 11 hours, less than about 4 hours, or less than about 3 hours.
  • a particular effect is or comprises a mean ti/2 of less than about 22 hours or less than about 15 hours.
  • a particular effect is or comprises a mean ti/2 of greater than about 2 hours, greater than about 3 hours, or greater than about 10 hours.
  • a particular effect is or comprises a particular mean accumulation ratio of Cmax (RaccCmax).
  • a mean RaccCmax is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • a particular effect is or comprises a mean RaccCmax of from about 0.6 to about 2.7, from about 0.6 to about 1.6, or from about 1.0 to about 1.6.
  • a particular effect is or comprises a mean RaccCmax of less than about 2.0, less than about 1.6, or less than about 1.4. In some embodiments, a particular effect is or comprises a mean RaccCmax of greater than about 0.5, greater than about 1.0, or greater than about 1.2.
  • a particular effect is or comprises a particular mean accumulation ratio of AUC (RaccAUC).
  • a mean RaccAUC is measured in a population of healthy subjects (e.g., healthy subjects in a fed or fasted state) who received multiple oral doses of Compound 1 (e.g., once daily (QD) or twice daily (BID) for, e.g., 14 days).
  • a particular effect is or comprises a mean RaccAUC of from about 0.8 to about 1.6 or from about 0.8 to about 1.2.
  • a particular effect is or comprises a mean RaccAUC of less than about 2.0, less than about 1.2, or less than about 1.0.
  • a particular effect is or comprises a mean RaccAUC of greater than about 0.5, greater than about 0.8, or greater than about 1.0.
  • the present disclosure provides methods of administering Compound 1 to a subject or population of subjects, according to a regimen established to achieve one or more particular effects in a population of subjects in one fed/fasted state, relative to a comparable population of subjects in another fed/fasted state.
  • a regimen is established to achieve one or more particular effects in a population of healthy subjects in one fed/fasted state, relative to a comparable population of healthy subjects in another fed/fasted state.
  • a regimen is established to achieve one or more particular effects in a population of healthy subjects in a fasted state, relative to a comparable population of healthy subjects in a fed state.
  • a regimen is established to achieve one or more particular effects in a population of healthy subjects in a fed state, relative to a comparable population of healthy subjects in a fasted state.
  • a reference population e.g., in a fasted or fed state
  • a composition providing Compound 1 e.g., the same composition providing Compound 1).
  • a population in a fasted state has received a composition providing Compound 1 after a period of fasting (e.g., an overnight fast of at least 10 hours).
  • a population in a fed state has received a composition providing Compound 1 after consumption of a meal (e.g., a high fat meal) within a period of time (e.g., within 30 minutes of receiving Compound 1).
  • the present disclosure provides methods of administering to a subject or population of subjects in need thereof an amount of Compound 1, wherein said amount, when administered as a single oral dose to a population of healthy subjects in one fed/fasted state, was established to achieve one or more particular effects relative to a comparable population of healthy subjects in another fed/fasted state.
  • the present disclosure provides methods of administering to a subject or population of subjects in need thereof an amount of Compound 1, wherein said amount, when administered as a single oral dose to a population of healthy subjects in a fasted state, was established to achieve one or more particular effects relative to a comparable population of healthy subjects in a fed state.
  • a particular effect is or comprises a difference in mean Cmax in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises an increased mean Cmax in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a mean Cmax in a population of subjects in a fasted state that is about 3 fold, about 3.5 fold, or about 4 fold greater than a mean Cmax in a population of subjects in a fed state.
  • a particular effect is or comprises a comparable mean Cmax in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state. In some embodiments, a particular effect is or comprises a mean Cmax in a population of subjects in a fasted state that is within about 50%, about 40%, or about 25% of a mean Cmax in a population of subjects in a fed state.
  • a particular effect is or comprises a difference in median Tmax in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a decreased median Tmax in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a median Tmax in a population of subjects in a fasted state that is about 2 fold, about 2.5 fold, or about 3 fold less than a median Tmax in a population of subjects in a fed state.
  • a particular effect is or comprises a lack of meaningful difference in mean AUCo-iast in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a comparable mean AUCo-iast in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a mean AUCo-iast in a population of subjects in a fasted state that is within about 10%, about 20%, or about 30% of a mean AUCo- iast in a population of subjects in a fed state.
  • a particular effect is or comprises a lack of meaningful difference in mean AUCo-inf in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a comparable mean AUCo-inf in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state. In some embodiments, a particular effect is or comprises a mean AUCo-inf in a population of subjects in a fasted state that is within about 10%, about 20%, or about 30% of a mean AUCo- inf in a population of subjects in a fed state.
  • a particular effect is or comprises a difference in mean Vz/F in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a decreased mean Vz/F in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a mean Vz/F in a population of subjects in a fasted state that is about 3 fold, about 3.5 fold, or about 4 fold less than a mean Vz/F in a population of subjects in a fed state.
  • a particular effect is or comprises a lack of meaningful difference in mean CL/F in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a comparable mean CL/F in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a mean CL/F in a population of subjects in a fasted state that is within about 10%, about 20%, or about 30% of a mean CL/F in a population of subjects in a fed state.
  • a particular effect is or comprises a difference in mean tm in a population of subjects (e.g., healthy subjects) in one fed/fasted state relative to a comparable population of subjects (e.g., healthy subjects) in another fed/fasted state.
  • a particular effect is or comprises a decreased mean tm in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state.
  • a particular effect is or comprises a mean tm in a population of subjects in a fasted state that is about 3 fold, about 3.5 fold, or about 4 fold less than a mean tm in a population of subjects in a fed state.
  • a particular effect is or comprises a comparable mean tm in a population of subjects in a fasted state relative to a comparable population of subjects in a fed state. In some embodiments, a particular effect is or comprises a mean tm in a population of subjects in a fasted state that is within about 10%, about 20%, or about 30% of a mean tm in a population of subjects in a fed state.
  • the present disclosure provides methods of administering Compound 1 to a subject or population of subjects, according to a regimen has been established to achieve one or more particular effects in a population of subjects suffering from a disease or disorder described herein (e.g., suffering from primary proteinuric kidney disease and/or primary glomerular disease and/or persistent proteinuria).
  • a regimen has been established to achieve one or more particular effects in a population of subjects suffering from a disease or disorder described herein (e.g., suffering from primary proteinuric kidney disease and/or primary glomerular disease and/or persistent proteinuria), relative to that observed for a comparable reference population that has not received Compound 1 (e.g., that has received a reference composition which does not deliver Compound 1).
  • a reference population has received an otherwise comparable reference composition that does not provide Compound 1 (e.g., a placebo as described herein).
  • composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, e.g., at a particular time point.
  • a composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, e.g., at a particular time point, relative to an appropriate reference as described herein (e.g., as is observed in a comparable population who has not received a composition providing Compound 1).
  • the present disclosure provides methods of administering to a subject or population of subjects in need thereof an amount of Compound 1, wherein said amount, when administered to a population of subjects suffering from a disease or disorder described herein (e.g., suffering from primary proteinuric kidney disease and/or primary glomerular disease and/or persistent proteinuria), has been established to achieve one or more particular effects.
  • a disease or disorder described herein e.g., suffering from primary proteinuric kidney disease and/or primary glomerular disease and/or persistent proteinuria
  • a particular effect is or comprises a particular mean or median percentage change in 24-hour urinary protein excretion (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median percent reduction in 24-hour urinary protein excretion (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median percent reduction in 24-hour urinary protein excretion (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1. In some embodiments, a particular effect is or comprises a mean or median percent reduction in 24-hour urinary protein excretion (e.g., at Week 12) of about 20% to about 40%.
  • a particular effect is or comprises a particular mean or median percentage change in 24-hour urinary albumin (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median percent reduction in 24-hour urinary albumin (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median percent reduction in 24-hour urinary albumin (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular proportion of subjects with complete remission in proteinuria.
  • a complete remission in proteinuria is defined as a 24-hour urinary protein excretion of less than 300 mg (e.g., at Week 12).
  • a particular effect is or comprises a greater proportion of subjects with complete remission in proteinuria in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular proportion of subjects with partial remission in proteinuria.
  • a partial remission in proteinuria is defined as (e.g., at Week 12) a 24-hour urinary protein excretion reduction of less than or equal to 50% from baseline and/or a 24-hour urinary protein excretion of less than 3.5 g/day if baseline 24-hour urinary protein excretion was greater than 3.5 g.
  • a particular effect is or comprises a greater proportion of subjects with partial remission in proteinuria in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a greater proportion of subjects with complete or partial remission in proteinuria in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular proportion of subjects with greater than or equal to 50% reduction in 24-hour urinary protein excretion from baseline (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater proportion of subjects with greater than or equal to 50% reduction in 24-hour urinary protein excretion from baseline (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular proportion of subjects with greater than or equal to 50% reduction in 24-hour urinary albumin excretion from baseline (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater proportion of subjects with greater than or equal to 50% reduction in 24-hour urinary albumin excretion from baseline (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median percentage change from baseline in creatinine clearance (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median percent increase from baseline in creatinine clearance (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median percent increase from baseline in creatinine clearance (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median change from baseline in serum albumin (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median increase from baseline in serum albumin (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median increase from baseline in serum albumin (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median change from baseline in fasting triglyceride levels (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median decrease from baseline in fasting triglyceride levels (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median decrease from baseline in fasting triglyceride levels (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median change from baseline in cholesterol levels (e.g., at Week 12). In some embodiments, a particular effect is or comprises a particular mean or median decrease from baseline in cholesterol levels (e.g., at Week 12). In some embodiments, a particular effect is or comprises a greater mean or median decrease from baseline in cholesterol levels (e.g., at Week 12) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median level of one or more plasma or urine biomarkers selected from plasma soluble TNF receptor 2 (sTNFR2), kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), urinary fibronectin, high molecular weight type IV collagen (HMW collagen IV), monocyte chemotactic protein-1 (MCP-1), and matrix metalloproteinase-7 (MMP-7).
  • plasma or urine biomarkers selected from plasma soluble TNF receptor 2 (sTNFR2), kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), urinary fibronectin, high molecular weight type IV collagen (HMW collagen IV), monocyte chemotactic protein-1 (MCP-1), and matrix metalloproteinase-7 (MMP-7).
  • a particular effect is or comprises a particular mean or median change in the level(s) of one or more plasma or urine biomarkers selected from plasma soluble TNF receptor 2 (sTNFR2), kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), urinary fibronectin, high molecular weight type IV collagen (HMW collagen IV), monocyte chemotactic protein- 1 (MCP-1), and matrix metalloproteinase-7 (MMP-7) in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1 relative to a comparable population who has not received a composition providing Compound 1.
  • plasma or urine biomarkers selected from plasma soluble TNF receptor 2 (sTNFR2), kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), urinary fibronectin, high molecular weight type IV collagen (HMW collagen IV), monocyte chemotactic protein- 1 (MCP-1),
  • a particular effect is or comprises a particular mean maximum concentration (Cmax).
  • Cmax mean maximum concentration
  • a mean or median Cmax is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular median time to maximum concentration (Tmax).
  • Tmax median time to maximum concentration
  • a median Tmax is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean area under drug concentration-time curve from time zero to last measurable concentration (AUCo-iast).
  • AUCo-iast mean area under drug concentration-time curve from time zero to last measurable concentration
  • a mean AUCo-iast is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean area under drug concentration-time curve from time zero to infinity (AUCo-inf).
  • AUCo-inf mean area under drug concentration-time curve from time zero to infinity
  • a mean AUCo-inf is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean area under the drug concentration-time curve over the dosing interval (AUCtau).
  • AUCtau is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean half-life (ti/2). In some embodiments, a mean ti/2 is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1. [0103] In some embodiments, a particular effect is or comprises a particular mean or median terminal elimination rate constant (K ei ). In some embodiments, a mean or median K ei is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median trough plasma concentration at steady-state measured at the end of a dosing interval before next administration (Ctrough).
  • a mean or median Ctrough is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean apparent total clearance (CL/F) on Day 1.
  • a mean CL/F is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean apparent total clearance at steady state (CL/Fss).
  • CL/Fss is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean apparent volume of distribution (Vz/F) on Day 1.
  • a mean Vz/F is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean apparent volume of distribution at steady state (Vz/Fss).
  • a mean Vz/Fss is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • a particular effect is or comprises a particular mean or median accumulation ratio.
  • a mean or median accumulation ratio is measured in a population of subjects suffering from primary glomerular disease and/or persistent proteinuria administered a composition providing Compound 1.
  • the present disclosure provides amounts of Compound 1 suitable to achieve one or more particular effects.
  • Compound 1 is administered in a daily dose of from about 50 mg to about 600 mg, from about 100 mg to about 600 mg, from about 200 mg to about 600 mg, from about 400 mg to about 600 mg, from about 50 mg to about 500 mg, from about 100 mg to about 500 mg, from about 250 mg to about 500 mg, from about 50 mg to about 250 mg, from about 100 mg to about 1000 mg, from about 200 mg to about 1000 mg, from about 500 mg to about 1000 mg, or from about 200 mg to about 500 mg.
  • Compound 1 is administered in a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a daily dose of about 50 mg, about 100 mg, about 200 mg, about 400 mg, about 500 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a twice daily dose of about 50 mg, about 100 mg, about 250 mg, or about 500 mg.
  • Compound 1 is administered in a dose of about 100 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 200 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 300 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 400 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 500 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 600 mg once daily.
  • Compound 1 is administered in a dose of about 50 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 100 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 150 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 200 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 250 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 300 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 350 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 400 mg twice daily.
  • provided methods further comprise reducing a dose of Compound 1 if a subject experiences a side effect (e.g., a gastrointestinal or liver side effect) and/or if a subject’s side effect persists.
  • one or more doses of Compound 1 are reduced by 100 mg if a subject experiences a side effect (e.g., a gastrointestinal or liver side effect).
  • a Compound 1 dosing regimen of 200 mg QD is reduced to 100 mg QD.
  • a Compound 1 dosing regimen of 400 mg QD is reduced to 300 mg QD.
  • a Compound 1 dosing regimen of 300 mg BID is reduced to 200 mg BID.
  • a Compound 1 dosing regimen of 300 mg BID is reduced to a once daily dose of 200 mg and a once daily dose of 300 mg (e.g., said doses separated by approximately 12 hours).
  • provided methods further comprise discontinuing administration of Compound 1 if a subject experiences a side effect and/or if a subject’s side effect persists after a dose reduction.
  • gastrointestinal side effects include diarrhea, nausea, vomiting, and/or abdominal cramps.
  • provided methods further comprise reducing the dose of Compound 1 if a subject experiences mild diarrhea (e.g., 4-6 extra stools per day) for 8 days or longer.
  • mild diarrhea e.g., 4-6 extra stools per day
  • anti motility therapy e.g., loperamide
  • provided methods further comprise discontinuing administration of Compound 1 if a subject experiences mild diarrhea (e.g., 4-6 extra stools per day) for an additional 7 days or longer after dose reduction.
  • provided methods further comprise reducing the dose of Compound 1 if a subject experiences severe diarrhea (e.g., more than 6 extra stools per day with clinical evidence of volume depletion or impact on activities of daily living). In some embodiments, such a subject is also treated with anti-motility therapy (e.g., loperamide). In some embodiments, provided methods further comprise discontinuing administration of Compound 1 if a subject experiences severe diarrhea (e.g., more than 6 extra stools per day with clinical evidence of volume depletion or impact on activities of daily living) for 14 days or longer.
  • severe diarrhea e.g., more than 6 extra stools per day with clinical evidence of volume depletion or impact on activities of daily living
  • provided methods further comprise reducing the dose of Compound 1 is a subject experiences nausea and/or vomiting for 8 days or longer. In some embodiments, such subjects are also treated with ondansetron. In some embodiments, provided methods further comprise discontinuing administration of Compound 1 if a subject experiences nausea and/or vomiting for 14 days or longer.
  • liver side effects include elevated liver function tests (e.g., AST or ALT increase to > 3x upper limit of normal or AST or ALT increase to > 2x baseline value if elevated at baseline).
  • provided methods comprise discontinuing administration of Compound 1 if a subject has elevated liver function tests for 2 or more days.
  • the present disclosure provides methods of treating diseases, disorders, and conditions (e.g., according to methods provided herein).
  • provided methods are useful for reducing fibrosis in a subject in need thereof.
  • provided methods are useful for treating a disease, disorder, or condition characterized by or otherwise associated with fibrosis.
  • the present disclosure encompasses the recognition that treating fibrosis (e.g., using provided methods) instead of the underlying etiology may allow for broadly applicable antifibrotic therapies. It will be appreciated that provided methods may be suitable for reducing fibrosis in a variety of tissues and/or organs; the present disclosure contemplates use of Compound 1 for treating diseases, disorders, and conditions characterized by or otherwise associated with fibrosis in any suitable tissue and/or organ. For example, in some embodiments, provided methods are suitable for treating diseases, disorders and conditions that are or comprise fibrosis of gastrointestinal tract, heart, kidney, lung, liver, muscle, pancreas, and/or skin.
  • provided methods are suitable for treating diseases, disorders, and conditions characterized by or otherwise associated with cysts (e.g., in the kidney, liver, pancreas, ovaries, spermatic duct, etc.). It will be appreciated that provided methods may be suitable for treating diseases, disorders, and conditions in which fibrosis is the sole or a predominant component, as well as those in which fibrosis is a secondary component (e.g., a symptom and/or result of an underlying disease, disorder, or condition). It will also be appreciated that there are a variety of sources or causes of fibrosis.
  • certain injuries can progress to development of fibrosis.
  • provided methods are useful for treating acute injuries (e.g., acute organ injuries, such as acute lung injury, acute liver injury, or acute kidney injury), as well as for treating chronic injuries (e.g., chronic organ injuries, such as chronic lung injury, chronic liver injury, or chronic kidney injury).
  • provided methods are useful for treating fibrosis associated with an acute injury, such as that incurred from trauma and/or surgery and/or infection (e.g., a viral infection).
  • provided methods are useful for acceleration of wound healing, reduction of post-surgical scarring, and/or reduction of adhesion formation.
  • provided methods are useful for treating damaged and/or ischemic organs, transplants, or grafts, as well as ischemia/reperfusion injury or post- surgical scarring.
  • provided methods are useful for promoting vascularization of a damaged and/or ischemic organ, transplant, or graft, ameliorating ischemia/reperfusion injury (e.g., in brain, heart, liver, or kidney), normalizing myocardial perfusion resulting from chronic cardiac ischemia or myocardial infarction, and/or developing or augmenting collateral vessel development after vascular occlusion or to ischemic tissues or organs.
  • provided methods are useful for treating pulmonary diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating pulmonary fibrosis. In some embodiments, provided methods are useful for treating pulmonary fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating interstitial lung diseases (e.g., fibrosing interstitial lung diseases). In some embodiments, provided methods are useful for treating pneumonias (e.g., idiopathic interstitial pneumonias). In some embodiments, provided methods are useful for treating idiopathic pulmonary fibrosis (IPF).
  • IPF idiopathic pulmonary fibrosis
  • provided methods are useful for treating pulmonary fibrosis associated with an infection (e.g., a bacterial, viral, or fungal infection).
  • an infection e.g., a bacterial, viral, or fungal infection.
  • provided methods are useful for treating pulmonary fibrosis associated with a viral infection (e.g., an influenza or coronavirus infection, such as COVID-19).
  • a fibrotic disease to be treated by methods of the present disclosure is pulmonary fibrosis.
  • Pulmonary fibrosis is a chronic, progressive, and ultimately fatal interstitial lung disease resulting from epithelial cell injury due to many factors.
  • activation of inflammatory cells and fibroblasts/myofibroblasts involves a cascade of cytokines/chemokines, growth factor network and deposit extracellular matrix, including collagen), which leads to pulmonary fibrosis and respiratory failure.
  • Pulmonary fibrosis causes high morbidity and mortality. At least five million people worldwide and -200,000 people in the United States suffer from pulmonary fibrosis. There is an unmet critical need for effective and affordable treatments for acute and chronic lung injuries.
  • pulmonary fibrosis Numerous endogenous and exogenous factors can provide primary stimuli for pulmonary fibrosis. Dust, silica, smoke, aerosolized toxins, infections and certain medicines have the potential to injure the lung and set the stage for the development of chronic pulmonary fibrosis. For example, viral infections may cause lung damage and/or otherwise develop into pulmonary fibrosis.
  • ARDS acute respiratory distress syndrome
  • Pulmonary fibrosis is associated with pronounced morbidity with high impact on economic burden.
  • the prevalence of pneumoconiosis (a disease caused by inhalation of dust and silica that causes inflammation and lung fibrosis) caused direct and indirect economic losses of around 28 billion yuan in China (4.3 billion US dollars) for 1 year.
  • a fibrotic disease to be treated by methods of the present disclosure is idiopathic pulmonary fibrosis.
  • Idiopathic pulmonary fibrosis is a chronic, irreversible, and progressive fibrotic disorder of the lower respiratory tract that typically affects adults over the age of 40.
  • Idiopathic interstitial pneumonias are diffuse parenchymal lung diseases, of which IPF is the most common and severe type of fibrotic lung disease. It is anatomically characterized by scarring of the lungs with a pattern of usual interstitial pneumonia (UIP) on high resolution computed tomography or histologic appearance on lung biopsy.
  • IPF Intra-pulmonary disease characterized by exertional dyspnea and cough.
  • Median survival following diagnosis of IPF ranges between 2 and 5 years, lower than that for many common cancers (Ley, B., et al. Am. J. Respir. Crit. Care Med. 2011;183:431-440; Seigel, R. L., et al. CA. Cancer J. Clin. 2016;66:7- 30).
  • Fibrotic process in IPF is progressive and, regardless of the nature of the initial injury, may follow a common pathway characterized by alveolar epithelial cell (AEC) dysfunction.
  • AEC alveolar epithelial cell
  • AECsl type I epithelial cells
  • AECs2 type II epithelial cells
  • TGF-bI transforming growth factor beta 1
  • PDGF platelet-derived growth factor
  • VEGF vascular endothelial growth factor
  • FGF fibroblast growth factor
  • the progression of fibrosis in IPF follows a common and complex path in which the AECs, fibroblasts, and endothelial cells produce an array of cytokines and growth factors that stimulate fibroblast proliferation and matrix synthesis.
  • provided methods are useful for treating hepatic diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating hepatic fibrosis (e.g., fibrotic liver disease). In some embodiments, provided methods are useful for treating cirrhosis. In some embodiments, provided methods are useful for treating hepatic fibrosis and/or cirrhosis secondary to, or otherwise associated with, an underlying indication.
  • hepatic fibrosis e.g., fibrotic liver disease
  • provided methods are useful for treating cirrhosis.
  • provided methods are useful for treating hepatic fibrosis and/or cirrhosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating hepatic fibrosis associated with hepatitis C, hepatitis B, delta hepatitis, chronic alcoholism, nonalcoholic steatohepatitis (NASH), extrahepatic obstructions (e.g., stones in bile duct), cholangiopathies (e.g., primary biliary cirrhosis or sclerosing cholangitis), autoimmune liver disease, or inherited metabolic disorders (e.g., Wilson’s disease, hemochromatosis, or alpha-1 antitrypsin deficiency).
  • NASH nonalcoholic steatohepatitis
  • a fibrotic disease to be treated by methods of the present disclosure is liver fibrosis.
  • Liver fibrosis is a scarring response of the liver to chronic liver injury; when fibrosis progresses to cirrhosis, morbid complications can develop.
  • end- stage liver fibrosis or cirrhosis is the seventh leading cause of death in the United States, and afflicts hundreds of millions of people worldwide; deaths from end-stage liver disease in the United States are expected to increase, mainly due to the hepatitis C epidemic.
  • liver disease In addition to the hepatitis C virus, many other forms of chronic liver injury also lead to end-stage liver disease and cirrhosis, including other viruses such as hepatitis B and delta hepatitis, chronic alcoholism, non-alcoholic steatohepatitis, extrahepatic obstructions (e.g., stones in the bile duct), cholangiopathies (e.g., primary biliary cirrhosis and sclerosing cholangitis), autoimmune liver disease, and inherited metabolic disorders (e.g., Wilson's disease, hemochromatosis, and alpha- 1 anti trypsin deficiency).
  • viruses such as hepatitis B and delta hepatitis, chronic alcoholism, non-alcoholic steatohepatitis, extrahepatic obstructions (e.g., stones in the bile duct), cholangiopathies (e.g., primary biliary cirrhosis and sclerosing
  • liver fibrosis has traditionally focused on eliminating a primary injury. For extrahepatic obstructions, biliary decompression is the recommended mode of treatment whereas patients with Wilson's disease are treated with zinc acetate. Treatments for other chronic liver diseases such as hepatitis B, autoimmune hepatitis and Wilson's disease are also associated with many side effects, while primary biliary cirrhosis, primary sclerosing cholangitis and non alcoholic fatty liver disease have no effective treatment other than liver transplantation.
  • provided methods are useful for treating renal diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating renal fibrosis. In some embodiments, provided methods are useful for treating renal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating renal fibrosis associated with renal failure, renal obstruction, renal trauma, renal transplantation, chronic kidney disease, diabetes, hypertension, radiocontrast nephropathy, immune-mediated glomerulonephri tides (e.g., lupus nephritis, ANCA-associated glomerulonephritides (e.g., Wegener’s granulomatosis, microscopic polyangiitis, or renal limited vasculitis), anti-GBM nephropathy, IgA nephropathy, membranous glomerulonephritis, or focal and segmental glomerulosclerosis), non-immune-mediated glomerulonephritides (e.g., polycystic kidney disease, collagen type III glomerulopathy, nail-patella syndrome, or Alport syndrome), minimal change disease, or nephrotic syndrome (e.g., steroid-resistant nephrotic syndrome).
  • provided methods are useful for treating nephrotic syndrome and/or diseases, disorders, or conditions associated with nephrotic syndrome (e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy).
  • nephrotic syndrome e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy.
  • provided methods are useful for treating a fibrotic disease of the kidney that is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, Alport syndrome, anti-globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), collagen type III glomerulopathy, nail- patella syndrome, or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria lupus nephritis
  • minimal change disease an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis
  • ANCA
  • provided methods are useful for treating a fibrotic disease of the kidney that is or comprises an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis.
  • ANCA-associated glomerulonephritis is selected from Wegener’s granulomatosis, microscopic polyangiitis (MPA), or renal limited vasculitis.
  • provided methods are useful for treating focal and segmental glomerulosclerosis (FSGS).
  • FSGS focal and segmental glomerulosclerosis
  • provided methods are useful for treating Alport syndrome.
  • provided methods are useful for treating polycystic kidney disease (e.g., autosomal dominant polycystic kidney disease or autosomal recessive polycystic kidney disease).
  • provided methods are useful for treating primary proteinuric kidney disease (PPKD).
  • PPKD primary proteinuric kidney disease
  • PGDs primary glomerular diseases
  • PGDs are among the leading causes of chronic kidney disease and end-stage kidney disease in the world. PGDs predominantly affect younger patients, significantly reducing their quality of life, productivity, and longevity.
  • FSGS, membranous nephropathy (MN), and IgA nephropathy are among the three most common primary glomerular diseases in adults. Accordingly, in some embodiments, provided methods are useful for treating FSGS. In some embodiments, provided methods are useful for treating MN. In some embodiments, provided methods are useful for treating IgA nephropathy.
  • CNIs calcineurin inhibitors
  • Some of the drugs have a narrow therapeutic index necessitating close monitoring of the drug levels.
  • Long-term use of CNIs is associated with hypertension, nephrotoxicity, and metabolic abnormalities such as diabetes and dyslipidemia.
  • cessation of calcineurin inhibitors results in the relapse of proteinuria (Meyrier, A. et al., Kidney International. 1994;45(5): 1446-56).
  • a significant number of patients eventually become either resistant to or dependent on these toxic agents.
  • Some of these glomerular diseases also recur after renal transplantation posing unique management problems (Choy, B.Y., et al., Am. J. Transplant. 2006;6(11):2535-42).
  • provided methods are useful for treating patients with proteinuria (e.g., persistent proteinuria). It is well established that higher rates of urinary protein excretion are associated with worse prognosis, and therapies that reduce proteinuria are desirable for improving renal outcomes. Patients with persistent proteinuria (e.g., who continue to have >
  • ESKD end-stage kidney disease
  • eGFR estimated glomerular filtration rate
  • Patients with persistent proteinuria also develop further complications of chronic kidney disease (CKD) such as dyslipidemia, cardiovascular disease, abnormalities in mineral-bone metabolism, and hypertension, resulting in significant increases in morbidity and mortality and utilization of health care resources.
  • CKD chronic kidney disease
  • RAAS renin-angiotensin-aldosterone system
  • ARB angiotensin-receptor blockers
  • RAAS blockers reduce proteinuria and improve clinical outcomes in proteinuric renal diseases regardless of the etiology.
  • Other standard of care recommendations include aggressive blood pressure control ( ⁇ 130/80), and HMG-CoA reductase inhibitors (e.g., statins) in patients with hyperlipidemia.
  • statins HMG-CoA reductase inhibitors
  • the inhibitors of the mineralocorticoid receptor and sodium glucose co-transporter-2 (SGLT-2) are increasingly being used in these patients as well.
  • provided methods are useful for treating primary glomerular diseases (e.g., FSGS, membranous nephropathy, or IgA nephropathy) and persistent proteinuria.
  • primary glomerular diseases e.g., FSGS, membranous nephropathy, or IgA nephropathy
  • persistent proteinuria e.g., FSGS, membranous nephropathy, or IgA nephropathy
  • PDGFRP Platelet- derived growth factor receptor beta
  • a kidney disease to be treated by methods of the present disclosure is nephrotic syndrome (NS).
  • NS is a group of rare renal diseases, including focal and segmental glomerulosclerosis (FSGS), minimal change disease (MCD), and membranous nephropathy.
  • FSGS focal and segmental glomerulosclerosis
  • MCD minimal change disease
  • membranous nephropathy is a rare disease that attacks the kidney’s filtering units (glomeruli) causing serious scarring which leads to permanent kidney damage and even failure (Fogo, A.B. Nat. Rev. Nephrol. 2015 Feb;l l(2):76-87, PMCID:PMC4772430). It will be appreciated that there are at least three types of FSGS.
  • Primary FSGS is FSGS that has no known cause (also referred to as idiopathic FSGS). Secondary FSGS is caused by one or more factors such as infection, drug toxicity, diseases such as diabetes or sickle cell disease, obesity, or other kidney diseases. Genetic FSGS (also called familial FSGS) is caused by one or more genetic mutations. Primary FSGS is idiopathic in nature. Manifestations of this disease include hypoalbuminemia and edema, lipid abnormalities and nephrotic range proteinuria. More than 5400 patients are diagnosed with FSGS every year (O’Shaughnessy, M.M., et al. Nephrol. Dial. Transplant 2018 Apr l;33(4):661-9).
  • a kidney disease to be treated by methods of the present disclosure is minimal change disease (MCD).
  • MCD is a kidney disease in which large amounts of protein are lost in the urine. It is one of the most common causes of the nephrotic syndrome worldwide. In children, MCD is usually primary (or idiopathic), but in adults, the disease is usually secondary. Secondary causes for MCD include allergic reactions, use of certain painkillers such as non-steroidal anti-inflammatory drugs (NSAIDs), tumors, or viral infections.
  • a kidney disease to be treated by methods of the present disclosure is membranous glomerulonephritis (MG or MGN), also known as membranous nephropathy (MN).
  • MG or MGN is a slowly progressive renal disease caused by immune complex formation in the glomerulus.
  • Immune complexes are formed by binding of antibodies to antigens in the glomerular basement membrane.
  • the antigens may be part of the basement membrane, or deposited from elsewhere by the systemic circulation.
  • a kidney disease to be treated by methods of the present disclosure is anti -neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis.
  • Anti neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis is a rapidly progressive renal disease and includes, e.g., Wegener's granulomatosis, microscopic polyangiitis, and renal limited vasculitis.
  • Wegener's granulomatosis is an organ- and/or life-threatening autoimmune disease of unknown etiology.
  • the classical clinical triad consists of necrotizing granulomatous inflammation of the upper and/or lower respiratory tract, necrotizing glomerulonephritis, and an autoimmune necrotizing systemic vasculitis affecting predominantly small vessels.
  • the detection of anti-neutrophil cytoplasmic antibodies directed against proteinase 3 (PR3-ANCA) is a highly specific indicator for Wegener's granulomatosis.
  • Microscopic polyangiitis is a disorder that causes blood vessel inflammation (vasculitis), which can lead to organ damage.
  • the kidneys, lungs, nerves, skin, and joints are the most commonly affected areas of the body.
  • MPA is diagnosed in people of all ages, all ethnicities, and both genders. The cause of this disorder is unknown.
  • Renal limited vasculitis is a type of anti-neutrophil cytoplasmic antibody (ANCA)- associated vasculitis that presents with only a renal manifestation; no other organs, including lungs, are involved.
  • ANCA anti-neutrophil cytoplasmic antibody
  • a kidney disease to be treated by methods of the present disclosure is lupus nephritis.
  • Lupus nephritis is inflammation of the kidney that is caused by the autoimmune disease systemic lupus erythematous (SLE). With lupus, the body's immune system targets its own body tissues; lupus nephritis occurs when lupus involves the kidneys.
  • a kidney disease to be treated by methods of the present disclosure is anti-globular basement membrane (anti-GBM) nephropathy.
  • Anti-globular basement membrane (anti-GBM) nephropathy is a disease that occurs as a result of injury to small blood vessels (capillaries) in the kidneys and/or lungs.
  • anti-GBM disease autoantibodies are targeted to the basement membrane in capillary blood vessels of the kidneys and lung, where they target and damage GBM.
  • a kidney disease to be treated by methods of the present disclosure is IgA nephropathy, also known as Berger’s disease.
  • IgA nephropathy is a kidney disease that occurs when IgA deposits build up in the kidneys, causing inflammation that damages kidney tissues.
  • IgA nephropathy affects the kidneys by attacking the glomeruli. The buildup of IgA deposits inflames and damages the glomeruli, causing the kidneys to leak blood and protein into the urine. The damage may lead to scarring of the nephrons that progresses slowly over many years. Eventually, IgA nephropathy can lead to end-stage kidney disease.
  • a kidney disease to be treated by methods of the present disclosure is collagen type III glomerulopathy.
  • Collagen type III glomerulopathy also known as collagenic or collagenofibrotic glomerulopathy, is characterized by pathological accumulation of collagen type III in glomeruli.
  • Collagen type III glomerulopathy presents either in childhood, often with a family history suggesting autosomal recessive inheritance, or in adults as a sporadic occurrence.
  • Proteinuria is a typical manifestation, with progression to end stage renal disease (ESRD) in approximately 10 years.
  • ESRD end stage renal disease
  • a kidney disease to be treated by methods of the present disclosure is nail-patella syndrome.
  • Nail-patella syndrome is a multi-organ disorder caused by mutations in the LMX1B gene. Nail-patella syndrome manifests with orthopedic and cutaneous deformities, as well as kidney complications due to development of structural lesions of collagen type III within glomerular basement membranes. Although the structural lesions may be asymptomatic, they are usually accompanied by proteinuria.
  • a kidney disease to be treated by methods of the present disclosure is Alport syndrome (AS).
  • AS is a genetic condition characterized by kidney disease, hearing loss, and eye abnormalities. Most affected individuals experience progressive loss of kidney function, usually resulting in end-stage kidney disease.
  • Alport syndrome is inherited in an X-linked manner and is caused by mutation(s) in the COL4A5 gene. In other cases, it can be inherited in either an autosomal recessive, or rarely in an autosomal dominant manner, and is caused by mutation(s) in the COL4A3 and/or COL4A4 genes.
  • Current therapies include hearing aid, hemodialysis, peritoneal dialysis and kidney transplantation.
  • a kidney disease to be treated by methods of the present disclosure is polycystic kidney disease (e.g., autosomal recessive polycystic kidney disease (ARPKD) - congenital hepatic fibrosis (CHF)).
  • ARPKD-CHF is a highly aggressive fibropolycystic disease that is characterized by the formation and expansion of fluid-filled cysts in the kidneys, enlargement of the kidneys and progressive fibrosis of both the kidney and the liver (Hartung, E.A., and Guay -Woodford, L.M. Pediatrics 2014 Sep;134(3):e833-e845; Gunay- Aygun, M., et al. J.
  • ARPKD-CHF congenital hepatic fibrosis
  • a kidney disease to be treated by methods of the present disclosure is or includes renal cysts.
  • Aberrant signaling by tyrosine kinases including platelet- derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) and their receptors (R), PDGFR and VEGFR/KDR, respectively, has been implicated in the formation and expansion of renal cysts.
  • PDGF platelet- derived growth factor
  • VEGF vascular endothelial growth factor
  • R vascular endothelial growth factor
  • a PDGF-driven ciliopathy and/or overexpression of PDGF in the cyst lining and adjacent tubules are thought to, in part, drive renal cystic disease (Torres, V.E., et al. Lancet 2007 Apr 14;369(9569): 1287-301; Park. J.H. et al.
  • VEGF-driven angiogenesis is also thought to contribute to the growth of renal cysts, and inhibition of VEGFR/KDR signaling is associated with decreased tubule cell proliferation, decreased cystogenesis, and blunted renal enlargement (Bello-Ruess, E., et al. Kidney Int. 2001 Jul;60(l):37-45; Schrijvers, B.F., et al. Kidney Int. 2004 Jun;65(6):2003-17). Nevertheless, the role of VEGF in fibropoly cystic disease is more controversial, with at least some reports suggesting that this growth factor might be associated with disease mitigation (Spirli, C., et al. Gastroenterology 2010 Jan;138(l):360-71).
  • provided methods are useful for treating dermal diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating dermal fibrosis. In some embodiments, provided methods are useful for treating dermal fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating scleroderma and/or systemic sclerosis (e.g., diffuse systemic sclerosis or limited systemic sclerosis).
  • systemic sclerosis e.g., diffuse systemic sclerosis or limited systemic sclerosis.
  • a fibrotic disease to be treated by methods of the present disclosure is scleroderma and/or systemic sclerosis (SSc).
  • Scleroderma which literally means hard skin, is a chronic fibrotic disorder of unknown etiology that affects the skin and other internal organs (SSc) (www.scleroderma.org).
  • SSc systemic sclerosis
  • Many patients who suffer from scleroderma/SSc also have loss of lung function.
  • Scleroderma/SSc and related diseases afflict approximately 400,000 to 990,000 people in the USA every year. Mortality and morbidity in scleroderma/SSc are very high and directly related to the extent of fibrosis of the involved organs (Hinchcliff, M.
  • Scleroderma/SSc can be classified in terms of the degree and location of the skin involvement and has been categorized into two major groups - diffuse and limited.
  • the diffuse form of scleroderma/SSc involves symmetric thickening of skin of the extremities, face and trunk. Organs affected include the esophagus, intestines, lungs, heart, and kidneys (Mayes, M. D. Semin. Cutan. Med. Surg. 1998 Mar;17(l):22-6; Jacobsen, L. et al. J. Am. Acad. Dermatol. 2003 Aug;49(2):323-5).
  • the limited form of scleroderma/SSc tends to be confined to the skin of fingers and face.
  • scleroderma/SSc The limited form of scleroderma/SSc is the CREST variant of scleroderma/SSc based on the clinical pattern of calcinosis with tiny deposits of calcium in the skin, Raynaud's phenomenon in the fingers, toes, nose, tongue, or ears, poor functioning of muscle of esophagus, sclerodactyly of the skin of the fingers or toes, and telangiectasias on the face, hands and mouth (Winterbauer, R.H. Bull. Johns Hopkins Hospital 1964;114:361-83; Wollheim, F.A. Classification of systemic sclerosis. Visions and reality. Rheumatology (Oxford) 2005).
  • fibrotic pathways are activated in scleroderma/SSc for reasons that are not completely understood.
  • the pathogenesis of fibrosis in scleroderma/SSc involves a complex set of interactions involving immune activation, microvascular damage and the activation of fibroblasts.
  • Scleroderma/SSc is characterized by excessive deposition of collagen in the skin and other involved organs and abnormalities of blood vessels (Jimenez, S. A., et al. Rheum. Dis.
  • TGFpi a multifunctional cytokine
  • PDGF downregulating growth factor
  • TGFp and PDGF are the most potent proteins involved in fibroblast proliferation, collagen gene expression and connective tissue (collagen) accumulation (Antoniades, H.N. Baillieres Clin. Endocrinol. Metab. 1991 Dec;5(4):595-613). Numerous other cytokines including VEGF, as well as cell-matrix interactions, also modify collagen expression and can influence the effects of TGFpi and PDGF (Trojanowska, M. Rheumatology (Oxford) 2008 Oct;47 Suppl 5:v2-4).
  • Persistent overproduction of collagen and other connective tissue results in excessive accumulation of ECM components leading to the formation of scar tissue (fibrosis) in the skin and other organs and is responsible for the progressive nature of scleroderma/SSc (Mauch, C. Rheum. Dis. Clin. North Am. 1990 Feb;16(l):93-107). This leads to thickness and firmness of involved areas.
  • the pathogenic cascade at different stages of scleroderma/SSc may have autoimmune, inflammatory, fibrotic and vascular components with systemic fibrosis and vasculopathy.
  • provided methods are useful for treating gastrointestinal diseases, disorders, or conditions.
  • provided methods are useful for treating gastrointestinal fibrosis (e.g., fibrosis of esophagus, stomach, intestines, and/or colon).
  • provided methods are useful for treating gastrointestinal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating inflammatory bowel disease (e.g., ulcerative colitis or Crohn’s disease), e.g., treating gastrointestinal fibrosis associated with inflammatory bowel disease.
  • IBD is primarily an inflammatory autoimmune disease
  • anti-fibrotic agents e.g., Compound 1 could be useful in treating IBD and, further could provide benefits beyond mere anti-fibrotic effects.
  • a disease to be treated by methods of the present disclosure is inflammatory bowel disease (IBD).
  • IBD is an inflammatory condition that comprises both ulcerative colitis (UC) and Crohn's disease (CD). While UC affects the entire colon, CD typically affects the ileum but can occur in any part of the GI tract.
  • IBD can manifest as acute or chronic colitis, characterized by recurrent intestinal inflammation accompanied by diarrhea and abdominal pain (Arivarasu, N., et al. Tissue Barriers 2018;6(2):el463897; Ponder, A. and Long, M.D. Clin. Epidemiol. 2013;5:237-47).
  • IBD Incidence of IBD is increasing worldwide and is an expanding global health problem (Amosy, E., et al. Clin. Med. Insights Gastroenterol. 2013;6:33-47). An estimated 2.5-3 million people in Europe are affected by IBD (Burisch, J., et al. J. Crohns Colitis 2013 May;7(4):322- 37). According to the Centers for Disease Control and Prevention (CDC), 3.1 million adults in this country were diagnosed with IBD in 2015, a substantial increase from the - 1.4 million adults diagnosed per 2008 reports (www.cdc.gov/IBD; www.cdc.gov/ibd/pdf/inflammatory- bowel-disease-an-expensive-disease.pdf).
  • IBD accounts for -1,300,000 physician visits and -92,000 hospitalizations each year in the United States. Of these, 75% patients diagnosed with CD and 25% patients diagnosed with UC and require surgery. Risk factors associated with IBD include environmental, genetic and immunologic factors (Abegunde, A.T., et al. World J. Gastroenterol. 2016 Jul 21;22(27):6296-6317; Frolkis, A., et al. Can. J. Gastroenterol. 2013 Mar; 27(3 ) : e28 -24) .
  • IBD ulcerative colitis
  • IBD is an autoimmune disease with excessive activation of the adaptive immune response.
  • Various factors including genetic factors alter the intestinal flora and trigger an inflammatory reaction, activate T cells, B cells, mast cells, macrophages and microglia, smooth muscle cells and fibroblasts in the colon, inducing mucosal disruption (Hildner, K., et al. Dig. Dis. 2016;34Suppl 1:40-7; Curciarello, R., et al. Front Med. (Lausanne) 2017 Aug 7;4:126).
  • Epithelial and endothelial damage release chemotactic factors promoting recruitment and activation of inflammatory cells, and release various cytokines including TNFa, and activate fibroblasts via TGFpi.
  • Activated fibroblasts i.e. myofibroblasts, secrete growth factors including platelet derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) (Scaldaferri, et al. Gastroenterology 2009 Feb;136(2):585-95.e5).
  • PDGF platelet derived growth factor
  • VEGF vascular endothelial growth factor
  • angiogenesis is also an important part of IBD pathogenesis in the colon of IBD patients.
  • Alkim, et al. demonstrated enhanced microvessel density in the intestinal tissue of both UC and CD patients, which correlated both the level of local VEGF expression and disease activity (Int. J. Inflam. 2015;2015:970890).
  • Anti-inflammatory drugs including 5-aminosalicylic acid (5-ASA)-based preparations, are often the first line of therapy in IBD (Segars, L.W., et al. Clin. Pharm. 1992 Jun;ll(6):514-28).
  • Anti-TNFa antibodies such as infliximab and adalimumab are also being used. Nevertheless, patients treated with adalimumab are at increased risk for serious infections and lymphoma (Dulai, P.S., et al. Clin. Gastroenterol. Hepatol. 2014 Sep; 12(9): 1443-51).
  • PDGF activates fibroblasts and IBD-fibroblasts proliferate more rapidly than normal fibroblasts; collagen secretion from IBD patients’ fibroblasts was increased compared to collagen secretion by normal fibroblasts.
  • IBD is also associated with increased circulating PDGF and the level of this growth factor has been reported to correspond with disease severity (Andrae, J., et al. Genes Dev. 2008 May 15;22(10): 1276-1312).
  • angiogenesis is a novel component of IBD pathogenesis and angiogenic activity is increased in IBD patients.
  • Serum VEGF levels were significantly higher in IBD patients compared to controls in several studies.
  • Griga et al. demonstrated that sources of increased serum VEGF were from inflamed intestinal tissue of IBD patients (Scand. J. Gastroenterol. 1998 May;33(5):504-8; Hepatogastroenterology 2002 Jan-Feb;49(43): 116-23; Hepatogastroenterology 1999 Mar-Apr;46(26):920-3; Eur. J. Gastroenterol. Hepatol. 1999 Feb; 11(2): 175-9).
  • VEGF expression was markedly increased in the inflamed mucosa of both CD and UC patients, when compared with normal mucosa of the same patient. Studies also showed that VEGF expression was increased in colon and was higher across all IBD groups (both CD and UC) when compared with healthy controls. Scaldaferri, et al. (2009) reported that VEGF receptor (VEGFR/KDR) levels were increased in intestinal samples of IBD patients, and in mice with experimental colitis.
  • VEGFR/KDR VEGF receptor
  • provided methods are useful for treating certain other diseases, disorders, or conditions.
  • provided methods are useful for treating cardiac fibrosis and/or fibrosis associated with cardiovascular system.
  • provided methods are useful for treating cardiac fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provide methods are useful for treating cardiac and/or cardiovascular fibrosis associated with ischemic heart disease, myocardial ischemia, athereosclerosis, myocardial perfusion (e.g., as a consequence of chronic cardiac ischemia or myocardial infarction), vascular occlusion, or restenosis.
  • a disease to be treated by methods of the present disclosure is ischemic heart disease.
  • Ischemic heart disease is a leading cause of morbidity and mortality in the US, afflicting millions of Americans each year at a cost expected to exceed $300 billion/year.
  • Numerous pharmacological and interventional approaches are being developed to improve treatment of ischemic heart disease including reduction of modifiable risk factors, improved revascularization procedures, and therapies to halt progression and/or induce regression of atherosclerosis.
  • atherosclerosis comprises a fibrotic component.
  • provided methods are useful for treating fibrosis associated with central nervous system (CNS) and/or one or more CNS-related diseases, disorders, or conditions.
  • CNS central nervous system
  • provided methods are useful for treating CNS-associated fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating fibrosis associated with cerebral infarction, stroke, or amyotrophic lateral sclerosis.
  • provided methods are useful for treating fibrosis associated with musculoskeletal system and/or one or more musculoskeletal diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating musculoskeletal- associated fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating fibrosis associated with muscular dystrophy.
  • provided methods are useful for treating pancreatic fibrosis.
  • provided methods are useful for treating pancreatic fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating fibrosis associated with pancreatitis.
  • one or more subjects or populations are selected to receive Compound 1 as described herein based on one or more markers and/or characteristics such as, for example, one or more risk factors of fibrosis or an associated disease, disorder or condition and/or one or more biomarkers, etc.
  • a subject or population thereof is selected to receive Compound 1 using technologies provided herein (e.g., based on assessment of one or more markers and/or characteristics, such as one or more biomarkers).
  • technologies are used to inform or determine one or more features of a therapeutic regimen (e.g., selection of subject(s) to receive a particular therapy (e.g., Compound 1 therapy) and/or dose thereof and/or timing of administration of such therapy).
  • assessment of one or more markers and/or characteristics is performed with respect to the same subject at a plurality of different time points. In some embodiments, assessment of one or more markers and/or characteristics is performed with respect to a particular patient prior to initiation of a particular therapeutic regimen (e.g., a Compound 1 therapeutic regimen) and/or prior to administration of a particular dose of therapy (e.g., Compound 1 therapy) in accordance with such therapeutic regimen.
  • a particular therapeutic regimen e.g., a Compound 1 therapeutic regimen
  • a particular dose of therapy e.g., Compound 1 therapy
  • a subject or population thereof is suffering from or susceptible to a disease, disorder, or condition described herein. In some embodiments, a subject or population thereof is suffering from or is susceptible to fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to a disease, disorder, or condition characterized by or otherwise associated with fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to fibrosis of gastrointestinal tract, heart, kidney, lung, liver, muscle, pancreas, and/or skin.
  • a subject or population thereof is suffering from or is susceptible to an acute injury (e.g., an acute organ injury, such as acute lung injury, acute liver injury, or acute kidney injury).
  • a subject or population thereof is suffering from or is susceptible to a chronic injury (e.g., a chronic organ injury, such as chronic lung injury, chronic liver injury, or chronic kidney injury).
  • a subject or population thereof is suffering from a traumatic injury.
  • a subject or population thereof has undergone, is undergoing, or will undergo an organ transplantation.
  • a subject or population thereof is suffering from or susceptible to a damaged and/or ischemic organ, transplant, or graft.
  • a subject or population thereof is suffering from or susceptible to ischemia/reperfusion injury. In some embodiments, a subject or population thereof is suffering from or susceptible to post-surgical scarring. In some embodiments, a subject or population thereof is suffering from a wound.
  • a subject or population thereof is suffering from or is susceptible to a pulmonary disease, disorder, or condition. In some embodiments, a subject or population thereof is suffering from or is susceptible to pulmonary fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to pulmonary fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to interstitial lung disease (e.g., fibrosing interstitial lung disease). In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic interstitial pneumonia. In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic pulmonary fibrosis.
  • interstitial lung disease e.g., fibrosing interstitial lung disease
  • a subject or population thereof is suffering from or is susceptible to idiopathic interstitial pneumonia. In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic
  • a subject or population thereof is suffering from or is susceptible to a hepatic disease, disorder, or condition.
  • a subject or population thereof is suffering from or is susceptible to hepatic fibrosis (e.g., fibrotic liver disease).
  • a subject or population thereof is suffering from or is susceptible to cirrhosis.
  • a subject or population thereof is suffering from or is susceptible to hepatic fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to hepatitis C, hepatitis B, delta hepatitis, chronic alcoholism, nonalcoholic steatohepatitis (NASH), extrahepatic obstructions (e.g., stones in bile duct), cholangiopathies (e.g., primary biliary cirrhosis or sclerosing cholangitis), autoimmune liver disease, or inherited metabolic disorders (e.g., Wilson’s disease, hemochromatosis, or alpha-1 antitrypsin deficiency).
  • a subject or population thereof is suffering from or is susceptible to a renal disease, disorder, or condition.
  • a subject or population thereof is suffering from or is susceptible to renal fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to renal fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to renal failure, renal obstruction, renal trauma, renal transplantation, chronic kidney disease, diabetes, hypertension, radiocontrast nephropathy, immune-mediated glomerulonephri tides (e.g., lupus nephritis, ANCA-associated glomerulonephritides (e.g., Wegener’s granulomatosis, microscopic polyangiitis, or renal limited vasculitis), anti-GBM nephropathy, IgA nephropathy, membranous glomerulonephritis, or focal and segmental glomerulosclerosis), non-immune-mediated glomerulonephritides (e.g., polyc
  • a subject or population thereof is suffering from or is susceptible to a fibrotic disease of the kidney that is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, Alport syndrome, anti-globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), collagen type III glomerulopathy, nail-patella syndrome, or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria lupus nephritis
  • minimal change disease an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomeruloneph
  • a subject or population thereof is suffering from or is susceptible to a fibrotic disease of the kidney that is or comprises an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis.
  • ANCA-associated glomerulonephritis is selected from Wegener’s granulomatosis, microscopic polyangiitis (MPA), or renal limited vasculitis.
  • a subject or population thereof is suffering from or is susceptible to collagen type III glomerulopathy or nail-patella syndrome.
  • a subject or population thereof is suffering from or is susceptible to nephrotic syndrome and/or diseases, disorders, or conditions associated with nephrotic syndrome (e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy).
  • a subject or population thereof is suffering from or is susceptible to focal and segmental glomerulosclerosis.
  • a subject or population thereof is suffering from or is susceptible to Alport syndrome.
  • a subject or population thereof is suffering from or is susceptible to polycystic kidney disease (e.g., autosomal dominant polycystic kidney disease or autosomal recessive polycystic kidney disease).
  • a subject or population thereof is suffering from or is susceptible to primary proteinuric kidney disease (e.g., as confirmed from a renal biopsy).
  • a subject or population thereof is suffering from or is susceptible to primary glomerular diseases (e.g., as confirmed from a renal biopsy).
  • a subject or population thereof is suffering from or susceptible to persistent proteinuria.
  • a subject or population thereof is suffering from or susceptible to primary glomerular disease (e.g., as confirmed from a renal biopsy) and persistent proteinuria.
  • a subject or population thereof is suffering from or susceptible to proteinuric chronic kidney disease.
  • a subject or population thereof is suffering from or is susceptible to focal and segmental glomerulosclerosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to membranous nephropathy. In some embodiments, a subject or population thereof is suffering from or susceptible to IgA nephropathy.
  • a subject or population thereof has one or more risk factors for primary glomerular disease and/or primary proteinuric kidney disease.
  • a subject or population thereof has one or more risk factors for primary glomerular disease and/or primary proteinuric kidney disease selected from proteinuria, renal dysfunction, hypertension, interstitial fibrosis on renal biopsy, and lack of response to therapy or relapse in proteinuria.
  • a subject or population thereof has one or more risk factors for primary glomerular disease (e.g., FSGS) selected from serum creatinine > 1.3 mg/dL, proteinuria > 3.5 g/day, lack of response to therapy, and/or a collapsing variant.
  • a subject or population thereof has one or more risk factors for primary glomerular disease (e.g., IgA nephropathy) selected from serum creatinine > 1.26 mg/dL, hypertension (e.g., >140/90 mmHg), and/or persistent (e.g., > 6 months) protein excretion > 1000 mg/day.
  • IgA nephropathy selected from serum creatinine > 1.26 mg/dL, hypertension (e.g., >140/90 mmHg), and/or persistent (e.g., > 6 months) protein excretion > 1000 mg/day.
  • a subject or population thereof has one or more risk factors for primary glomerular disease (e.g., membranous nephropathy) selected from serum creatinine > 1.5 mg/dL, progressive decline in eGFR > 25% over prior 2 years, and/or severe disabling or life-threatening nephrotic syndrome (with e.g., serum albumin ⁇ 2.5 g/dL, refractory edema, and/or thromboembolic event).
  • primary glomerular disease e.g., membranous nephropathy
  • serum creatinine > 1.5 mg/dL e.g., progressive decline in eGFR > 25% over prior 2 years
  • severe disabling or life-threatening nephrotic syndrome with e.g., serum albumin ⁇ 2.5 g/dL, refractory edema, and/or thromboembolic event.
  • a subject or population thereof has a risk factor for primary glomerular disease (e.g., FSGS, IgA nephropathy, and/or membranous nephropathy) of eGFR ⁇ 70 mL/min/1.73m 2 .
  • FSGS primary glomerular disease
  • IgA nephropathy IgA nephropathy
  • membranous nephropathy membranous nephropathy
  • a subject or population thereof has an estimated glomerular filtration rate (eGFR) of greater than or equal to 40 mL/min/1.73m 2 , e.g., calculated using Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI). In some embodiments, a subject or population thereof has an estimated glomerular filtration rate (eGFR) of greater than or equal to 30 mL/min/1.73m 2 , e.g., calculated using Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI).
  • CKD-EPI Chronic Kidney Disease Epidemiology Collaboration
  • a subject or population thereof has a urinary protein excretion of greater than or equal to 1 g/day on a 24-hour urine collection.
  • a subject or population thereof has stable blood pressure (e.g., less than or equal to 140/90 mmHg) for at least 12 weeks. In some embodiments, a subject or population thereof is on a stable antihypertensive regimen and has stable blood pressure (e.g., less than or equal to 140/90 mmHg) for at least 12 weeks.
  • a subject or population thereof is receiving or has received standard of care therapy. In some embodiments, a subject or population thereof is receiving or has received a maximally tolerated or recommended dose of standard of care therapy. In some embodiments, a subject or population is receiving or has received standard of care therapy and has been stable on said therapy for at least 12 weeks. In some embodiments, a subject or population thereof is resistant to standard of care therapy. In some embodiments, a subject or population thereof has relapsed after receiving standard of care therapy. In some embodiments, standard of care therapy comprises immunosuppressive or immunomodulatory agents.
  • standard of care therapy comprises renin-angiotensin-aldosterone system (RAAS) blockers, such as, e.g., ACE inhibitors or angiotensin-receptor blockers (ARBs).
  • RAAS renin-angiotensin-aldosterone system
  • standard of care therapy comprises immunosuppressive agents and RAAS blockers (e.g., ACE inhibitors or ARBs).
  • standard of care comprises blood pressure control (e.g., to ⁇ 130/80 mmHg) and/or HMG-CoA reductase inhibitors (i.e., statins), particularly in patients with hyperlipidemia.
  • a subject or population thereof is receiving or has received an immunosuppressive or immunomodulatory agent. In some embodiments, a subject or population thereof is receiving or has received therapy comprising an immunosuppressive or immunomodulatory agent and has been stable on said therapy for at least 12 weeks.
  • a subject or population thereof is receiving or has received a RAAS blocker. In some embodiments, a subject or population thereof is receiving or has received an ACE inhibitor. In some embodiments, a subject or population thereof is receiving or has received an ARB. In some embodiments, a subject or population thereof is receiving or has received a maximally tolerated or recommended dose of a RAAS blocker (e.g., an ACE inhibitor or ARB). In some embodiments, a subject or population thereof is receiving or has received an inhibitor of mineralocorticoid receptor. In some embodiments, a subject or population thereof is receiving or has received an inhibitor of sodium glucose co-transporter-2 (SGLT-2). In some embodiments, a subject or population thereof is receiving or has received therapy comprising a RAAS blocker (e.g., an ACE inhibitor or ARB) or SGLT-2 inhibitor and has been stable on said therapy for at least 12 weeks.
  • a RAAS blocker e.g., an ACE inhibitor or ARB
  • SGLT-2 inhibitor
  • a subject or population thereof is receiving or has received therapy comprising an immunosuppressive or immunomodulatory agent and one or more of a RAAS blocker (e.g., ACE inhibitor or ARB), mineralocorticoid receptor inhibitor or SGLT-2 inhibitor.
  • a RAAS blocker e.g., ACE inhibitor or ARB
  • mineralocorticoid receptor inhibitor e.g., mineralocorticoid receptor inhibitor or SGLT-2 inhibitor.
  • the subject or population thereof has been stable on said therapy for at least 12 weeks.
  • a subject or population thereof is not receiving non-steroidal anti-inflammatory agents (NSAIDS). In some embodiments, a subject or population thereof is not receiving non-steroidal anti-inflammatory agents (NSAIDS) chronically.
  • NSAIDS non-steroidal anti-inflammatory agents
  • a subject or population thereof is negative for hepatitis B virus, hepatitis C virus, and/or human immunodeficiency virus. In some embodiments, a subject or population thereof does not have a history of treated hepatitis C virus.
  • a subject or population thereof does not have any one or more hematologic abnormalities selected from hemoglobin ⁇ 8 g/dL, platelets ⁇ 50,000, and absolute neutrophil count (ANC) ⁇ 1000 cells/pL. In some embodiments, a subject or population thereof does not have hemoglobin AIC > 8.5%.
  • a subject or population thereof does not have any one or more liver function results selected from aspartate aminotransferase (AST), alanine aminotransferase (ALT), or total bilirubin > 2x upper limit of normal (ULN).
  • AST aspartate aminotransferase
  • ALT alanine aminotransferase
  • UPN upper limit of normal
  • a subject or population thereof has no known predisposition to bleeding.
  • a subject or population thereof does not require fibrinolysis, full-dose therapeutic anti coagulation (e.g., vitamin K antagonists, dabigatran, heparin, hirudin, etc.), or high dose antiplatelet therapy.
  • a subject or population thereof has no history of hemorrhagic central nervous system events, e.g., within 12 months. In some embodiments, a subject or population thereof has no history of active gastrointestinal bleeding, e.g., within 6 months. In some embodiments, a subject or population thereof has no history of thrombotic events (including, e.g., stroke or transient ischemic attack), e.g., within 12 months.
  • a subject or population thereof is not diagnosed with Type I diabetes mellitus. In some embodiments, a subject or population thereof has never had (e.g., in the last 5 years) a renal biopsy showing histopathological evidence of diabetic kidney disease. In some embodiments, a subject or population thereof is not diagnosed with anti-PLA2R antibody- associated membranous nephropathy.
  • a subject or population thereof has not had a myocardial infarction or unstable angina, e.g., within 6 months.
  • a subject or population thereof has no history of solid organ or hematopoietic cell transplantation. In some embodiments, a subject or population thereof is not on an organ transplant waiting list.
  • a subject or population thereof has no history or presence of any form of cancer except excised basal or squamous cell carcinoma of the skin, e.g., within 2 years.
  • a subject or population thereof is not suffering from renal disease secondary to systemic disease, including but not limited to systemic lupus erythematosus, anti-neutrophil cytoplasmic antibodies-associated diseases, anti-glomerular basement disease, secondary forms of focal segmental glomerulosclerosis, renal diseases associated with para- proteinemias, C3 glomerulopathy, and diabetic kidney disease.
  • a subject or population thereof is not suffering from a known systemic disorder that requires or is expected to require systemic glucocorticoids or immune modulators.
  • a subject or population thereof has not received treatment with anti-CD20 monoclonal antibodies, e.g., within 6 months.
  • a subject or population thereof has a BMI of greater than or equal to 40 kg/m 2 .
  • a subject or population thereof is suffering from or is susceptible to a dermal disease, disorder, or condition. In some embodiments, a subject or population thereof is suffering from or is susceptible to dermal fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to dermal fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to scleroderma and/or systemic sclerosis (e.g., diffuse systemic sclerosis or limited systemic sclerosis).
  • systemic sclerosis e.g., diffuse systemic sclerosis or limited systemic sclerosis
  • a subject or population thereof is suffering from or is susceptible to a gastrointestinal disease, disorder, or condition.
  • a subject or population thereof is suffering from or is susceptible to gastrointestinal fibrosis (e.g., fibrosis of esophagus, stomach, intestines, and/or colon).
  • a subject or population thereof is suffering from or is susceptible to gastrointestinal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to inflammatory bowel disease (e.g., ulcerative colitis or Crohn’s disease).
  • a subject or population thereof is suffering from or is susceptible to cardiac fibrosis and/or fibrosis associated with cardiovascular system. In some embodiments, a subject or population thereof is suffering from or is susceptible to cardiac fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to ischemic heart disease, myocardial ischemia, atherosclerosis, myocardial perfusion (e.g., as a consequence of chronic cardiac ischemia or myocardial infarction), vascular occlusion, or restenosis.
  • a subject or population thereof is suffering from or is susceptible to fibrosis associated with central nervous system (CNS) and/or one or more CNS- related diseases, disorders, or conditions.
  • CNS central nervous system
  • a subject or population thereof is suffering from or is susceptible to CNS-associated fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to cerebral infarction, stroke, or amyotrophic lateral sclerosis.
  • a subject or population thereof is suffering from or is susceptible to fibrosis associated with musculoskeletal system and/or one or more musculoskeletal diseases, disorders, or conditions.
  • a subject or population thereof is suffering from or is susceptible to musculoskeletal-associated fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to muscular dystrophy.
  • a subject or population thereof is suffering from or is susceptible to pancreatic fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to pancreatic fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to pancreatitis.
  • the present disclosure also provides methods related to treatment of fibrotic and related disease(s) (e.g., as described herein, such as kidney fibrotic disease(s) or lung fibrotic disease(s)) and selecting, identifying, and/or characterizing patients likely to benefit from a treatment with Compound 1 as described herein.
  • fibrotic and related disease(s) e.g., as described herein, such as kidney fibrotic disease(s) or lung fibrotic disease(s)
  • the present disclosure is based in part on the discovery that certain biomarkers can distinguish patients who are likely to respond to therapy, for example because the drivers of their disease (e.g., as described herein, such as kidney disease or lung disease) correspond with a mechanism of action of Compound 1.
  • a patient to be treated with a method of the present disclosure has an altered level of one or more gene products or one or more proteins (or fragments thereof) that are part of the mechanism of action of Compound 1 (e.g., down regulated by Compound 1).
  • a biomarker is a component of a biological sample that may be detected and/or quantified when present in the biological sample.
  • a patient to be treated with a method of the present disclosure has an elevated level of gene expression (e.g., COL3A1 expression and/or COL1A1 expression).
  • a patient to be treated with a method of the present disclosure has an elevated level of protein expression (e.g., COL3A1 protein and/or COL1 A1 protein) or elevated level of a fragment thereof.
  • an elevated level of expression e.g., of COL3A1 and/or COL1 Al
  • a gene product e.g., mRNA expressed from a COL3A1 gene and/or COL1A1 gene
  • protein e.g., COL3A1 protein and/or COL1 Al protein
  • a non-italicized name will be used to refer to both the gene and protein product.
  • a level of COL3A1 and/or COL1A1 corresponds to a level of gene expression (e.g., RNA expression, e.g., mRNA expression).
  • a level of COL3A1 and/or COL1 Al corresponds to a level of collagen protein expression.
  • collagen protein expression includes expression of a protein in any form, including, a procollagen polypeptide (e.g., type III procollagen or type I procollagen), a collagen protein (e.g., type III collagen or type I collagen), a preprocollagen polypeptide, an amino-terminal procollagen polypeptide (e.g., pN-type III collagen (“PIIINP”) or pN-type I collagen (“PINP”)), collagen fibril, collagen fiber, and/or any fragment or degradation product thereof.
  • a procollagen polypeptide e.g., type III procollagen or type I procollagen
  • collagen protein e.g., type III collagen or type I collagen
  • preprocollagen polypeptide e.g., an amino-terminal procollagen polypeptide (e.g., pN-type III collagen (“PIIINP”) or pN-type I collagen (“PINP”)), collagen fibril, collagen fiber, and/or any fragment or degradation product thereof.
  • PIIINP pN
  • a level of COL3 Al refers to a gene product (e.g., RNA, e.g., mRNA) or a protein product (e.g., a prepropolypeptide, propolypeptide, amino- terminal propolypeptide, protein, fibril, fiber, or any fragment or degradation product thereof).
  • a gene product e.g., RNA, e.g., mRNA
  • a protein product e.g., a prepropolypeptide, propolypeptide, amino- terminal propolypeptide, protein, fibril, fiber, or any fragment or degradation product thereof.
  • a level of COL1A1 refers to a gene product (e.g., RNA, e.g., mRNA) or a protein product (e.g., a prepropolypeptide, propolypeptide, amino-terminal propolypeptide, protein, fibril, fiber, or any fragment or degradation product thereof).
  • a gene product e.g., RNA, e.g., mRNA
  • a protein product e.g., a prepropolypeptide, propolypeptide, amino-terminal propolypeptide, protein, fibril, fiber, or any fragment or degradation product thereof.
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease comprising: (i) obtaining or determining a level of expression of a biomarker (e.g., COL3 Al and/or COL1 Al) in a biological sample from the patient; (ii) comparing the level of biomarker expression with a threshold level (e.g., a predetermined mean or median level of a population of healthy subjects), wherein if the expression level of the biomarker is different from the threshold level (e.g., above the threshold level or below the threshold level), administering to the patient an effective amount of Compound 1, or a pharmaceutical composition thereof.
  • obtaining a level of expression of a biomarker may comprise obtaining knowledge of a level that has been determined previously (e.g., obtaining said level from a report
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease comprising: (i) obtaining or determining a level of expression of a biomarker (e.g., COL3A1 and/or COL1 Al) in a biological sample from the patient; (ii) comparing the level of biomarker expression with a threshold level (e.g., a predetermined mean or median level of a population of healthy subjects), wherein if the expression level of the biomarker is above the threshold level, administering to the patient an effective amount of Compound 1, or a pharmaceutical composition thereof.
  • a biomarker e.g., COL3A1 and/or COL1 Al
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease comprising administering an effective amount of Compound 1, or a pharmaceutical composition thereof, to a patient that has been determined to have an altered (e.g., elevated or reduced) level of a biomarker (e.g., COL3 Al and/or COL1 Al).
  • a biomarker e.g., COL3 Al and/or COL1 Al
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease comprising administering an effective amount of Compound 1, or a pharmaceutical composition thereof, to a patient that has been determined to have an elevated level of a biomarker (e.g., COL3 Al and/or COL1 Al).
  • a biomarker e.g., COL3 Al and/or COL1 Al.
  • an elevated level of a biomarker comprises a level that is above that of a corresponding threshold level.
  • a reduced level of COL3A1 and/or COL1 Al comprises a level that is below a corresponding threshold level.
  • the level of biomarker (e.g., COL3A1 and/or COL1 Al) was obtained from or previously determined from a biological sample from the patient.
  • the method further comprises obtaining or determining a level of a biomarker (e.g., COL3A1 and/or COL1 Al) in a biological sample from the patient.
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney or the lung
  • a fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney or the lung
  • a biomarker e.g., COL3A1 and/or COL1A1
  • an elevated level of a biomarker comprises a level that is above a corresponding threshold level.
  • the level of biomarker (e.g., COL3A1 and/or COL1A1) was obtained from or previously determined from a biological sample from the patient.
  • the method further comprises obtaining or determining a level of a biomarker (e.g., COL3A1 and/or COL1 Al) in a biological sample from the patient.
  • a threshold expression level corresponds to a predetermined mean or median level of COL3A1 and/or COL1A1 of a population of healthy subjects (e.g., healthy human subjects). In some embodiments, a threshold expression level corresponds to a predetermined normal range of COL3A1 and/or COL1A1 of a population of healthy subjects (e.g., healthy human subjects).
  • the expression level of COL3 Al is at least 20% higher than the corresponding threshold expression level. In some embodiments, the expression level of COL1 Al is at least 20% higher than the corresponding threshold expression level. In some embodiments, the expression levels of both COL3A1 and COL1 Al are at least 20% higher than the respective corresponding threshold expression levels.
  • the expression level of COL3 Al is at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, or at least 50% higher than the corresponding threshold expression level.
  • the expression level of COL1 Al is at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, or at least 50% higher than the corresponding threshold expression level.
  • the expression levels of both COL3A1 and COL1 Al are at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, or at least 50% higher than the respective corresponding threshold expression levels.
  • a biological fluid may be or comprise an intracellular fluid, an extracellular fluid, an intravascular fluid (blood plasma), an interstitial fluid, a lymphatic fluid, and/or a transcellular fluid.
  • a biological tissue or sample may be obtained, for example, by aspirate, biopsy (e.g., fine needle or tissue biopsy), swab (e.g., oral, nasal, skin, or vaginal swab), scraping, surgery, washing or lavage (e.g., bronchoalveolar, ductal, nasal, ocular, oral, uterine, vaginal, or other washing or lavage).
  • the biological sample (e.g., from which an expression level of a biomarker is obtained or has been determined) is a renal biopsy sample, a urine sample or a blood sample.
  • an expression level of a protein biomarker e.g., a prepropeptide, propeptide, amino-terminal propeptide, protein, fibril or fiber, e.g., of COL3A1 and/or of COL1 Al
  • a protein biomarker e.g., a prepropeptide, propeptide, amino-terminal propeptide, protein, fibril or fiber, e.g., of COL3A1 and/or of COL1 Al
  • an expression level of a genetic biomarker e.g., a gene product, e.g., RNA, e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • a genetic biomarker e.g., a gene product, e.g., RNA, e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • RNA e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • the biological sample (e.g., from which an expression level of a biomarker is or has been determined) is a lung biopsy sample, a bronchoalveolar lavage fluid (BALF) sample or a blood sample.
  • a protein biomarker e.g., a prepropeptide, propeptide, amino-terminal propeptide, protein, fibril or fiber, e.g., of COL3A1 and/or of COL1 Al
  • a protein biomarker e.g., a prepropeptide, propeptide, amino-terminal propeptide, protein, fibril or fiber, e.g., of COL3A1 and/or of COL1 Al
  • an expression level of a genetic biomarker e.g., a gene product, e.g., RNA, e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • a genetic biomarker e.g., a gene product, e.g., RNA, e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • a genetic biomarker e.g., a gene product, e.g., RNA, e.g., mRNA, e.g., mRNA encoding COL3A1 and/or COL1A1
  • an expression level of a biomarker is obtained from or determined in a renal biopsy sample from a patient.
  • an expression level of a biomarker e.g., COL3A1 and/or COL1 Al
  • a gene biomarker e.g., RNA, e.g., mRNA
  • the biological sample is a renal biopsy sample and the expression level of COL3 Al is an mRNA level and/or the expression level of COL1A1 is an mRNA level.
  • the biological sample is a renal biopsy sample and the expression level of COL3 Al is an mRNA level and/or the expression level of COL1 A1 is an mRNA level.
  • a biomarker e.g., mRNA level
  • Methods for measuring and/or determining levels of a biomarker (e.g., mRNA level) in renal biopsy samples are known in the art, for example, Genovese et ah, Biomark Insights . 2016 May 22;ll:77-84.
  • an expression level of a biomarker is obtained from or determined in a urine sample from a patient.
  • the biological sample is a urine sample and the expression level of COL3 A1 is a level of COL3 A1 protein or fragments thereof and/or the expression level of COL1 A1 is a level of COL1 A1 protein or fragments thereof.
  • Methods for measuring and/or determining levels of a biomarker (e.g., a level of collagen protein and/or fragment thereof) in urine samples are known in the art, for example, Soylemezoglu et ah, Nephrol Dial Transplant. 1997 Sep;12(9):1883-9.
  • an expression level of a biomarker is obtained from or determined in a blood sample from a patient.
  • the biological sample is a blood sample and the expression level of COL3 A1 is a level of COL3 A1 protein or fragments thereof and/or the expression level of COL1 A1 is a level of COL1 A1 protein or fragments thereof.
  • Methods for measuring and/or determining levels of a biomarker (e.g., a level of collagen protein and/or fragment thereof) in blood samples are known in the art.
  • an expression level of a biomarker is obtained from or determined in a lung biopsy sample from a patient.
  • an expression level of a biomarker e.g., COL3A1 and/or COL1 Al
  • a gene biomarker e.g., RNA, e.g., mRNA
  • the biological sample is a lung biopsy sample and the expression level of COL3 Al is an mRNA level and/or the expression level of COL1A1 is an mRNA level.
  • the biological sample is a lung biopsy sample and the expression level of COL3 Al is an mRNA level and/or the expression level of COL1 Al is an mRNA level.
  • Methods for measuring and/or determining levels of a biomarker (e.g., mRNA level) in lung biopsy samples are known in the art.
  • an expression level of a biomarker is obtained from or determined in a BALF sample from a patient.
  • the biological sample is a BALF sample and the expression level of COL3 Al is a level of COL3 Al protein or fragments thereof and/or the expression level of COL1 Al is a level of COL1 Al protein or fragments thereof.
  • Methods for measuring and/or determining levels of a biomarker (e.g., a level of collagen protein and/or fragment thereof) in BALF samples are known in the art, for example, Wattiez, R. and Falmagne, P., J Chromatogr B Analyt Technol Biomed Life Sci. 2005 Feb 5;57(9):2005-l 1.
  • the present disclosure provides methods of administering Compound 1 to a subject or population of subjects described herein, according to a regimen established to achieve one or more desirable outcomes.
  • the fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney or the lung
  • the fibrotic disease is stabilized (i.e., does not worsen) and/or is ameliorated (i.e., one or more symptoms improve) in a patient treated with Compound 1.
  • treatment of a patient with Compound 1 reduces a level of one or more biomarkers (e.g., COL1 A1 and/or COL3A1).
  • treatment of a patient with Compound 1 reduces proteinuria.
  • a regimen has been established to achieve one or more desirable outcomes, relative to that observed for a comparable reference population that has not received Compound 1 (e.g., that has received a placebo), such as those described herein.
  • a reference composition may be or may have been administered at the same intervals and/or in the same amounts as a composition providing Compound 1.
  • a regimen has been established to achieve one or more desirable outcomes, relative to that observed for a comparable reference population that does not have an altered level of a biomarker (e.g., an elevated level of COL3A1 and/or COL1 Al).
  • a patient that expresses a biomarker treated with Compound 1 has an improved outcome (e.g., improved stability and/or amelioration of the fibrotic disease) relative to a patient with the same disease that does not express an elevated level of the biomarker.
  • treatment of a patient with Compound 1 reduces a level of one or more biomarkers (e.g., COL1 Al and/or COL3A1).
  • treatment of a patient with Compound 1 reduces proteinuria.
  • a composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, e.g., at a particular time point, such as regimens described herein. In some embodiments, a composition providing Compound 1 is administered according to a regimen established to achieve a particular effect, such as regimens described herein. [0230] In some embodiments, a patient with a fibrotic disease of the kidney exhibits increased renal COL3 A1 expression that is correlated with urine protein to creatinine ratio. In some embodiments, a patient with a fibrotic disease of the kidney exhibits COL3 A1 expression that is inversely correlated with eGFR.
  • a kidney disease that is or comprises focal and segmental glomerulosclerosis (FSGS).
  • a patient has FSGS and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a patient has FSGS and also exhibits (i) increased renal COL3 A1 expression that is correlated with urine protein to creatinine ratio and/or (ii) COL3 A1 expression that is inversely correlated with eGFR.
  • provided methods are useful in treating a kidney disease that is or comprises primary proteinuric kidney disease (PPKD).
  • PPKD primary proteinuric kidney disease
  • a patient has PPKD and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • provided methods are useful in treating a kidney disease that is or comprises primary glomerular disease (PGD).
  • PGD primary glomerular disease
  • a patient has PGD and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises steroid resistant nephrotic syndrome (SRNS).
  • SRNS steroid resistant nephrotic syndrome
  • a patient has SRNS and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • provided methods are useful in treating a kidney disease that is or comprises proteinuria.
  • a patient has proteinuria and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises lupus nephritis.
  • a patient has lupus nephritis and an elevated level of COL3A1 and/or COL1A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises minimal change disease.
  • a patient has minimal change disease and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises ANCA-associated glomerulonephritis.
  • a patient has ANCA-associated glomerulonephritis and an elevated level of COL3A1 and/or COL1A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises anti-GBM nephropathy.
  • a patient has anti-GBM nephropathy and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises IgA nephropathy.
  • a patient has IgA nephropathy and an elevated level of COL3A1 and/or COL1A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises membranous glomerulonephritis (MG).
  • MG membranous glomerulonephritis
  • a patient has MG and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises polycystic kidney disease (e.g., autosomal dominant polycystic kidney disease (ADPKD)).
  • ADPKD autosomal dominant polycystic kidney disease
  • a patient has polycystic kidney disease (e.g., ADPKD) and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • ADPKD polycystic kidney disease
  • COL3A1 and/or COL1 A1 e.g., mRNA and/or protein
  • provided methods are useful in treating a kidney disease that is or comprises chronic kidney disease.
  • a patient has chronic kidney disease and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • COL3A1 and/or COL1 A1 e.g., mRNA and/or protein
  • provided methods are useful in treating a kidney disease that is or comprises collagen type III glomerulopathy.
  • a patient has collagen type III glomerulopathy and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • a biological sample from the patient e.g., renal biopsy sample, urine sample, blood sample, etc.
  • provided methods are useful in treating a kidney disease that is or comprises nail-patella syndrome.
  • a patient has nail-patella syndrome and an elevated level of COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein), for example, as determined in or obtained from a biological sample from the patient (e.g., renal biopsy sample, urine sample, blood sample, etc.).
  • COL3A1 and/or COL1 A1 e.g., mRNA and/or protein
  • provided methods are useful in treating a lung disease that is or comprises idiopathic pulmonary fibrosis.
  • a patient has idiopathic pulmonary fibrosis and an elevated level of one or more biomarkers, e.g., identified as described in Example 22 (optionally, e.g., COL3A1 and/or COL1 A1 (e.g., mRNA and/or protein)), for example, as determined in or obtained from a biological sample from the patient (e.g., lung biopsy sample, BALF sample, blood sample, etc.).
  • biomarkers e.g., identified as described in Example 22
  • COL3A1 and/or COL1 A1 e.g., mRNA and/or protein
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease comprising: (i) obtaining or determining a level of expression of one or more gene products or proteins in a biological sample from the patient, wherein the one or more gene products or proteins are selected from: COL3A1 and COL1A1; (ii) comparing the expression level of the one or more gene products or proteins with that of a corresponding threshold level, and (iii) if the expression level of COL3 A1 and/or the expression level of COL1 A1 is above the threshold expression level, then administering to the patient an effective amount of Compound 1 or a pharmaceutical composition thereof, and if the expression level of COL3A1
  • a fibrotic disease e.g., a fibrotic disease as described herein, e.g., a fibrotic disease of the kidney or the lung
  • a fibrotic disease e.g., a fibrotic disease as described herein, e.g., a fibrotic disease of the kidney or the lung
  • a biomarker e.g., COL3A1 and/or COL1 Al
  • the method comprising: (i) administering an effective amount of Compound 1 or a pharmaceutical composition thereof, and (ii) monitoring a level of biomarker (e.g., COL3A1 and/or COL1A1) subsequent to administration (e.g., after a duration of a day, week, two weeks, month, two months, 3 months, etc.).
  • the level of biomarker e.g., COL3A1 and/or COL1 Al
  • further treatment with Compound 1 is discontinued.
  • the level of biomarker e.g., COL3A1 and/or COL1 Al
  • the dose of Compound 1 administered to the patient is increased.
  • the level of biomarker e.g., COL3A1 and/or COL1 Al
  • treatment with Compound 1 is continued (e.g., subsequent doses are administered).
  • a fibrotic disease e.g., a fibrotic disease as described herein, e.g., a fibrotic disease of the kidney or the lung
  • a fibrotic disease is stabilized (i.e., does not worsen) and/or is ameliorated (i.e., one or more symptoms improve) in a patient treated with Compound 1.
  • treatment of a patient with Compound 1 reduces a level of one or more biomarkers (e.g., COL1 Al and/or COL3A1).
  • administration of Compound 1 to the patient (i) reduces the expression of renal COL3A1 , (ii) reduces the expression of renal COL1A1 , (iii) reduces renal COL3A1 accumulation, (iv) reduces renal COL1A1 accumulation, or (v) any combination thereof.
  • administration of Compound 1 to the patient (i) reduces the expression of pulmonary COL3A1 , (ii) reduces the expression of pulmonary COL1A1 , (iii) reduces pulmonary COL3A1 accumulation, (iv) reduces pulmonary COL1 A1 accumulation, or (v) any combination thereof.
  • a patient to be treated with a method of the present disclosure exhibits proteinuria.
  • a method for treatment with Compound 1 that includes: (i) receiving a report listing the expression level of one or more biomarkers (e.g., COL3A1 and/or COL1 Al) for a patient with a fibrotic disease (e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney or the lung), (ii) receiving a request for reimbursement of the screening and/or of a particular therapeutic regimen; and (iii) approving payment and/or reimbursement for Compound 1 therapy if the report indicates the level of biomarker is above a threshold level.
  • a fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney or the lung
  • the present disclosure provides certain biomarkers that can distinguish subjects (e.g., subjects suffering from or at risk of fibrosis or associated diseases, disorders, and conditions) who are more likely than others to respond to therapy with Compound 1.
  • the present disclosure provides insight that certain biomarkers can distinguish patients who are likely to respond to therapy, for example because the drivers of their fibrotic disease correspond with the mechanism of action of Compound 1.
  • an altered level of one or more gene products or proteins that are part of the mechanism of action of Compound 1 e.g., down- or up-regulated by Compound 1).
  • one or more biomarkers comprise an elevated level of COL3A1 expression and/or COL1 Al expression.
  • a patient with an altered level of one or more biomarkers may have an improved response to treatment with Compound 1 relative to a patient that does not have a level of the biomarker that meets the threshold criteria.
  • one biomarker is used to characterize subjects; in some embodiments, more than one biomarker (e.g., two, three, etc.) is used to characterize subjects.
  • a biomarker is differentially present in a sample taken from a subject of one status as compared with another status (e.g., more responsive to Compound 1 therapy vs. less responsive to Compound 1 therapy).
  • a biomarker is differentially present in a sample taken from the same subject at two or more different time points, i.e., when the status of the subject has changed from one time point to another.
  • a biomarker is detected and/or quantified in a tissue sample (e.g., from a biopsy, such as a kidney or lung biopsy) and/or in a biological fluid (e.g., blood, urine, BALF, etc.).
  • a biomarker is a level of mRNA that is detected and/or quantified in a kidney tissue sample, e.g., obtained from a kidney biopsy.
  • a biomarker is detected and/or quantified in a urine sample (e.g., a level of a protein or protein fragment).
  • a biomarker is detected and/or quantified in a blood sample (e.g., a level of a protein or protein fragment). In some embodiments, a biomarker is detected and/or quantified in a BALF sample (e.g., a level of a protein or protein fragment). In some embodiments, a biomarker is a level of mRNA that is detected and/or quantified in a lung tissue sample, e.g., obtained from a lung biopsy.
  • a biomarker may include one or more of a peptide, protein, nucleic acid (e.g., polynucleotide, DNA, RNA, etc.), polysaccharide (e.g., lectins or sugars), lipid, enzyme, small molecule, ligand, receptor, antigen, or antibody.
  • a biomarker comprises a protein.
  • a biomarker comprises a nucleic acid (e.g., mRNA).
  • detection of a threshold level of one or more biomarkers are used to select and/or characterize patients who may be responsive to Compound 1 therapy.
  • levels of one or more biomarkers in a sample from a subject are compared to a threshold level.
  • a biomarker is considered increased if the level is increased relative to a threshold level (e.g., increased by at least about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 100%, or more).
  • a threshold level is determined from a population of healthy volunteers (e.g., a mean or median level from a population of healthy volunteers).
  • a method includes an in vitro method for determining a level of a biomarker.
  • in vitro methods for determining a level of a biomarker include, but are not limited to, a chemiluminescence assay, enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay, colorimetric assay, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), High Performance Liquid Chromatography (“HPLC”), and/or PCR (e.g., real-time PCR).
  • HPLC High Performance Liquid Chromatography
  • a method for detecting a level of a biomarker includes chromatographic and/or MS methods.
  • Exemplary methods include, but are not limited to, gas chromatography (GC), liquid chromatography/mass spectroscopy (LC-MS), gas chromatography/mass spectroscopy (GC-MS), nuclear magnetic resonance (NMR), magnetic resonance imaging (MRI), Fourier Transform InfraRed (FT-IR), and inductively coupled plasma mass spectrometry (ICP-MS).
  • the present disclosure encompasses the recognition that COL1 A1 and/or COL3A1 are useful biomarkers in the methods provided herein. In some embodiments, the present disclosure provides insight that increased levels of COL1 A1 and/or COL3 A1 expression may be useful in selecting and/or characterizing patients for Compound 1 therapy.
  • COL1 A1 is a biomarker useful in the methods provided herein.
  • COL3 A1 is a biomarker useful in the methods provided herein.
  • an elevated level of COL1A1 and/or COL3A1 corresponds to a level of gene expression (e.g., RNA, e.g., mRNA).
  • an elevated level of COL1A1 and/or COL3A1 corresponds to a level of collagen protein expression.
  • an elevated level of COL1A1 and/or COL3A1 protein is or includes, e.g., a corresponding procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino- terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product thereof.
  • an increased level of COL1 A1 and/or COL3A1 is above a threshold level (e.g., a predetermined median or mean level). In some embodiments, an increased level of COL1 A1 and/or COL3A1 is more than about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 100%, or more of a threshold level. In some embodiments, an increased level of COL1 A1 and/or COL3A1 is more than about 0.5, about 1.0, about 1.5, or about 2.0, or more standard deviations above a threshold level.
  • a threshold level e.g., a predetermined median or mean level. In some embodiments, an increased level of COL1 A1 and/or COL3A1 is more than about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 100%, or more of a threshold level. In some embodiments, an
  • subjects who are selected for Compound 1 therapy based on increased levels of COL1 A1 and/or COL3A1 achieve improved outcomes (e.g., compared to subjects who do not meet one or more selection criteria for Compound 1 therapy).
  • the present disclosure provides methods of treating a disease, disorder, or condition characterized by increased expression of COL1 A1 and/or COL3A1, comprising administering Compound 1 to a subject or population of subjects in need thereof as described herein.
  • such a disease, disorder, or condition is a renal disease, disorder, or condition (e.g., a renal disease, disorder, or condition associated with fibrosis) as described herein.
  • a renal disease, disorder, or condition is characterized by increased expression of COL1A1 and/or COL3A1.
  • such a disease, disorder, or condition is a pulmonary disease, disorder, or condition (e.g., a pulmonary disease, disorder, or condition associated with fibrosis) as described herein.
  • a pulmonary disease, disorder, or condition is characterized by increased expression of COL1A1 and/or COL3A1.
  • the present disclosure provides methods comprising administering Compound 1 to a subject in need thereof, wherein the subject has been determined to have increased level(s) of COL1A1 and/or COL3A1.
  • a subject has been determined to have increased level(s) of COL1 A1 and/or COL3A1 in a renal biopsy sample and/or a urine sample.
  • a subject has been determined to have increased level(s) of COL1 A1 and/or COL3 A1 in a biological sample other than a renal biopsy sample and/or a urine sample.
  • a subject has been determined to have increased level(s) of COL1A1 and/or COL3A1 in aBALF sample.
  • the present disclosure provides methods comprising determining level(s) of COL1A1 and/or COL3A1 by: obtaining a biological sample from a subject; performing an assay on the sample to determine level(s) of COL1 A1 and/or COL3A1 in the sample; and comparing the determined level(s) to a threshold level.
  • a sample has increased level(s) of COL1 A1 and/or COL3A1 compared to a threshold level
  • Compound 1 is administered to the subject.
  • if a sample does not have increased level(s) of COL1 A1 and/or COL3A1 compared to a threshold level then Compound 1 is not administered to the subject.
  • the present disclosure provides methods comprising determining level(s) of COL1 A1 and/or COL3A1 in a biological sample from a subject; comparing the determined level(s) to a threshold level; and identifying the subject as in need of therapeutic intervention when the sample is determined to have increased level(s) of COL1 A1 and/or COL3A1 compared to the threshold level.
  • a subject is identified as in need of therapeutic intervention with Compound 1 therapy as described herein.
  • a provided method further comprises administering Compound 1 to a subject (e.g., a subject identified as in need of therapeutic intervention) as described herein.
  • provided technologies are useful for monitoring subjects (e.g., monitoring status of subjects over time and/or monitoring therapy).
  • the present disclosure provides methods comprising determining level(s) of COL1A1 and/or COL3A1 in each of a plurality of biological samples obtained at different time points from a single patient; and comparing the determined level(s) from a first time point with that from at least one later time point.
  • the present disclosure provides methods comprising determining level(s) of COL1 A1 and/or COL3A1 from a biological sample obtained from a subject for whom level(s) of COL1 A1 and/or COL3A1 have previously been obtained at least once; and comparing the determined level(s) with the previously obtained level(s).
  • a first time point and one or more later time points are separated from one another by a reasonably consistent interval.
  • a significant change in the determined level over time indicates a change in the subject’s status.
  • a significant change in a determined level over time is a change (e.g., an increase or a decrease) of at least about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 100%, or more compared to a threshold level.
  • a significant change in a determined level over time is a change of more than about 0.5, about 1.0, about 1.5, or about 2.0, or more standard deviations away from a threshold level.
  • such methods further comprise administering Compound 1 therapy to a subject (e.g., a subject who has been determined to have moved from a non- responsive to a responsive state) as described herein.
  • a subject e.g., a subject who has been determined to have moved from a non- responsive to a responsive state
  • provided methods are useful for monitoring therapy (e.g., efficacy and/or other indicators of response).
  • a sample from a first time point was obtained from a subject prior to administration of Compound 1
  • a sample from a second time point was obtained from a subject after administration of Compound 1.
  • Compound 1 therapy is continued.
  • Compound 1 therapy is discontinued, or dosage amount and/or frequency of Compound 1 therapy is increased.
  • a method includes an in vitro method for determining a level of a biomarker.
  • a method for determining a level of a biomarker can be or include a chemiluminescence assay, enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay, colorimetric assay, UV spectroscopy, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), High Performance Liquid Chromatography (“HPLC”), and/or PCR (e.g., quantitative PCR and/or real-time PCR).
  • a chemiluminescence assay e.g., enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay,
  • a level of a biomarker corresponds to a level of gene expression (e.g., RNA, e.g., mRNA) and is quantified using methods known in the art.
  • a method of determining a level of expression of a biomarker gene can be or include a chemiluminescence assay, UV spectroscopy, hybridization assay (e.g., Fluorescent in Situ Hybridization (FISH), e.g., RNA- FISH), enzymatic assay, enzyme immunoassay (e.g., ELISA), multiplex assay, electrophoresis assay, radioassay, colorimetric assay, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), High Performance Liquid Chromatography (“HPLC”), and/or PCR (e.g., quantitative PCR and/or real-time PCR).
  • FISH Fluorescent in Situ Hybridization
  • RNA- FISH e.g., RNA- FISH
  • enzymatic assay e.g., enzyme immunoassay (e.g., ELISA), multiplex
  • a level of a biomarker corresponds to a level of protein (e.g., procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product thereof) and is quantified using methods known in the art.
  • protein e.g., procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product thereof
  • a method of determining a level of expression of a biomarker protein can be or include a chemiluminescence assay, enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay, colorimetric assay, UV spectroscopy, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), or High Performance Liquid Chromatography (“HPLC”).
  • HPLC High Performance Liquid Chromatography
  • a method for treating a fibrotic disease of the kidney comprising: administering an effective amount of a Compound 1 or a pharmaceutical composition thereof, to a patient that has been determined to have an elevated level of COL3 A1 and/or COL1A1.
  • a method for treating a patient diagnosed with or at risk for a fibrotic disease of the kidney comprising:
  • the expression level of COL3A1 is at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, or at least 50% higher than the corresponding threshold expression level and/or the expression level of COL1 A1 is at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, or at least 50% higher than the corresponding threshold expression level.
  • fibrotic disease of the kidney is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, anti-globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), collagen type III glomerulopathy, nail-patella syndrome, or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria lupus nephritis
  • minimal change disease an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis
  • anti-GBM anti-globular basement membrane
  • ANCA anti-neutrophil cytoplasmic antibody
  • MPA microscopic polyangiitis
  • fibrotic disease of the kidney is or comprises focal and segmental glomerulosclerosis (FSGS).
  • FSGS focal and segmental glomerulosclerosis
  • a method for treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease of the kidney comprising:
  • fibrotic disease of the kidney is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, anti-globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), collagen type III glomerulopathy, nail-patella syndrome, or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria lupus nephritis
  • minimal change disease an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis
  • anti-GBM anti-globular basement membrane
  • composition providing Compound 1, as described herein, can be administered in accordance with methods provided herein.
  • a composition providing Compound 1 is a composition comprising Compound 1 (in a pharmaceutically acceptable form as described herein), formulated together with one or more pharmaceutically acceptable carriers.
  • a composition providing Compound 1 is or comprises Compound 1 present in unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population.
  • compositions providing Compound 1 may be specially formulated for administration in solid or liquid form, including those adapted for the following: oral administration, for example, drenches (aqueous or non-aqueous solutions or suspensions), capsules, tablets, e.g., those targeted for buccal, sublingual, and systemic absorption, boluses, powders, granules, pastes for application to the tongue; parenteral administration, for example, by subcutaneous, intramuscular, intravenous or epidural injection as, for example, a sterile solution or suspension, or sustained-release formulation; topical application, for example, as a cream, ointment, or a controlled-release patch or spray applied to the skin, lungs, or oral cavity; intravaginally or intrarectally, for example, as a pessary, cream, or foam; sublingually; ocularly; transdermally; or nasally, pulmonary, and to other mucosal surfaces.
  • oral administration for example, drenches (aqueous or non
  • composition providing Compound 1 is formulated for oral administration (e.g., in a capsule form). In some embodiments, a composition providing Compound 1 is administered orally.
  • a composition providing Compound 1 is administered as one or more unit dosage forms.
  • a composition providing Compound 1 is administered as one or more solid unit dosage forms (e.g., one or more capsules or tablets).
  • a composition providing Compound 1 is administered as one or more immediate release solid unit dosage forms.
  • Compound 1 is administered as one or more oral unit dosage forms.
  • a composition providing Compound 1 is a capsule. In some embodiments, a composition providing Compound l is a tablet.
  • Compound 1 is administered as a capsule comprising 10 mg, 50 mg, or 250 mg of Compound 1 In some embodiments, Compound 1 is administered as a capsule comprising 10 mg, 50 mg, or 250 mg of Compound 1 with no excipients. In some embodiments, Compound 1 is administered as a capsule comprising 50 mg of Compound 1 (e.g., with no excipients). In some embodiments, Compound 1 is administered as a capsule comprising 250 mg of Compound 1 (e.g., with no excipients). [0287] In some embodiments, Compound 1 is administered as a capsule comprising 100 mg of Compound 1 (e.g., a Form A Compound 1 Hydrochloride Trihydrate). In some embodiments, Compound 1 is administered as a capsule comprising 200 mg of Compound 1 (e.g., a Form A Compound 1 Hydrochloride Trihydrate). In some such embodiments, a capsule comprising Compound 1 has no excipients.
  • Compound 1 is administered as a capsule comprising 10 mg
  • unit dosage forms e.g., tablets or capsules
  • one unit dosage form e.g., tablet or capsule
  • a suitable dose as described herein e.g., a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg
  • more than one (e.g., 2, 3, 4, 5, etc.) unit dosage forms e.g., tablets or capsules
  • a suitable dose as described herein e.g., a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg.
  • each unit dosage form when multiple unit dosage forms are administered, each unit dosage form contains the same amount of Compound 1, in order to provide a suitable dose as described herein; in some embodiments, when multiple unit dosage forms are administered, each unit dosage form contains different amounts of Compound 1, in order to provide a suitable dose as described herein.
  • a suitable number of unit dosage forms are administered in order to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • one unit dosage form is administered to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • more than one (e.g., 2, 3, or 4) unit dosage forms are administered to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • a composition providing Compound 1 is administered as a single dose. In some embodiments, a composition providing Compound 1 is administered at regular intervals. Administration at an “interval,” as used herein, indicates that the therapeutically effective amount is administered periodically (as distinguished from a one-time dose). In some embodiments, a composition providing Compound 1 is administered bimonthly (Q2M), monthly (QM), twice monthly (BIM), triweekly (Q3W), biweekly (Q2W), weekly (QW), twice weekly (BIW), thrice weekly (TIW), daily (QD), twice daily (BID), thrice daily (TID), or four times a day (QID) in accordance with methods provided herein. In some embodiments, a composition providing Compound 1 is administered twice daily (BID). In some embodiments, a composition providing Compound 1 is administered once daily (QD).
  • the present disclosure encompasses the recognition that, in some embodiments, administration of Compound 1 once daily is desirable for, e.g., patient convenience and/or compliance. Alternatively and/or additionally, the present disclosure encompasses the recognition that, in some embodiments, administration of Compound 1 twice daily may be desirable to achieve certain outcomes (e.g., to meet a particular threshold blood concentration, etc.).
  • a composition providing Compound 1 is administered in a suitable number of unit dosage forms (e.g., tablets or capsules) in order to provide a dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, or about 600 mg once daily.
  • a composition providing Compound 1 is administered in a suitable number of unit dosage forms (e.g., tablets or capsules) in order to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, or about 400 mg twice daily.
  • a composition providing Compound 1 is administered at regular intervals indefinitely. In some embodiments, a composition providing Compound 1 is administered at regular intervals for a defined period of time. In some embodiments, a composition providing Compound 1 is administered at regular intervals for at least 12 weeks. [0294] In some embodiments, a composition providing Compound 1 is administered to a subject in a fed state (e.g., after a meal, such as within 1 hour, 45 minutes, 30 minutes, or 15 minutes of a meal). In some embodiments, Compound 1 is administered to a subject in a fasted state (e.g., after a fast of at least 4 hours, at least 6 hours, at least 8 hours, at least 10 hours, at least 12 hours, or at least 16 hours).
  • composition providing Compound 1 is administered to a subject at least 30 minutes apart from any other medications.
  • Nintedanib is approved by the FDA for treatment of idiopathic pulmonary fibrosis, for treatment chronic fibrosing interstitial lung diseases with a progressive phenotype and for slowing the rate of decline in pulmonary function in patients with systemic sclerosis-associated interstitial lung disease (OFEV ® [package insert] Ridgefield, CT: Boehringer Ingelheim Pharmaceuticals, Inc.; March 2020).
  • Nintedanib is an oral tyrosine kinase inhibitor and has a recommended dosage of 150 mg twice daily approximately 12 hours apart taken with food.
  • Nintedanib is also referred to as liT-indole-6-carboxylic acid, 2,3-dihydro-3-[[[4-[methyl[(4- methyl-l-piperazinyl)acetyl]amino]phenyl]amino]phenylmethylene]-2-oxo-, methyl ester, (3Z)-, ethanesulfonate.
  • Nintedanib is reported to reach maximum plasma concentrations approximately 2 to 4 hours after oral administration as a soft gelatin capsule under fed conditions.
  • the absolute bioavailability of a 100 mg dose was 4.7% (90% CL 3.62 to 6.08) in healthy volunteers.
  • nintedanib exposure increased by approximately 20% compared to administration under fasted conditions (90% CL 95.3% to 152.5%) and absorption was delayed (median tmax fasted: 2.00 hours; fed: 3.98 hours), irrespective of food type.
  • the effective half-life of nintedanib in patients with IPF was 9.5 hours (gCV 31.9%). Total plasma clearance after intravenous infusion was high (CL: 1390 mL/min; gCV 28.8%).
  • Urinary excretion of unchanged drug within 48 hours was about 0.05% of the dose after oral and about 1.4% of the dose after intravenous administration; the renal clearance was 20 mL/min.
  • Fig. 2 of Wind, S., et al. Clin. Pharmacokinet. 2019;58:1131-47 A trough concentration for producing 80% of Umax (ECxo) on FVC decline was estimated to be > 10 ng/mL.
  • Fig. 6 of Wind, S., et al. Clin. Pharmacokinet. 2019;58:1131-47 was estimated to be > 10 ng/mL.
  • Nintedanib is not recommended for use in patients with moderate or severe hepatic impairment. Elevated liver enzymes (e.g., ALT, AST, and bilirubin) and drug-induced liver injury have occurred with nintedanib. Diarrhea, nausea, and vomiting have occurred with nintedanib. Nintedanib may cause fetal harm. Arterial thromboembolic events have occurred with nintedanib. Bleeding events have occurred with nintedanib. Gastrointestinal perforation has been reported with nintedanib. See OFEV ® [package insert] Ridgefield, CT: Boehringer Ingelheim Pharmaceuticals, Inc.; March 2020.
  • the present disclosure also encompasses the recognition that provided technologies may provide different (e.g., improved) properties and/or outcomes, compared to one or more other known therapies (e.g., nintedanib).
  • Compound 1 may display different (e.g., improved) pharmacokinetic, pharmacodynamic, safety and/or efficacy characteristics, compared to one or more other known therapies (e.g., nintedanib).
  • a representative procedure for obtaining Compound 1 is as follows.
  • the quantities of materials used are approximate and may be increased or decreased in unison to obtain a larger or smaller lot size.
  • Conditions such as time or temperature are approximate and may be used as targets.
  • Step 1 Acetic anhydride (5.40 kg) and methyl 2-oxo-2,3-dihydro-lH-pyrrolo[2, 3- b]pyridine-6-carboxylate (1.0 kg) were added to a reactor at room temperature and stirred to combine. Trimethylorthobenzoate (1.90 kg) was added to the reaction mixture. The mixture was then heated to 105 °C and stirred for 1 hr. The reaction was cooled to 40 °C and isopropyl alcohol (3.14 kg) was added. The reaction was cooled further to 5 °C and stirred for 4 hr. The mixture was then filtered, and the product washed with isopropyl alcohol twice.
  • Step 2 Methanol (7.12 kg), methyl (E)-l -acetyl-3 -(methoxy(phenyl)methylene)-2- oxo-2, 3-dihydro-lH-pyrrolo[2,3-b]pyridine-6-carboxylate (1.0 kg), and A f -(4-aminophenyl)-N- methyl-2-(4-methylpiperazin-l-yl)acetamide (0.78 kg) in methanol (0.79 kg) were added to a reactor at room temperature and stirred to combine. The reaction mixture was then heated to 63 °C and stirred for 4 hrs. The reaction mixture was cooled to 5 °C and stirred for 4 hrs.
  • Step 3 Purified water (2.50 kg) and methyl (Z)-3-(((4-(N-methyl-2-(4- methylpiperazin-l-yl)acetamido)phenyl)amino)(phenyl)methylene)-2-oxo-2,3-dihydro-lH- pyrrolo[2,3-b]pyridine-6-carboxylate (1.0 kg) were added to a reactor at room temperature and stirred to combine. Acetone (1.975 kg) was then added, followed by iron-free HC1 (1.20 mol eq.). The mixture was stirred for 1 hr at 30 °C, and then filtered through a micron filter and washed with purified water (0.2 kg).
  • Compound 1 was formulated in a capsule for oral administration.
  • the capsule formulation included a Size 00 Swedish orange capsule containing Compound 1 (10 mg, 50 mg, or 250 mg) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose), iron oxide as a coloring agent, and titanium dioxide as an opacifier.
  • Capsule formulations were prepared as follows. First, an optional sieving step was performed to deagglomerate the active agent if needed. Then, Compound 1 was filled into HPMC capsules, using either an automated Xelodose machine (e.g., for 10 mg and 50 mg capsules) or a semiautomated process (e.g., for 250 mg capsules). All capsules were polished or dedusted, either by an inline deduster (e.g., for 10 mg and 50 mg capsules) or a separate capsule polisher (e.g., for 250 mg capsules).
  • an automated Xelodose machine e.g., for 10 mg and 50 mg capsules
  • a semiautomated process e.g., for 250 mg capsules. All capsules were polished or dedusted, either by an inline deduster (e.g., for 10 mg and 50 mg capsules) or a separate capsule polisher (e.g., for 250 mg capsules).
  • Compound 1 Hydrochloride Form A was formulated in a capsule for oral administration.
  • the capsule formulation included a Swedish Orange capsule containing Compound 1 Hydrochloride Form A (100 mg or 200 mg dose) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose), iron oxide as a coloring agent, and titanium dioxide as an opacifier.
  • Capsule formulations were prepared as described above.
  • Example 2 A Phase 1, Randomized, Double-Blind, Placebo-controlled, Single and Multiple Ascending Dose Study to Determine the Safety, Tolerability, Pharmacokinetics, and Food Effect of Compound 1 in Healthy Adult Participants Objectives
  • Primary Endpoint The frequency and severity of treatment-emergent adverse events (TEAEs), including clinically significant abnormal vital signs, electrocardiograms (ECGs), laboratory test results, and physical examination findings.
  • TEAEs treatment-emergent adverse events
  • ECGs electrocardiograms
  • Plasma PK endpoints include:
  • Urine PK endpoints include:
  • Cohort A3 was a food effect cohort, and participants in Cohort A3 only returned to the CRU on Day 14 and, following a 14-day washout, received a second single dose of their assigned treatment on Day 15 following consumption of a high fat meal. For Cohort A3 only, a second period of confinement applied from Day 14 until completion of the 72-hour post-dose assessments on Day 18.
  • Part A participants returned to the CRU for a follow-up visit 7 days ( ⁇ 1 day) after their final dose of study drug.
  • SAD the decision to escalate a dose or modify a dose was determined by the SMC following review of the 7-day blinded safety and available PK data from the preceding cohort.
  • Compound 1 dose will not exceed 1200 mg or a dose level that leads to mean C max >800 ng/mL or mean AUCo-iast > 4000 ng*h mL.
  • Compound 1 dose will not exceed 1000 mg per day or a dose level that leads to mean C max >800 ng/mL or mean AUCo-iast > 4000 ng*h mL.
  • Part A up to 48 healthy volunteers; Part B: up to 32 healthy volunteers).
  • Participants must have a minimum body weight of 50 kg and a body mass index (BMI) between >18.0 and ⁇ 32.0 kg/m 2 at screening.
  • BMI body mass index
  • Participants must have clinical laboratory values within normal range as specified by the testing laboratory, unless deemed not clinically significant by the investigator or delegate.
  • Double contraception is defined as a condom and one other form of the following:
  • FSH follicle-stimulating hormone
  • Periodic abstinence e.g., calendar, ovulation, symptothermal, post-ovulation- methods
  • withdrawal are not considered highly effective methods of birth control. Participant complete abstinence for the duration of the study and for 1 months after last study treatment is acceptable.
  • WOCBP must have negative pregnancy test at screening and Day 1 and be willing to have additional pregnancy test as required.
  • GI gastrointestinal
  • atrophic gastritis atrophic gastritis
  • lactose intolerance atrophic gastritis
  • H. Helicobacter
  • Fever body temperature > 38 °C
  • symptomatic viral or bacterial infection within 2 weeks prior to screening.
  • HCV hepatitis C antibody
  • HBV hepatitis B surface antigen
  • HCV human immunodeficiency virus
  • Participants with a positive toxicology screening panel urine test including qualitative identification of barbiturates, tetrahydrocannabinol (THC), amphetamines, benzodiazepines, opiates, and ***e
  • Alcohol breath test Participants with a history of substance abuse or dependency or history of recreational intravenous (IV) drug use over the last 5 years (by self-declaration).
  • Participant is unwilling to abstain from alcohol beginning 48 hours prior to admission to the CRU and during confinement period.
  • Use of any prescription drugs other than hormonal contraception: OCPs, long-acting implantable hormones, injectable hormones, a vaginal ring or an HID), over-the- counter (OTC) medication, herbal remedies, supplements or vitamins within 1 week prior to dosing and during course of study without prior approval of the investigator and medical monitor.
  • Simple analgesia nonsteroidal anti-inflammatory drug (NSAID)
  • paracetamol may be permitted at discretion of investigator.
  • Compound 1 was provided as a powder in capsule formulation for oral administration.
  • the formulation was a Size 00 Swedish orange capsule containing drug substance (10 mg, 50 mg, or 250 mg) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose (HPMC)), iron oxide as coloring agent, and titanium dioxide as an opacifier.
  • HPMC hydroxypropylmethyl cellulose
  • the drug product was stored at room temperature (15 °C - 25 °C).
  • Compound 1 is in a pharmacological class of tyrosine kinase inhibitors (TKI).
  • TKI tyrosine kinase inhibitors
  • Compound 1 is an orally bioavailable small molecule dual kinase inhibitor of platelet-derived growth factor receptors (PDGFR) and vascular endothelial growth factor receptors (VEGFR2). Duration of Treatment
  • Part A In Part A (SAD), participants randomized to active treatment in cohorts Al, A2, A4, A5, and A6 received a single oral dose of Compound 1 administered once on Day 1 only. Participants randomized to active treatment in Cohort A3 (Food Effect) received a single oral dose of Compound 1 on Days 1 and 15.
  • PartB In Part B (MAD), participants randomized to active treatment received oral Compound 1 administered twice daily for 7 consecutive days (Day 1 to Day 7) or once daily for 14 consecutive days (Day 1 to Day 14), to be determined on completion of Part A (SAD) of the study.
  • the placebo capsules were visually matching to active study drug and were composed of the same capsule shell but were filled with silicified microcrystalline cellulose.
  • Silicified microcrystalline cellulose is a pharmaceutical excipient composed of co-process microcrystalline cellulose and colloidal silicon dioxide.
  • Safety The safety and tolerability of single and repeat-doses of Compound 1 was investigated according to the following specific assessments: vital signs (systolic and diastolic blood pressure, pulse rate, body temperature, and respiratory rate), 12-lead ECG, clinical laboratory tests (hematology, biochemistry, coagulation, and urinalysis), physical examination, and assessment of TEAEs.
  • ITT Intent-to-Treat
  • Safety Population All participants who received any amount of study drug (Compound 1 or placebo) were included in the Safety population. The Safety population was used for the summaries of all safety assessments. Participants were analyzed according to treatment received.
  • Pharmacokinetic Population All participants who received any amount of active study drug (Compound 1) and have sufficiently evaluable concentration-time profile to allow determination of at least one PK parameter were included in the PK population. An evaluable PK profile was determined at the discretion of the pharmacokineticist following examination of subjects with dosing or protocol deviations that could potentially affect the PK profile. The PK population was used for the summaries of all PK data.
  • Concomitant medications were coded using the World Health Organization (WHO) drug dictionary Version B3 September 2018 Drug Global. Concomitant medications were listed by participant and summarized by anatomical therapeutic class and preferred name.
  • WHO World Health Organization
  • Pharmacokinetics Plasma Compound 1 concentrations, actual blood sampling times, and PK parameters were listed by treatment and protocol specified time point and summarized using descriptive statistics for PK data as outlined above for each scheduled time point by treatment arm. Individual and mean Compound 1 concentration-time profiles were also presented graphically for each treatment. Pharmacokinetic parameters were computed from the individual plasma Compound 1 concentrations using a non-compartmental approach.
  • the following plasma Compound 1 non-compartmental PK parameters were estimated, as appropriate: Cmax, Tmax, AUCo-iast, AUCo-inf, AUCtau (calculated for MAD cohorts only), Kei, ti/2, CL/F (SAD cohorts), CL/Fss (MAD cohorts), Vz/F (SAD cohorts, Vz/Fss (MAD cohorts), RA (calculated for MAD cohorts only).
  • Urine collection time, volume collected, and Compound 1 concentration (Aeu-a) duration of each sampling interval was listed for each participant and summarized by nominal sampling time point and treatment using descriptive statistics (Number of subjects [N], arithmetic mean, SD, CV%, geometric mean, median, minimum and maximum). Individual and mean Compound 1 cumulative urinary excretion-time profiles for each treatment were also presented graphically. Where urine was collected for PK analysis, the following parameters are calculated, as appropriate: Aeu-t2, CLR, CL/F, and Feu-t2.
  • Standard extrapolation for administration to humans would predict that a mean Cmax > 800 ng/mL or mean AUCo-iast > 4000 ng*h/mL would be observed at a human dose of approximately 24 mg/kg/day (i.e., approximately 1440 mg/day for a 60 kg human). Yet, during the clinical study described above, Cmax and AUCo-iast approached their limits at doses approx. 2.4-fold lower than predicted using standard techniques, and Compound 1 exposure in humans was approximately 6-8 times greater than in monkeys. For example, as described below, subjects given a 600 mg dose (i.e., approx.
  • Example 24 describes a protocol for a Phase I study with different cohorts based on certain preliminary PK findings.
  • FIG. 1 summarizes mean oral blood PK profile over time from SAD cohorts in a fasted state.
  • Table 4 summarizes mean oral blood PK parameters from SAD cohorts in a fasted state as of a first data cutoff.
  • Compound 1 was rapidly absorbed with a median Tmax of 1-2 h post-dose. With 400 mg and 600 mg doses, a decline in plasma concentration was observed with mean ti/2 of approx. 10-30 h. Moderate inter-subject variability was observed in 400 mg and 600 mg doses, with approx. 28-44% variability in Cmax and AUC. Cmax increased dose-proportionally up to 400 mg and slightly over-proportionally from 400 mg to 600 mg.
  • Plasma AUC increased 1.6-fold with a 1.5-fold increase in dose (i.e., from 400 mg to 600 mg).
  • Blood Cmax in human healthy volunteers given a 600 mg dose was greater than Cmax of maximum tolerated dose in a monkey 7-day QD study, where blood Cmax in monkey was 605 ng/mL (male) and 780 ng/mL (female) and blood AUC in monkey was 5720 ng*h/mL (male) and 6450 ng*h/mL (female).
  • FIG. 2 and Table 5 summarize results of food effect cohorts (3 A and 3B) as of a first data cutoff.
  • FIG. 2 shows mean oral plasma PK profiles over time in SAD food effect cohorts receiving 200 mg Compound 1. A negative food effect was observed, with 3.6-fold lower Cmax when Compound 1 was administered with food, yet little change in AUCo-iast was observed.
  • Compound 1 pharmacokinetics were characterized following single and multiple oral doses to male Sprague Dawley rats and to male and female cynomolgus monkeys in pharmacokinetic and toxicology studies. Single intravenous doses were administered to rats and monkeys for the purpose of estimating bioavailability of oral doses of Compound 1. Table 6 summarizes pharmacokinetics of Compound 1 observed in rats and monkeys following single intravenous or oral doses.
  • Exposure as assessed by AUCo-iast increased with increasing dose; however, the increase was greater than dose proportional in rats and nearly dose proportional in the monkey.
  • the oral bioavailability of Compound 1 across a dose range of 10 to 100 mg/kg ranged from 4 to 10% in both species.
  • Compound 1 When administered intravenously, Compound 1 was highly distributed outside of the systemic circulation and its volume of distribution values of 2.3 to 18.2 L/kg exceeded total body water in rats and monkeys.
  • Example 4 Compound 1 Inhibits PDGFRjl Phosphorylation in Hepatic Stellate Cells
  • Human Hepatic Stellate Cells (ScienCell Research Labs, Carlsbad, CA, Cat# 5300) were grown in Stellate Cell Medium (ScienCell Research Labs, Carlsbad, CA) in a 37 °C incubator with an atmosphere of 5% C02/95% air. Cells were maintained in tissue culture flasks, plated in 6-well tissue culture plates (at 1 million cells per well), allowed to adhere and grown to 80% confluence. Cells were starved overnight in Stellate Cell Medium without serum. From a 10 mM stock solution of Compound 1 in DMSO, 10-fold serial compound dilutions were prepared. Diluted Compound 1 solutions in DMSO were added at final concentrations of 10 mM, 1 pM,
  • PDGF-BB R&D Systems Catalog # 220-BB
  • lysis buffer RIPA lysis buffer, Sigma R0278
  • the lysis buffer also contained Phosphatase Inhibitor Cocktail 1 (Sigma P2850) and Phosphatase Inhibitor Cocktail 2 Sigma P5726).
  • Example 5 Compound 1 Inhibits KDR Phosphorylation in HUVEC Cells
  • HUVEC Human Umbilical Vein Endothelial Cells (HUVEC) (ATCC Catalog # CRL1730) cells were grown in EBM-2 (Endothelial Cell Growth Basal Medium-2, Lonza Catalog # 00190860) supplemented with EGM-2 Endothelial SingleQuots Kit (Lonza Catalog # CC-4176) in a 37°C incubator with an atmosphere of 5% C02/95% air. Cells were maintained in tissue culture flasks, plated in 6-well tissue culture plates (at 1 million cells per well), allowed to adhere and grown to 80% confluence. Cells were starved overnight in EBM-2 without serum. From a 10 mM stock solution of Compound 1 in DMSO, 10-fold serial compound dilutions were prepared. Diluted Compound 1 solutions in DMSO were added at final concentrations of 10 mM, 1 pM,
  • VEGF165 R&D Systems Catalog # 293-VE
  • lysis buffer RIPA lysis buffer, Sigma R0278.
  • the lysis buffer also contained Phosphatase Inhibitor Cocktail 1 (Sigma P2850) and Phosphatase Inhibitor Cocktail 2 (Sigma P5726).
  • Protein concentrations in the lysates were determined using the DC protein determination kit (Biorad 500-0113 and 500-0114) and 25 pg protein was loaded for Western analysis on 10 lane gels (Invitrogen Catalog # NP0321). In a control lane, MagicMarkTM XP Western Protein Standard was loaded (Invitrogen Catalog # LC5602). Proteins were transferred to Nitrocellulose membranes (Invitrogen Catalog # 77010).
  • Blots were blocked with non-fat dry-milk (Biorad Catalog # 70-6404) and incubated overnight at 2-8 °C with primary antibodies against phospho-VEGF Receptor 2 (Tyrl 175) (Cell Signaling Catalog # 2478) or GAPDH (Cell Signaling # 2118) at a 1 : 1000 dilution. Blots were washed several times and then incubated with an anti-rabbit IgG, HRP-linked secondary antibody at a 1:1000 fold dilution (Cell Signaling Catalog # 7074).
  • Example 6 Compound 1 Decreases Expression of Certain Fibrosis and Inflammation Related Markers
  • Compound 1 is in a pharmacological class of tyrosine kinase inhibitors (TKI) and has a formula of: (Z)-methyl 3-(((4-(N-methyl-2-(4-methylpiperazin-l-yl)acetamido)phenyl)amino) (phenyl)methylene)-2-oxo-2,3-dihydro-lH-pyrrolo[2,3-b]pyridine-6-carboxylate as yellow solid.
  • TKI tyrosine kinase inhibitors
  • the present example describes Bio-Map studies of Compound 1 in cell culture systems using DiscoverX technology platform and that were profiled in a KINOMEscan (Ambit/Di scoveRx/Eurofms, CA) biochemical assay.
  • Exemplary culture systems analyzed were: (1) Small airway epithelial cells + lung fibroblasts (SAEMyoF); (2) Lung fibroblasts only (MyoF); and (3) Renal proximal tubular epithelial cells + lung fibroblasts (REMyoF). When activated, these cell systems mimic pathological conditions such as chronic inflammation, fibrosis, and matrix remodeling.
  • Co-cultures of epithelial cells and fibroblasts or fibroblasts alone were activated with TGFpi (10 ng/mL) + TNF alpha (lOng/mL) and treated with Compound 1 at 100 nM, 500 nM, 2.5mM and 13mM concentrations and incubated for 48 hours. Expression of numerous critical anti-inflammatory and anti-fibrotic marker proteins were evaluated.
  • Compound 1 treatment decreased levels of several inflammation-related markers including monocyte chemotactic protein (MCP-1), macrophage colony stimulating factor (M-CSF), soluble interleukin 8 (sIL-8), Interferon induced T cell alpha chemoattractant (I-TAC), IL-8, Interferon gamma induced Protein- 10 (IP- 10).
  • MCP-1 monocyte chemotactic protein
  • M-CSF macrophage colony stimulating factor
  • sIL-8 soluble interleukin 8
  • I-TAC Interferon induced T cell alpha chemoattractant
  • IL-8 Interferon gamma induced Protein- 10
  • Compound 1 treatment also decreased fibrosis-related markers including N-cadherin, a-SMA, Collagen I and III, TIMP-1, plasminogen activator inhibitor (PAI-1), matrix metalloprotease, MMP-1, MMP-9, tissue and urokinase plasminogen activator (tPA and uPA), soluble VEGF and epidermal growth factor (EGFR) as shown in FIG.
  • fibrosis-related markers including N-cadherin, a-SMA, Collagen I and III, TIMP-1, plasminogen activator inhibitor (PAI-1), matrix metalloprotease, MMP-1, MMP-9, tissue and urokinase plasminogen activator (tPA and uPA), soluble VEGF and epidermal growth factor (EGFR) as shown in FIG.
  • the present example demonstrates that Compound 1 induced dose-dependent decrease in expression of a number of different inflammation and fibrosis-related markers.
  • the present example demonstrates that a mixed cell culture comprising renal epithelial cells and activated myofibroblasts treated with Compound 1 (13 mM) reduced expression of COL1 and COL3.
  • Example 7 TGFpi-Induced Collagen Production in Normal Human Lung Fibroblasts
  • NHLF Normal Human Lung Fibroblasts
  • FGBM Fibroblast Growth Basal Medium
  • FGM2 Fibroblast Growth Medium-2 SingleQuots Supplements and Growth Factors
  • FGM2 Fibroblast Growth Medium-2 SingleQuots Supplements and Growth Factors
  • TGFpi R&D Systems; Catalog 240-B
  • Treatments were performed in duplicate wells and cells that were not treated with compound and/or TGFpi were used as controls.
  • the percentage inhibition was calculated using cells treated with TGFpi as 0% inhibition and cells not treated with compound and TGFpi as 100% inhibition.
  • collagen production was inhibited by 104% and 86%, respectively.
  • the inhibition of collagen production at both Compound 1 concentrations was statistically significant (p ⁇ 0.01).
  • Example 8 Compound 1 Ameliorates Fibrosis in Bleomycin-induced Injury Model of Idiopathic Pulmonary Fibrosis
  • mice Male C57BL/6 mice ( ⁇ 20 to 25 g) were anaesthetized for a short period of time using ketamine (100 mg/kg) and xylazine (10 mg/kg).
  • mice Two weeks after bleomycin instillation, a subgroup of animals was sacrificed and the establishment of lung fibrosis was confirmed (initially as determined by increased lung weight).
  • FIG. 7A is a graph showing lung to body weight ratio (mg/g).
  • Compound 1 significantly reduced hydroxyproline, a major component and marker of collagen deposition and fibrosis.
  • Two weeks after bleomycin exposure animals treated with bleomycin had increased lung hydroxyproline levels compared to sham-treated animals, which indicates development of pulmonary fibrosis. Animals then treated with Compound 1 for three weeks had reduced hydroxyproline levels compared to vehicle-treated animals (FIG. 7B).
  • Compound 1 also significantly reduced fibrosis score (Ashcroft) on histopathology. Lung tissue sections were stained with H&E, and slides were scored for histopathological damage using the Ashcroft scale. Two weeks after bleomycin exposure, animals treated with bleomycin had increased histopathological damage compared to sham-treated animals, indicative of development of pulmonary fibrosis. Animals treated with Compound 1 for three weeks had reduced histopathological damage compared to vehicle-treated animals (FIG. 7C).
  • TGFpi transforming growth factor beta
  • Example 9 Compound 1 Reduces Fibrosis in Inducible TGFpi Mouse Model of Lung Fibrosis
  • mice which expresses TGFpi in the lung via an externally regulatable, triple transgenic system using a doxycycline-inducible promoter.
  • a doxycycline-inducible promoter See Lee, C. G., et al. Proc. Am. Thorac. Soc. 2006 Jul;3(5):418-23; Lee C. G., et al. J. Exp. Med. 2004 Aug 2;200(3)377-89.
  • Eight-to-ten week old TGFpi positive female mice (approximately 25 g of body weight) were induced (fed) with doxycycline (dox) (0.5 mg/ml dox and 20 g/L sucrose in drinking water) for 4 weeks.
  • dox doxycycline
  • the TGFpi transgene was under the control of a dox-inducible promoter.
  • age and gender matched TGFpi negative mice from the transgenic mouse breeding colony not fed with dox were included as control mice (sham).
  • Marked loss of normal bronchial tree architecture due to TGFpi overexpression was evidenced after 4 weeks of dox compared to sham mice using micro-CT images.
  • lung hydroxyproline (HYP) for collagen content, picrosirius red (PSR) staining for collagen signal, and the histopathological observations from (H&E) slides by Ashcroft scoring and IHC staining for alpha SMA were performed. All histological quantitations were performed using a Bioquant image analysis program.
  • Compound 1 reduced lung fibrosis score (Ashcroft) in dox-fed TGFpi transgenic mice.
  • Dox feeding of TGFpi transgenic mice for 4 weeks resulted in a significant increase in lung fibrotic score (Ashcroft score based on H&E staining), compared to normal mice (FIG. 8A).
  • Further dox feeding for an additional 4 weeks resulted in more increases in lung fibrotic score, as shown by the vehicle group.
  • a reduction in hydroxyproline, a component of collagen is indicative of a reduction in fibrosis.
  • Compound 1 significantly decreases aSMA, an early lung fibrosis marker.
  • aSMA is a marker of myofibroblast activation, an early step in fibrosis development.
  • Example 10 Compound 1 Has Anti-Fibrotic Effects Across Multiple Organ Systems in Bleomycin Systemic Sclerosis Mouse Model
  • SC subcutaneous
  • the right lung and right kidney were also fixed in formalin for histopathological evaluation.
  • Dermal thickness measurements were made from H&E-stained tissue sections by measuring the distance from the epidermis to the dermal junction using Bioquant planimetric software.
  • Lung and kidney hydroxyproline (HYP) assays were performed to determine tissue collagen content. Skin, lung and kidney histopathological fibrotic scores were determined by two independent observers, and the scores were averaged.
  • Picrosirius red (PSR) staining of kidney sections was also performed to determine renal collagen deposition. A subset of animals were randomly chosen from each group for histological staining. All histological quantitations were performed using Bioquant image analysis software.
  • Scleroderma is characterized by thickening of the skin caused by accumulation of collagen.
  • dermal thickness was determined by measuring the distance between the epidermal-dermal junction and the dermal-subcutaneous fat junction. This was done in H&E-stained slides prepared from a skin biopsy. After 4 weeks of subcutaneous bleomycin injections (Bleo Pre-Rx Group), dermal thickness was significantly increased compared to sham-injected animals (Sham Group). This indicates significant skin thickening and scleroderma at the time of initiation of compound treatment.
  • Therapeutic treatment of Compound 1 for 5 weeks 25 mg/kg, PO, BID) (Bleo Compound 1 Group) significantly decreased dermal thickness compared to the bleomycin- exposed vehicle cohort (Bleo Vehicle Group).
  • FIG. 9A summarizes these results.
  • Hydroxyproline is a major component of collagen, and it is used to indicate tissue collagen deposition and the development of fibrosis.
  • pulmonary hydroxyproline levels were determined. After 4 weeks of subcutaneous bleomycin injections, lung hydroxyproline levels were significantly increased compared to sham treated animal (Sham vs Bleo PreRx Groups), indicating the development of pulmonary fibrosis.
  • Treatment of bleomycin exposed animals with Compound 1 for 5 weeks (25 mg/kg, PO, BID) (Bleo Compound 1 Group) significantly decreased lung hydroxyproline levels compared to bleomycin exposed vehicle treated animals (Bleo Vehicle), indicating anti -fibrotic activity of Compound 1.
  • FIG. 9C summarizes these results.
  • Urine was again collected (for 24 hours) from day 10-11 after first PAN injection (referred to as Day 11 urines).
  • Day 11 urines On day 11 after first PAN administration GFR was measured by computing FITC-sinistrin decay (Medibeacon) and animals were sacrificed. The left kidney was cut coronally, immersed in formalin (4%) and submitted for sectioning and mounting on slides for subsequent analysis (PAS stain for determining glomerular diameter) and anti-COL 3 antibody IHC. All microscopic analyses (40X) were conducted by an observer blinded to the treatment groups. Glomerular diameter was determined from digital microscopic pictures using ImageJ image analysis software to perform planimetry. Proteinuria was determined from urine samples.
  • every endpoint was not measurable in each animal within a group due to logistical limitations. In such cases, randomly selected animals were queried for data. For instance, no urine was collected for certain animals due to the limited number of metabolic cages available, and only a few animals in each cohort were randomly selected for GFR measurement due to the limited number of probes available for this measurement. The missing animals are marked as NA (not available) in the relevant data tables.
  • the highly sensitive FITC-sinistrin clearance method was employed in conscious animals. This method is capable of measuring the elimination kinetics of FITC-sinistrin using an optical device (Medibeacon, Germany) that can monitor fluorescence trans-cutaneously.
  • FITC-sinistrin clearance is a readout of glomerular filtration rate.
  • the optical device measures fluorescence emitted by FITC-sinistrin through the skin and was temporarily secured on the back of the animal.
  • a single dose of FITC-sinistrin (5 mg/100 g body weight FITC-sinistrin) was injected into the tail vein and the measurement with the optical device was performed for 1.5 hr. The data were downloaded and analyzed using the MPD lab software (Medibeacom, Germany).
  • Glomerular Filtration Rate determination by the FITC- sinistrin method was determined for three animals per experimental group.
  • Glomerular filtration rate was determined immediately prior to terminal sacrifice using FITC-sinistrin decay kinetics as a measurement of renal clearance.
  • GFR Glomerular filtration rate
  • FIG. IOC in Sham (Control) animals, administration of FITC-sinistrin results in gradual decline in fluorescence, indicative of normal renal clearance.
  • animals treated with PAN and Vehicle a significant decrease in the clearance of FITC-sinistrin was observed, showing marked renal dysfunction in the PAN Vehicle animals. Animals treated with PAN and Compound 1 had markedly enhanced clearance of FITC-sinistrin compared to PAN Vehicle animals.
  • Collagen 3 (COL-3) expression a measure of scar, was determined in glomeruli by quantitative immuno-histochemistry.
  • the COL-3 expression level was markedly elevated in glomeruli in the kidneys from PAN Vehicle animals compared to Sham treated animals (FIG. 10E).
  • the COL-3 level was significantly reduced with Compound 1 treatment (FIG. 10E) compared to PAN Vehicle treatment.
  • Example 12 Compound 1 Reduces Proteinuria and Fibrosis in DOCA/Salt Model of Renal Injury and Fibrosis
  • Induction of renal dysfunction was started on day 1, at which point experimental animals received a first subcutaneous dose of deoxycorticosterone acetate (DOCA) at 30 mg/kg and animals were switched from normal drinking water (tap water) to drinking water with 1% NaCl. Animals were kept on the 1% NaCl drinking water for the remainder of the experiment. Animals received weekly subcutaneous doses of DOCA; 30 mg/kg in weeks 1, 2, 3 and 4 and 15 mg/kg in weeks 5 and 6. Control groups were subcutaneously injected with PEG400 and received regular tap water ad libitum. After two weeks, overnight urine was collected in metabolic cages and a subset of animals was sacrificed to establish baseline values.
  • DOCA deoxycorticosterone acetate
  • Animals were monitored several times daily. Three animals in the DOCA + Salt + Vehicle group (Group 3) had unscheduled sacrifice (one each on days 29, 34, and 35). Body weight, serum and kidneys were collected for those animals and samples were processed along with the samples from other animals in the group.
  • kidneys were collected and their weights recorded. At terminal sacrifice, the kidney weights from animals of DOCA + Vehicle group (Group 4) were significantly more than the kidney weights in Sham animals (Group 3). The kidney weight in DOCA + Compound 1 group (Group 5) was not significantly different from the DOCA + Vehicle group (Group 4).
  • the weight of the kidney was also expressed as a percentage of the total body weight of each individual animal.
  • the % kidney weight from animals of DOCA + Vehicle group (Group 4) was significantly higher than the % kidney weights in Sham animals (Group 3).
  • Compound 1 increased body weight and decreased kidney weight compared to vehicle, albeit not statistically significant.
  • the % kidney weight (taken both body weight and kidney weight into consideration) in DOCA + Compound 1 group (Group 5) was significantly reduced compared to the DOCA + Vehicle group (Group 4).
  • Urines were analyzed for protein content and the total amount of protein produced over 24 hours was calculated from the protein concentration and the urine volume produced in 24 hours (FIG. 11 A).
  • a statistically significant elevation in proteinuria was observed in DOCA + Salt animals (Group 2) compared to Sham animals (Group 1), indicating renal dysfunction in the DOCA + Salt animals at the start of compound or vehicle treatment.
  • a statistically significant elevation in proteinuria was again observed in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), indicative of continued renal dysfunction.
  • DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in proteinuria compared to DOCA + Vehicle treated animals (Group 4). This is consistent with an improvement in renal function by Compound 1 treatment.
  • Urines were analyzed for albumin concentration and the total amount of albumin produced over 24 hours was calculated from the albumin concentration and the urine volume produced in 24 hours (FIG. 1 IB).
  • elevated albuminuria was observed in DOCA + Salt animals (Group 2) compared to Sham animals (Group 1), albeit that this effect was not statistically significant.
  • a statistically significant elevation in albuminuria was observed in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), indicative of renal dysfunction.
  • DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in albuminuria compared to DOCA + Vehicle treated animals (Group 4). This is consistent with the proteinuria data and with an improvement in renal function by Compound 1 treatment.
  • Urines were also analyzed for creatinine levels and the albumin-to-creatinine-ratio (ACR) was calculated (FIG. 11C). Data for ACR were similar to the overall albumin production data. As before, DOCA-exposed animals treated with Compound 1 (Group 5), were found to have statistically significant reduction in ACR compared to DOCA + Vehicle treated animals (Group 4). This is consistent with the proteinuria data and the albuminuria data, and with an improvement in renal function by Compound 1 treatment.
  • Kidney Injury Molecule- 1 (KIM1) (also known as Hepatitis A virus cellular receptor
  • HAVcr-1 T-cell immunoglobulin and mucin domain 1
  • TIM1 T-cell immunoglobulin and mucin domain 1
  • Hydroxyproline is a major component of collagen, and tissue hydroxyproline levels are considered a good indicator of tissue collagen content and therefore of tissue fibrosis.
  • the hydroxyproline content of renal samples was determined and expressed as the total hydroxyproline content per kidney (FIG. 1 IE).
  • a statistically significant elevation in renal hydroxyproline content was seen in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), consistent with marked renal fibrosis at study end.
  • DOCA-exposed animals treated with Compound 1 were found to have statistically significant reduction in renal hydroxyproline content compared to DOCA + Vehicle treated animals (Group 4). This indicates that Compound 1 treatment prevents the development of renal fibrosis in DOCA + Salt treated animals.
  • Picrosirius red-staining in the slides was quantified using Bioquant Image Analysis software. The data was expressed as arbitrary units of staining intensity and also expressed as a % of control in relation to the average staining intensity of the control group (Group 1). As shown in FIG. 11G, in pre-randomization animals, no increase in picrosirius red staining intensity was observed in DOCA + Salt animals (Group 2) compared to Sham control animals (Group 1). This indicates lack of renal fibrosis at the time of compound of vehicle treatment initiation. This lack of histologically observable fibrosis in the renal samples from Group 2 is consistent with the hydroxyproline data in this group.
  • DOCA + Vehicle treated animals (Group 4) had markedly increased insterstitial picrosirius red staining compared to Sham control animals (Group 3). This indicates marked collagen deposition and renal fibrosis (tubulointerstitial fibrosis) as a result of DOCA + Salt treatment.
  • animals that received DOCA + Salt that were treated with Compound 1 (Group 5) showed statistically significantly reduced picrosirius red staining intensity compared to DOCA + Vehicle animals (Group 4). This is consistent with the observed differences in renal hydroxyproline in the various experimental groups and shows anti-fibrotic effects of Compound 1 in this model.
  • the weight of the obstructed kidney was also expressed as a percentage of the body weight of each individual animal.
  • the percentage kidney weight for animals from the UUO + Vehicle group (Group 2) was elevated compared to Control animals (Group 1). There was no statistically significant difference between the UUO + Compound 1 group (Group 3) and the other experimental groups.
  • kidneys were fixed in 10% neutral buffered formalin for at least 48 hours before preparation for histology. Kidneys tissues were grossed, processed, dehydrated, embedded in paraffin and cut into 5 pm sections.
  • Compound 1 was evaluated in the Unilateral Ureteral Obstruction (UUO) model of renal fibrosis in the mouse.
  • Primary endpoints for evaluating compound effects were renal damage score, collagen deposition (as determined by picrosirius red staining) and formation of myofibroblasts (as determined by a-smooth muscle actin staining).
  • Compound 1 had statistically significant beneficial effects on renal damage, picrosirius red staining and alpha-SMA staining.
  • the PCK rat which has a mutation in PKHD1 (ARPKD gene/ phenotypically resembles both human ARPKD-CHF and human PKD in that it exhibits large renal cysts, and renal and hepatic scarring.
  • the PCK rat model is a well-established and well-characterized model that resembles human polycystic kidney and liver disease (Lager, D. L, et al. Kidney Int. 2001 Jan;59(l): 126-36). Results described in this Example are also described in Paka, P., et al. World J. Nephrol. 2018 Sept 8;7(5):96-107).
  • PCK rats that were treated with either vehicle or with Compound 1 did not differ in their body weights from age and sex matched wild-type (WT) Sprague Dawley rats.
  • SCr Serum creatinine
  • BUN Serum creatinine
  • WT wild-type rats
  • Kidneys were weighed at sacrifice and kidney weights are given in absolute weight (g) and as a percentage of body weight (FIG. 13C and FIG. 13D, respectively). Kidneys from PCK Vehicle rats were markedly larger than kidneys from wild-type (WT) rats. Compound 1 treatment in PCK rats reduced kidney size and the % kidney weight compared to PCK Vehicle animals.
  • Hydroxyproline is a major component of collagen, and it is used to indicate tissue collagen deposition and fibrosis. Hydroxyproline levels were determined in kidney samples of 14-week old animals (FIG. 13E). Kidney hydroxyproline levels were markedly increased in PCK Vehicle rats compared to wild-type (WT) rats. Compound 1 treatment significantly reduced renal hydroxyproline levels compared to PCK Vehicle animals, indicating reduced renal collagen deposition as a result of Compound 1 treatment.
  • Increased urine protein excretion can be due to diseases of the glomerulus, an increased quantity of proteins in serum (overflow proteinuria) or low reabsorption at proximal tubule.
  • Urine protein excretion was determined in urines collected over 24 hours immediately prior to sacrifice (FIG. 13H). Protein excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced proteinuria in PCK rats compared to PCK Vehicle animals.
  • Albumin is a major plasma protein. Usually, only trace amounts of it are present in urine, whereas larger amounts occur in the urine of patients with kidney disease. Urine albumin concentrations were determined in urines collected over 24 hours immediately prior to sacrifice (FIG. 131). Urine albumin production was found to be strongly elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced urine albumin production in PCK rats compared to Vehicle treated PCK animals.
  • Lipocalin-2 (LCN2) is also known as neutrophil gelatinase-associated lipocalin (NGAL) and is a protein that is expressed in neutrophils and at low levels in normal kidneys (Cowland, J. B., et al. Genomics 1997 Oct 1;45(1): 17-23).
  • NGAL neutrophil gelatinase-associated lipocalin
  • Urine NGAL excretion was determined for 14-week old animals (FIG. 13J).
  • NGAL excretion was found to be markedly elevated in PCK Vehicle rats compared to wild-type (WT) rats, albeit that the increase was not statistically significant by one way ANOVA followed by Tukey’s test. This is probably the result of high variability in NGAL excretion amongst the PCK Vehicle animals.
  • Compound 1 treatment reduced NGAL excretion in PCK rats compared to Vehicle treated animals, albeit that this decrease was also not statistically significant when tested by ANOVA followed by Tukey’s test.
  • HAVcr-1 also known as KIM-1 (KIM1, kidney injury molecule 1) is highly upregulated in injured kidneys in response to various types of renal insults (Bonventre, J. V. Trans. Am. Clin.
  • KIM-1 excretion was found to be strongly elevated in PCK Vehicle rats compared to wild-type (WT) rats, indicating severe renal injury, in particular tubular injury (Bonventre 2014). The elevated KIM-1 excretion is consistent with tubular cyst formation and tubular damage known to exist in the PCK rat (Lager 2001).
  • Compound 1 treatment markedly reduced KIM-1 excretion in PCK rats compared to Vehicle treated animals (FIG. 13K), which indicates mitigation of renal damage, and in particular tubular damage.
  • Interleukin 18 (IL-18 or IL18) is also known as interferon-gamma inducing factor and is a pro-inflammatory cytokine.
  • Urine levels of IL-18 are associated with urine protein excretion and have been used as a biomarker for the progression of diabetic nephropathy (Zhang, D., et ak, PeerJ. 2019;7:e7079; Nakamura, A., et al. Diabetes Care 2005 Dec;28(12):2890-5). Urine IL-18 excretion was determined for 14-week old WT and PCK animals (FIG. 13L).
  • IL-18 excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced IL-18 excretion in PCK rats compared to Vehicle treated animals.
  • Cystatin C or cystatin 3 is used as a biomarker of kidney function (Dharnidharka, V. R., et al. Am. J. Kidney Dis. 2002 Aug;40(2):221-6; Roos, J. F., et al. Clin. Biochem. 2007 Mar;40(5-6):383-91).
  • Urine Cystatin C excretion was determined for 14-week old animals (FIG. 13M).
  • Cystatin C excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced Cystatin C excretion in PCK rats compared to Vehicle treated animals. These results are indicative of an improvement of renal function, consistent with the observation that Compound 1 reduced serum BUN and SCr levels in PCK rats compared to PCK Vehicle treated animals.
  • TNBS (2,4,6-trinitrobenzenesulfonic acid)/ethanol induces a severe illness characterized by bloody diarrhea and a dramatic loss of body weight during the first week with increased colon damage score (Mateus, V., et al. Clin. Exp. Gastroenterol. 2018;11:325-334).
  • TNBS-induced colitis characteristic of acute colitis and the early phase of IBD, is a commonly utilized animal model in rodents that shares significant properties with human Crohn's disease. Advantages of this model include reproducibility and technical simplicity (Filipescu, I. E., et al. PLoS One 2018 Aug 23;13(8):e0202929).
  • TNBS-induced colitis is a well-validated animal model to assess compounds with potential therapeutic effects such as anti-TNFa, corticosteroids, natural compounds and traditional medicine (Mateus 2018; Filipescu 2018; Wirtz, S. et al. Nat. Protoc. 2017 Jul; 12(7): 1295-1309). Compound 1 was therefore tested in the well-established model of TNBS-induced IBD/ Acute Colitis.
  • 5 TNBS-treated mice, as well as 2 sham mice were sacrificed to demonstrate significant TNBS-induced colonic inflammation and pathology, including decreased body weight, reduced colon length and colon gross morphological macroscopic damage.
  • mice were randomized to vehicle and Compound 1 at 5, 15 and 45 mg/kg Compound 1 Hydrochloride trihydrate, PO, BID for 4 days. After 4 days of Compound 1 or vehicle treatment, mice were sacrificed. Body weight, colon length and colon weight were recorded. The middle and distal part of colon were fixed in 10% neutral buffered formalin.
  • Colon histopathology was determined from H&E-stained tissue sections by two independent observers using a previously published scoring system.
  • the score for each individual component (0-3) of the Histopathological Score is shown in FIG. 14E and the composite (total) Colon Histopathological Score (the sum of individual component scores; 0-12) is shown in FIG. 14F. Histopathological Scores showed that Compound 1 treatment at 15 and 45 mg/kg markedly improved colon histology compared to pre-treatment and vehicle treated animals.
  • Colon tissue sections were stained with Alcian blue to evaluate crypt architecture/goblet cells. The degree of Alcian blue staining was quantified to determine crypt damage and goblet cell loss (FIG. 14G, group averages shown). Compound 1 treatment at 15 and 45 mg/kg significantly reduced TNBS-induced goblet cell loss compared to pre-treatment and vehicle treated animals.
  • MPO Myeloperoxidase
  • Example 16 Compound 1 Decreases Acetic Acid (AA) Induced Colitis in Mice
  • Male CD-I mice were infused with 4% AA in saline in 150 m ⁇ volume through rectal enema under isoflurane anesthesia.
  • AA or sham
  • Compound 1 treatment significantly mitigated the decrease in colon length (FIG. 15E), gross morphological colon damage (FIG. 15F) and histopathological colon damage (FIG. 15G) by decreasing mucosal wall thickness, and preserving and crypt/villi architecture compared to vehicle cohort (FIG. 15H).
  • Example 17 Compound 1 Decreases Chronic Colitis in IBD Mouse Model [0468] Efficacy of Compound 1 in a dextran sulfate sodium (DSS) induced chronic colitis/IBD model was evaluated.
  • DSS dextran sulfate sodium
  • Ratio Efficacy of Compound 1 in a dextran sulfate sodium (DSS) induced chronic colitis/IBD model was evaluated.
  • Adult male CD-I mice were fed with 3% DSS in drinking water for 5 days with alternate cycles of normal drinking water for 5 days and continued a total of 3 DSS cycles in 4 weeks. This model has been established to induce chronic colitis/IBD.
  • a small group (n 6) with normal drinking water was included as a sham control. Gross morphological and histopath
  • DSS-Vehicle cohort had the shortest colon length; DSS-Compound 1 cohort had significantly increased colon length compared to the DSS-vehicle cohort (FIG. 16A).
  • Compound 1 treatment decreased macroscopic colon damage score in terms of appearance, inflammation and rectal bleeding (FIG. 16B), decreased colonic hydroxyproline (FIG. 16C) and histopathological injury score based on crypt damage, epithelial erosion and inflammatory cell infiltration as seen in H&E stained colon sections (FIG. 16D). These results indicate that Compound 1 could prevent progression of inflammation-induced fibrosis.
  • PCK rat which has a mutation in PKHD1 (ARPKD gene) phenotypically resembles both human ARPKD-CHF and human PKD in that it exhibits large renal cysts, and renal and hepatic scarring, and is recognized model for polycystic kidney disease (e.g., as described in Example 14).
  • Male PCK rats (PCK/CrljCrl-pkhdlpck/Crl; Charles River Laboratories were randomized to vehicle or test compounds (25 mg/kg Compound 1 Hydrochloride trihydrate, BID, PO) at 6.5 weeks of age following confirmation of frank disease and then sacrificed at 13.5 weeks of age.
  • Age-matched male Sprague-Dawley rats served as wild-type controls. Proteinuria was recorded daily and the results are summarized in Table 13.
  • Compound 1 demonstrated a 2.9 fold greater reduction in proteinuria than nintedanib.
  • mice Streptozotocin (STZ)-induced diabetic mice were randomized to vehicle or test compounds 12 weeks after STZ induction. Results of kidney hydroxyproline analysis are summarized in Table 15. Compound 1 demonstrated 8.3 fold greater reduction in kidney hydroxyproline than nintedanib.
  • FIG. 17C Sirius red staining (FIG. 17C) and TGFpi staining (FIG. 17D) were reduced significantly compared to the vehicle cohort and showed better efficacy even when compared to the pirfenidone (PFD) cohort (an advanced clinical compound).
  • the present disclosure encompasses a recognition that in different patients, disease driving pathways may have divergent outcomes or converge on a common disease related outcome. For example, as depicted in FIG. 18, in some instances, different individuals may have distinct genomic responses that all result in a common kidney disease endpoint, while in other instances, different individuals may have a shared genomic response that leads to different kidney disease endpoints. For example, distinct disease-driving pathways in different individuals may lead to a common outcome of a fibrotic kidney disease, such as, e.g., FSGS. As such, different patients may respond to different therapies and interventions for the same disease.
  • the present disclosure provides the insight that identifying and neutralizing the particular disease drivers of a given patient may provide an early and effective way to manage kidney disease endpoints. Specifically, the present example determined the signalosome in various models of Focal and Segmental Glomerulosclerosis (FSGS) to identify biomarkers for treatment with Compound 1.
  • FSGS Focal and Segmental Glomerulosclerosis
  • Exemplary and etiologically distinct murine models of FSGS were used to characterize the kidney transcriptome and proteome that are associated with therapeutic activity of Compound 1.
  • kidneys were harvested.
  • kidney tissue was preserved in RNAlater (Sigma Aldrich) for glomerular isolation by manual microdissection.
  • RNAlater Sigma Aldrich
  • glomeruli were isolated by magnetic bead extraction.
  • RNA was extracted from glomerular preparations.
  • RNA extraction, RNA sequencing, data normalization and filtering were then performed. Differential expression analysis was performed using DESeq2 and EdgeR (M. F Love, et ak, (December 5, 2014) Genome Biol. 15: 550; M. D.
  • Hierarchical cluster dendrograms were generated to determine the ability of overall transcriptional profiles to recapitulate treatment groups. Rat and murine genes were converted to the corresponding human orthologs using the NCBI homolog (Build 64) and Genomatix annotated ortholog databases. Signalosome of rodent models treated with Compound 1 was analyzed and compared with transcriptome of renal dysfunction in human patients to thereby identify biomarkers of patients to be treated with Compound 1.
  • PAN puromycin aminonucleoside
  • rat puromycin aminonucleoside nephropathy (PAN) model of proteinuric kidney disease exhibit proteinuria that is mitigated by treatment with Compound 1, but not with sham.
  • Animals were sacrificed on Day 21 after PAN administration.
  • Mean arterial pressure (MAP) was evaluated prior to sacrifice and was not substantially different between vehicle treated and Compound 1 treated animals (data not shown).
  • MAP Mean arterial pressure
  • Kidneys were harvested and one of the two kidneys was used for analysis of the glomerular transcriptome. The other kidneys were homogenized for determination of COL1 A1 and COL3 A1 expression.
  • Characteristics of this model include the development of severe proteinuria and glomerulosclerosis, which reflects several aspects of the pathogenesis of focal and segmental glomerulosclerosis (FSGS).
  • FSGS focal and segmental glomerulosclerosis
  • the present disclosure provides the insight that disease-driving networks in animal (e.g., rodent) models of kidney disease may prove beneficial in those human patients that share the same transcriptional elements.
  • animal e.g., rodent
  • lupus nephritis lupus nephritis
  • COL1A1 and COL3A1 have been identified as disease-associated nodes in human FSGS. See, e.g., Canadas-Garre, M., et al. J. Transl. Med. (2016) 16:292; Grgic, T, et al. Kidney Int. 2014 Dec; 86(6): 1116-1129; Schwab, K., et al. Am. J. Nephrol. 2004;24:438-447. Furthermore, increased renal COL3 Al expression is associated with increased renal dysfunction in human FSGS.
  • Compound 1 interactome network include MAP2K5, MAP3K3, MAPK7, PRKAR1A, PIK3R1, SMPD1, AXL, FYN, PLXND1, CDC42PB, VAT1, RPS6KA2, RBPMS, CAV1, BGN, NPR1, FGFR1, EFEMP2, COL6A1, HEG1, PCGF2, COL6A2, MYH11, PDGFRA, PDGFRB, and KIT.
  • the present disclosure identified biomarkers in rodent models of kidney disease and in human patients that correspond with the Compound 1 mechanism of action.
  • the present disclosure encompasses a recognition that patients with fibrotic kidney diseases who have an altered level of one or more of these biomarkers may benefit from treatment with Compound 1.
  • the present example describes treatment of fibrotic disease of the kidney in a patient.
  • a nephropathy patient with suspected glomerular disease i.e., suspected of having FSGS or MCD
  • a renal biopsy For example, mRNA is extracted from single glomeruli (see Menon et ah, JCI Insight. 2020, 5(6):el33267), and sequenced to quantify levels of certain biomarkers associated with Compound l’s mechanism of action.
  • mRNA is extracted from single glomeruli (see Menon et ah, JCI Insight. 2020, 5(6):el33267), and sequenced to quantify levels of certain biomarkers associated with Compound l’s mechanism of action.
  • COL1 A1 and COL3A1 expression are quantified. If the subject has a level of COL3A1 and/or COL1A1 that is above a threshold level, then the subject is to be treated with Compound 1.
  • An exemplary threshold level is a level that is about 20% or more above a predetermined normal range or mean or median level in a healthy patient. If a subject does not have a level of biomarker above the threshold level, then alternate treatment is determined.
  • a patient to be treated with Compound 1 the kidney disease is stabilized and/or ameliorated.
  • Example 21 Puromycin Aminonucleoside Administration in Rat is Associated with Collagen Type III Glomerulopathy
  • Urine protein was determine using the Bradford assay.
  • Renal homogenates were subjected to analysis for COL3A1 and peptidylprolyl isomerase A (housekeeping gene) mRNA.
  • qPCR was performed on a Thermofisher Quant-Studio 3 Real-Time PCR system, each sample was diluted three-fold, and qPCR reaction was performed in triplicate for all tissue samples. Renal slices were stained with periodic acid Schiff for morphometric analysis or with Collagen Type III (Col3al) Antibody (BioCompare) for immunohistochemical analysis.
  • COL3 A1 mRNA level was elevated ⁇ 4-fold in the PAN cohort (FIG. 24A). There was as significant and direct association between the fold-increase in renal COL3 A1 mRNA level and the corresponding urine protein value (FIG. 24B). Immunohistochemical analysis indicated deposition of collagen type III restricted to the glomerular mesangium.
  • the present example demonstrates that an increase in COL3A1 mRNA is directly associated with an increase in urine protein in a rat model of kidney disease.
  • collagen type III deposition is restricted to the glomerulus, which may be indicative of a robust COL3A1 transcriptomic network in that compartment, compared to, e.g., the tubulointerstitium.
  • the present disclosure encompasses a recognition that in different patients, disease driving pathways may have divergent outcomes or converge on a common disease related outcome. For example, in some instances, different individuals may have distinct genomic responses that all result in a common lung disease endpoint, while in other instances, different individuals may have a shared genomic response that leads to different lung disease endpoints. For example, distinct disease-driving pathways in different individuals may lead to a common outcome of a fibrotic lung disease, such as, e.g., IPF. As such, different patients may respond to different therapies and interventions for the same disease.
  • the present disclosure provides the insight that identifying and neutralizing the particular disease drivers of a given patient may provide an early and effective way to manage lung disease endpoints. Specifically, the present example determined the signalosome in various models of idiopathic pulmonary fibrosis (IPF) to identify biomarkers for treatment with Compound 1
  • IPPF idiopathic pulmonary fibrosis
  • Two exemplary and etiologically distinct models of pulmonary fibrosis are used to characterize the lung transcriptome and proteome that are associated with therapeutic activity of Compound 1. Dosing of Compound 1 starts 2 weeks after disease induction with bleomycin or S1O2 instillation, and dosing is continued via the oral route for 4 weeks. Lung hydroxyproline (HYP) and histopathology are determined. BAL inflammatory cell infiltrates, BAL protein, MPO, BAL cytokines, tissue cytokines, and lung fibrotic markers are evaluated by IHC stainings and transcriptomic analysis by performing real-time PCR. A cytokine/chemokine/growth factor array on minimally invasive BALF samples from the animal models.
  • HEP Lung hydroxyproline
  • BAL inflammatory cell infiltrates, BAL protein, MPO, BAL cytokines, tissue cytokines, and lung fibrotic markers are evaluated by IHC stainings and transcriptomic analysis by performing real-time
  • the present example describes treatment of fibrotic disease of the lung in a patient.
  • a sample of BALF is obtained from a patient with suspected or confirmed pulmonary disease (i.e., suspected of having idiopathic pulmonary fibrosis).
  • mRNA is extracted from the sample, and sequenced to quantify levels of certain biomarkers associated with Compound l’s mechanism of action.
  • expression of biomarkers identified using the method of Example 22 are quantified. If the subject has a level of a biomarker that is above a threshold level, then the subject is to be treated with Compound 1.
  • An exemplary threshold level is a level that is about 20% or more above a predetermined normal range or median level in a healthy patient. If a subject does not have a level of biomarker above the threshold level, then alternate treatment is determined.
  • the lung disease is stabilized and/or ameliorated.
  • Example 24 A Phase 1, Randomized, Double-Blind, Placebo-controlled, Single and Multiple Ascending Dose Study to Determine the Safety, Tolerability, Pharmacokinetics, and Food Effect of Compound 1 in Healthy Adult Participants
  • Primary Endpoint The frequency and severity of treatment-emergent adverse events (TEAEs), including clinically significant abnormal vital signs, electrocardiograms (ECGs), laboratory test results, and physical examination findings.
  • TEAEs treatment-emergent adverse events
  • ECGs electrocardiograms
  • Plasma PK endpoints include:
  • Urine PK endpoints include:
  • Cohort A3 was a food effect cohort, and participants in Cohort A3 only returned to the CRU on Day 14 and, following a 14-day washout, received a second single dose of their assigned treatment on Day 15 following consumption of a high fat meal. For Cohort A3 only, a second period of confinement applied from Day 14 until completion of the 72-hour post-dose assessments on Day 18. [0510] All Part A participants returned to the CRU for a follow-up visit 7 days ( ⁇ 1 day) after their final dose of study drug. For all cohorts in Part A (SAD), the decision to escalate a dose or modify a dose was determined by the SMC following review of the 7-day blinded safety and available PK data from the preceding cohort.
  • SAD Part A
  • Compound 1 dose will not exceed 1200 mg or a dose level that leads to mean whole blood C max >600 ng/mL or mean whole blood AUCo-i ast > 2000 ng*h/mL.
  • C max and AUCi ast values were extrapolated from the mean plasma C max of 800 ng/mL and the mean plasma AUCi ast of 4000 ng*hr/mL observed in non-human primates at the NOAEL of 75 mg/kg/day.
  • Part B Multiple Ascending Dose (MAD)
  • the Compound 1 dose level tested in Part B did not exceed 1000 mg per day or a dose level that led to a mean whole blood Cmax > 600 ng/mL or mean whole blood AUCo-iast > 2000 ng*h/mL.
  • Cmax and AUCo-iast values were extrapolated from the mean plasma Cmax of 800 ng/mL and the mean plasma AUCiast of 4000 ng*h/mL observed in non-human primates at the no observed adverse effect level (NOAEL) of 75 mg/kg/day.
  • Compound 1 dose will not exceed 1000 mg per day or a dose level that leads to mean whole blood C max >600 ng/mL or mean whole blood AUCo-ias t > 2000 ng*h/mL.
  • C max and AUCi ast values were extrapolated from the mean plasma C max of 800 ng/mL and the mean plasma AUCias t of 4000 ng*hr/mL observed in non-human primates at the NOAEL of 75 mg/kg/day.
  • Part D Single-Dose Food-Effect Cohort
  • Part A up to 48 healthy volunteers; Part B: up to 32 healthy volunteers; Part C: up to 16 healthy volunteers; Part D: up to 8 healthy volunteers).
  • Participants must have a minimum body weight of 50 kg and a body mass index (BMI) between >18.0 and ⁇ 32.0 kg/m 2 at screening.
  • BMI body mass index
  • Participants must have clinical laboratory values within normal range as specified by the testing laboratory, unless deemed not clinically significant by the investigator or delegate.
  • Double contraception is defined as a condom and one other form of the following:
  • FSH follicle-stimulating hormone
  • Periodic abstinence e.g., calendar, ovulation, symptothermal, post-ovulation- methods
  • withdrawal are not considered highly effective methods of birth control. Participant complete abstinence for the duration of the study and for 1 months after last study treatment is acceptable.
  • WOCBP must have negative pregnancy test at screening and Day 1 and be willing to have additional pregnancy test as required.
  • Acceptable methods of contraception include the use of condoms and the use of an effective contraceptive for the female partner that includes: OCPs, long-acting implantable hormones, injectable hormones, a vaginal ring or an IUD. Participants with same sex partners (abstinence from penile-vaginal intercourse) are eligible when this is their preferred and usual lifestyle.
  • GI gastrointestinal
  • atrophic gastritis atrophic gastritis
  • lactose intolerance atrophic gastritis
  • H. Helicobacter
  • Fever body temperature > 38 °C
  • symptomatic viral or bacterial infection within 2 weeks prior to screening.
  • HCV hepatitis C antibody
  • HBVAg hepatitis B surface antigen
  • HAV human immunodeficiency virus
  • Participants with a positive toxicology screening panel urine test including qualitative identification of barbiturates, tetrahydrocannabinol (THC), amphetamines, benzodiazepines, opiates, and ***e), or alcohol breath test.
  • any prescription drugs other than hormonal contraception: OCPs, long-acting implantable hormones, injectable hormones, a vaginal ring or an HID
  • over-the- counter (OTC) medication herbal remedies, supplements or vitamins within 1 week prior to dosing and during course of study without prior approval of the investigator and medical monitor.
  • Simple analgesia nonsteroidal anti-inflammatory drug (NSAID)
  • NSAID nonsteroidal anti-inflammatory drug
  • paracetamol may be permitted at discretion of investigator.
  • CYP3 A4 cytochrome P450 3 A4
  • any strong inhibitors or inducers of CYP3 A4 starting from 2 weeks prior to first dose of study drug and until end of study assessments.
  • Compound 1 was provided as a powder in capsule formulation for oral administration.
  • the formulation was a Size 00 Swedish orange capsule containing drug substance (50 mg or 250 mg) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose (HPMC)), iron oxide as coloring agent, and titanium dioxide as an opacifier.
  • HPMC hydroxypropylmethyl cellulose
  • the drug product was stored at room temperature (15 °C - 25 °C).
  • Compound 1 is in a pharmacological class of tyrosine kinase inhibitors (TKI).
  • TKI tyrosine kinase inhibitors
  • Compound 1 is an orally bioavailable small molecule dual kinase inhibitor of platelet-derived growth factor receptors (PDGFR) and vascular endothelial growth factor receptors (VEGFR2). Duration of Treatment
  • Part A In Part A (SAD), participants randomized to active treatment in cohorts Al, A2, A4, A5, and A6 received a single oral dose of Compound 1 administered once on Day 1 only. Participants randomized to active treatment in Cohort A3 (Food Effect) received a single oral dose of Compound 1 on Days 1 and 15.
  • PartB In Part B (MAD), participants randomized to active treatment received oral Compound 1 administered twice daily for 7 consecutive days (Day 1 to Day 7) or twice daily for 14 consecutive days (Day 1 to Day 14), to be determined on completion of Part A (SAD) of the study.
  • Part C In Part C (MAD QD), participants randomized to active treatment received oral Compound 1 administered once daily for 14 consecutive days (Day 1 to Day 14).
  • the placebo capsules visually matched to active study drug and were composed of the same capsule shell but were filled with silicified microcrystalline cellulose.
  • Silicified microcrystalline cellulose is a pharmaceutical excipient composed of co-process microcrystalline cellulose and colloidal silicon dioxide.
  • Safety The safety and tolerability of single and repeat-doses of Compound 1 was investigated according to the following specific assessments: vital signs (systolic and diastolic blood pressure, pulse rate, body temperature, and respiratory rate), 12-lead ECG, clinical laboratory tests (hematology, biochemistry, coagulation, and urinalysis), physical examination, and assessment of TEAEs.
  • descriptive statistics e.g. arithmetic mean, standard deviation [SD], median, minimum and maximum
  • SD standard deviation
  • Frequency summaries e.g. number of observed and percentage of each categories
  • ITT Intent-to-Treat
  • Safety Population All participants who received any amount of study drug (Compound 1 or placebo) were included in the Safety population. The Safety population was used for the summaries of all safety assessments. Participants were analyzed according to treatment received.
  • Pharmacokinetic Population All participants who received any amount of active study drug (Compound 1) and have sufficiently evaluable concentration-time profile to allow determination of at least one PK parameter were included in the PK population. An evaluable PK profile was determined at the discretion of the pharmacokineticist following examination of subjects with dosing or protocol deviations that could potentially affect the PK profile. The PK population was used for the summaries of all PK data.
  • Concomitant medications were coded using the World Health Organization (WHO) drug dictionary Version B3 September 2018 Drug Global. Concomitant medications were listed by participant and summarized by anatomical therapeutic class and preferred name. [0550] Physical examinations at each visit were listed for each participant and summarized using descriptive statistics at each visit by treatment arm.
  • WHO World Health Organization
  • Pharmacokinetics Plasma Compound 1 concentrations, actual blood sampling times, and PK parameters were listed by treatment and protocol specified time point and summarized using descriptive statistics for PK data as outlined above for each scheduled time point by treatment arm. Individual and mean Compound 1 concentration-time profiles were also presented graphically for each treatment. Pharmacokinetic parameters were computed from the individual plasma Compound 1 concentrations using a non-compartmental approach.
  • the following plasma Compound 1 non-compartmental PK parameters were estimated, as appropriate: Cmax, Tmax, AUCo-iast, AUCo-inf, AUCtau (calculated for MAD cohorts only), Kei, ti/2, CL/F (SAD cohorts), CL/Fss (MAD cohorts), Vz/F (SAD cohorts, Vz/Fss (MAD cohorts), RA (calculated for MAD cohorts only).
  • Urine collection time, volume collected, and Compound 1 concentration (Aeu-t2) duration of each sampling interval was listed for each participant and summarized by nominal sampling time point and treatment using descriptive statistics (Number of subjects [N], arithmetic mean, SD, CV%, geometric mean, median, minimum and maximum). Individual and mean Compound 1 cumulative urinary excretion-time profiles for each treatment were also presented graphically. Where urine was collected for PK analysis, the following parameters are calculated, as appropriate: Aeu-t2, CLR, CL/F, and Feu-t2.
  • Example 25 Compound 1 Ameliorates Fibrosis in Alport Syndrome Mouse Model
  • Protein-to-creatinine ratio (PCR). Creatinine standard was prepared at 20, 10, 5, 2.5, 1.25, 0.625 and 0.3125 mg/dL. Urine samples were diluted 20-fold with water. Then 25 pL of samples were added to the 96 well plate and pipet 50 pL of samples, water as the blank, or standards into wells in the clear plate. 50 pL of the DetectX® Creatinine Reagent was added according to manufacturer instructions, (Arbor Assays). Urine protein was calculated for mg/mL. Then, urine protein of the sample was divided by the urine creatinine to determine the protein to creatinine ratio (PCR) and expressed as mg/mg.
  • PCR Protein-to-creatinine ratio
  • Serum BUN Serum samples were diluted 5-fold with distilled water. BUN standard was prepared at 10, 5, 2.5, 1.25, 0.625, 0.3125, and 0.156 mg/dL. 50 pL of samples or appropriate standards into 96 wells in the plate. 50 pL of water was used as the zero standard. Then, 75 pL of Color Reagent A to each well and then added 75 pL of Color Reagent B to each well using a multichannel pipet. The plates were incubated at room temperature for 30 minutes and then the optical density was read at 450 nm according to manufacturer's instructions (Arbor Assays).
  • Serum Creatinine 20 pL of the serum samples were analyzed for serum creatinine levels using LC-MS/MS. Briefly, after spiking in creatinine-d3 as internal standard, mouse serum samples were purified by protein precipitation with acetonitrile. Creatinine was analyzed by LC- MS/MS. Chromatographic separation was performed with a Shimadzu Prominence system on a Primesep 200 (SIELC Technologies) column with a gradient elution (0.1% formic acid in water as mobile phase A and 0.1% formic acid in methanol as mobile phase B). MRM detection (114.05 to 44.2 for creatinine and 117.05 to 47.2 for creatinine-d3) was carried out with AB Sciex API-3200 triple quadrupole mass spectrometer. Creatinine was expressed as mg/dL.
  • hydroxyproline Assay The hydroxyproline (HYP) content from kidney tissues was determined as described by Woessner et al (Archives of Biochemistry and Biophysics, Vol. 93, Issue , May 1961, 440-447).
  • Kidney sections were fixed in 10% formalin for at least 48 hr before preparation for histology. Kidney tissues were processed, dehydrated, embedded in paraffin, and cut into sequential 5 pm sections. Kidney sections were subjected to haematoxylin- eosin (H&E) and Masson’s Trichrome staining using a Trichrome Stain Vector Kit (Sigma). IHC staining using mouse antibodies for Collagen- 1 (Sigma), TGFpi (Cell Signaling), and aSMA (Sigma) were performed by following standard laboratory procedures. Photomicrographs were taken using a microscope (Leica) equipped with a computerized digital camera.
  • Trichrome Staining Working Weigeret’s Iron Hematoxylin Solution was prepared according to label instructions. The slides were deparaffmized and hydrated with deionized water. Then slides were placed in pre-Bouin’s Solution, at room temperature overnight. Slides in a Coplin Jar were washed in running tap water to remove yellow color from sections. The slides were then stained in Working Weigert’s Iron Hematoxylin solution for 5 minutes. The slides were washed in running tap water for 5 minutes. The slides were then stained in Trichrome Stain AB Solution (Sigma) for 5 minutes and placed in 0.5% Acetic acid for 1 minute. The slides were rinsed, dehydrated through alcohol, cleared in xylenes and then mounted.
  • PSR Picrosirius Red Staining. Sections of paraffin-embedded tissue were deparaffmized by sequential washes in xylene (2 times for 5 minutes), 100% ethanol (2 times for 5 minutes), 90% ethanol (once for 5 minutes), 80% ethanol (once for 5 minutes) and 70% ethanol (once for 5 minutes). Samples were then washed in water for 5 minutes. Samples were then incubated in a solution of saturated picric acid containing 0.2% Sirius Red dye at room temperature for 35 min with shaking. Samples were washed with tap water until the wash was colorless. Samples were then dehydrated with serial 5 minute washes in water, 70% ethanol,
  • IHC Stainings for Fibrotic Markers Formalin fixed kidney tissue sections were used for IHC staining. Endogenous peroxidases were quenched with 3.0% hydrogen peroxide in methanol for 30 min. Sections were further blocked with 3.0% bovine serum albumin (BSA) in PBS, exposed to 0.5 % Triton X-100 for 1 h to reduce non-specific antibody binding and incubated with primary antibodies of anti -mouse mouse aSMA antibody (Sigma), Collagen- 1 antibody (Sigma) and TGFB1 (Santa Cruz) at 4 °C overnight.
  • BSA bovine serum albumin
  • the sections were washed with PBS three times, incubated with HRP-conjugated secondary antibodies (Cell Signaling) for 1 hour followed by washing 3 times with lx PBS. The sections were then incubated with a HRP substrate (Invitrogen) for 2-3-minutes to obtain clean staining. After dehydration with a series of increasingly concentrated ethanol solutions, sections were mounted with neutral gum. Images were captured using a Nikon microscope equipped with a computer.
  • Renal injury was evaluated in H&E stained kidney sections, and a renal damage score was assigned on a numerical scale from 0 (normal/no injury) to 4 (severe injury) by two independent observers.
  • analysis consisted of estimating the total amount of tubular and glomerular damages in the cortical regions of the entire kidney section. Injuries included the disruption of normal tissue micro architecture, hemorrhage and monocyte infiltration, inflammation, casts formation, and necrosis and sclerosis.
  • Compound 1 did not decrease serum BUN or serum creatinine (SCr) in Alport mice (FIG. 26G and FIG. 26H, respectively). Treatment with Compound 1 for five weeks significantly decreased kidney fibrosis, as judged by hydroxyproline (HYP) levels, in Alport mice (FIG. 261). Compound 1 also significantly reduced renal histopathological injury score in Alport mice (FIG. 26J).
  • Compound 1 treatment decreased kidney Trichrome staining, an indication of decreased collagen and fibrosis, significantly compared to the vehicle cohort (FIG. 26K).
  • IHC stainings for kidney fibrotic markers demonstrated that Compound 1 decreased TGFpi, aSMA, and Collagen- 1 in Alport mice compared to vehicle (FIG. 26L, FIG. 26M, and FIG. 26N, respectively).
  • Picrosirius red (PSR) staining for collagen showed that Alport mice treated with Compound 1 had significantly reduced collagen as compared to Alport mice treated with vehicle (FIG. 260).
  • Example 26 Effect of Compound 1 in a Passive Heymann Nephritis Model
  • Compound 1 was tested in the Passive Heymann Nephritis model of renal dysfunction in rats.
  • renal dysfunction was mediated by intravenous administration of Anti-FXl A serum to rats, resulting in podocyte-localized deposition of antibodies, whereby immune deposits accumulate and obscure the slit diaphragm (Spicer et ah, J. Immunol. 2007 Jul 1; 179(1): 172-8). This results in podocyte foot process effacement and proteinuria.
  • the model has nearly identical pathology to human membranous nephropathy and gl omerul onephriti s .
  • Urine Analysis Overnight urines were collected using Tecniplast Metabolic Cage Systems for rodents (Tecniplast, Catalogue #3700M071). Animals were kept in metabolic cages for 24 hours. Urine samples were analyzed for protein content using the Bradford Assay (Sigma Cat #B6916-500ML) according to the manufacturer’s protocols. Urine samples were undiluted or diluted to 10-fold or 20-fold with water to be within the assay’s acceptable range. Creatinine was measured using the Quantichrom Creatinine Assay Kit (Bioassay Systems). A standard curve was generated to determine urine sample concentrations. The protein to creatinine ratio was determined and expressed as mg/mg.
  • Serum Analysis Sera were collected at sacrifice and processed in BD Microtainer serum collection tubes according to the manufacturer’s instructions. Sera were stored at -20 °C until analysis. Serum creatinine, cholesterol, and triglycerides were analyzed using validated analytical procedures for human samples. Serum BUN was measured according to the manufacturer’s protocols (Arbor Assays, Catalogue # K024-H).
  • Kidney tissue was collected, and mass determined at the time of tissue collection. Samples were stored at -80 °C until analysis. The hydroxyproline (HYP) content from kidney tissues were determined. Kidney tissue was subjected to hydrolysis in 10 N HC1 for 18 hr at 110 °C, and aliquots were incubated with chloramine T solution, perchloric acid and Ehrlich's reagent at 60 °C for 20 minutes. Absorbance was measured at 550 nm, and the amount of HYP was determined using a hydroxyproline standard curve. Total hydroxyproline is expressed as micrograms per kidney.
  • Body mass was measured at time of sacrifice, day 93.
  • the nintedanib-treated animals had significantly reduced body mass, reduced kidney mass and increased kidney mass to body mass ratio compared to the vehicle cohort (***p ⁇ 0.0001, One-way ANOVA, Tukey).
  • the Compound 1 treatment groups did not have a significant change in body mass (FIG. 27B), kidney mass (FIG. 27C), or the kidney to body mass ratio (FIG. 27D) compared to vehicle or sham.
  • Serum cholesterol was significantly increased (p ⁇ 0.05) in the nintedanib group compared to Sham, 50 mg/kg Compound 1, and 15 mg/kg Compound 1 groups (p ⁇ 0.05, One Way ANOVA) (FIG. 27G). No statistically significant changes in serum cholesterol were noted with Compound 1 treatment. Serum triglycerides were significantly reduced in the 50 mg/kg group compared to vehicle (*p ⁇ 0.05, One way ANOVA, Tukey), whereas the 100 mg/kg and 15 mg/kg groups did not have a statistically significant change in serum triglycerides (FIG. 27H). Serum creatinine (FIG. 271) and BUN (FIG. 27J) did not change significantly in the vehicle or Compound 1 treatment groups compared to sham. However, nintedanib treatment caused a significant increase in serum creatinine and BUN.
  • PAS staining was performed to semi-quantitatively determine the level of glomerulosclerosis of glomeruli. Representative images are shown in FIG. 27M, and glomerular damage scores are shown in FIG. 27N. There was a significant increase in glomerular injury in the vehicle group compared to sham. There was a significant reduction in glomerular damage with Compound 1 treatment at 50 mg/kg and 15 mg/kg. The Nintedanib group had global glomerulosclerosis.
  • Example 27 A Phase 2, Multicenter, Double-Blind, Randomized, Placebo-controlled Study of Safety and Efficacy of Compound 1 in Patients with Primary Glomerular Disease and Persistent Proteinuria
  • the primary objective of this study is to evaluate efficacy of Compound 1 in patients with primary glomerular disease and persistent proteinuria while on standard of care (SOC), as measured by a reduction in the 24-hour urinary protein excretion.
  • SOC standard of care
  • the secondary objective of this study is to evaluate the safety and tolerability of Compound 1 in patients with primary glomerular disease and persistent proteinuria.
  • the eligible patients are randomized in a 1 : 1 : 1 ratio to three dose levels of Compound 1 (200 mg once daily (QD), 400 mg QD, 300 mg twice daily (BID)) or placebo administered daily for 12 weeks.
  • Compound 1 200 mg once daily (QD), 400 mg QD, 300 mg twice daily (BID)
  • BID 300 mg twice daily
  • the study drugs are administered QD or BID orally to patients according to their assignment to QD or BID dosing schedule. All patients take the study drug either once-daily in the morning or twice-daily within 30 minutes of meals and at least 30 minutes apart from any other drugs. The study ends when the last patients have completed 12 weeks of study drug and the follow-up period of 4 weeks.
  • the study period is from the time of signing of the Informed Consent until their last visit.
  • Adverse events are collected during all the study periods and are considered under treatment until 4 weeks after the drug discontinuation.
  • Inclusion Criteria Approximately 100 patients (25 patients per study arm) are planned for this study. Diagnosis and Main Criteria for Inclusion: Inclusion Criteria:
  • ACEi Angiotensin-converting enzyme inhibitor
  • ARB Angiotensin II receptor blocker
  • Renin-angiotensin-aldosterone system (RAAS) blockers and Sodium glucose co transporter 2 (SGLT-2) inhibitors must be stable for at least 12 weeks prior to screening and projected to remain stable through week 16.
  • RAAS Renin-angiotensin-aldosterone system
  • SGLT-2 Sodium glucose co transporter 2
  • Immunosuppressive or immunomodulatory therapy must be stable for at least 12 weeks prior to screening and projected to remain stable through study week 16.
  • HBV Hepatitis B
  • HBV Hepatitis C
  • HBV human immunodeficiency virus
  • ANC absolute neutrophil count
  • NSAIDS non-steroidal anti-inflammatory agents
  • antiplatelet therapy e.g. acetyl salicylic acid up to 325 mg/d, or clopidogrel at 75 mg/d, or equivalent doses of other antiplatelet therapy.
  • Renal disease secondary to systemic disease including but not limited to: systemic lupus erythematosus, anti -neutrophil cytoplasmic antibodies-associated diseases, anti- glomerular basement disease, secondary forms of focal segmental glomerulosclerosis, renal diseases associated with para-proteinemias, C3 glomerulopathy, and diabetic kidney disease
  • Compound 1 is an orally active tyrosine kinase inhibitor. Three dose levels (200 and 400 mg QD and 300 mg BID) are tested in this study. The study drug and the placebo are administered once or twice daily orally after breakfast and dinner and at least 30 minutes apart from any concomitant medications.
  • Compound 1 is administered as a Form A hydrochloride trihydrate salt in a Hypromellose Swedish Orange-colored capsule containing no excipients.
  • Ingredients of the capsule shell are hypromellose (hydroxypropylmethyl cellulose also known as HPMC), iron oxide as a coloring agent, and titanium dioxide as an opacifier.
  • hypromellose hydroxypropylmethyl cellulose also known as HPMC
  • iron oxide as a coloring agent
  • titanium dioxide as an opacifier
  • Reference Therapy Patients receive the standard of care including maximally tolerated/recommended dose of ACEi or ARB but not both. Where applicable, the doses of mineralocorticoid receptor blockers, direct renin inhibitors, and SGLT-2 inhibitor doses must be stable for at least 12 weeks and anticipated to remain stable during the study period.
  • Placebo capsules are composed of the same capsule shell used for the active but are filled with silicified microcrystalline cellulose.
  • Silicified microcrystalline cellulose is a pharmaceutical excipient composed of co-processed microcrystalline cellulose and colloidal silicon dioxide.

Abstract

L'invention concerne des procédés de réduction de la fibrose par l'administration de méthyle (Z)-3-(((4- (N-méthyle-2-(4-méthylpipérazin-1-yl)acétamido)phényl)amino)(phényl)méthylène)-2-oxo-2,3-dihydro-1H-pyrrolo[2,3-b]pyridine-6-carboxylate.
PCT/US2021/039899 2020-07-01 2021-06-30 Réduction de la fibrose et traitement de maladies, de troubles et d'états associés WO2022006278A1 (fr)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150306078A1 (en) * 2012-01-26 2015-10-29 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US20180008581A1 (en) * 2012-01-26 2018-01-11 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US20200102301A1 (en) * 2018-09-18 2020-04-02 Goldfinch Bio, Inc. Pyridazinones and methods of use thereof
WO2021102161A1 (fr) * 2019-11-20 2021-05-27 Angion Biomedica Corp. Méthodes de traitement d'une maladie inflammatoire de l'intestin

Patent Citations (4)

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Publication number Priority date Publication date Assignee Title
US20150306078A1 (en) * 2012-01-26 2015-10-29 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US20180008581A1 (en) * 2012-01-26 2018-01-11 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US20200102301A1 (en) * 2018-09-18 2020-04-02 Goldfinch Bio, Inc. Pyridazinones and methods of use thereof
WO2021102161A1 (fr) * 2019-11-20 2021-05-27 Angion Biomedica Corp. Méthodes de traitement d'une maladie inflammatoire de l'intestin

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Title
LAPOINTE JEAN-FRANÇOIS, LAURENT HARVEY, SARYA AZIZ, HEATHER JORDAN, ROBERT A HEGELE, PIERRE LEMIEUX : "A Single-dose, Comparative Bioavailability Study of a Formulation containing OM3 as Phospholipid and Free Fatty Acid to an Ethyl Ester Formulation in the Fasting and Fed States", CLINICAL THERAPEUTICS, vol. 41, no. 3, 22 February 2019 (2019-02-22), XP055897436, DOI: 10.1016/j.clinthera.2019.01.017 *

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