OA20540A - Compounds, compositions, and methods for modulating CDK9 activity. - Google Patents

Abstract

Inhibitors of CDK9 that are pyrazolo[1,5a]pyrimidine derivatives and salts thereof, corresponding to formula (I) :

Description

COMPOUNDS, COMPOSITIONS, AND METHODS FOR

MODULATING CDK9 ACTIVITY

CROSS-REFERENCE TO RELATED APPLICATIONS

This application daims the benefit of U.S, Provisional Patent Application Nos. 62/752,635, 62/884,993, and 62/910,058; filed on October 30, 2018, August 9, 2019, and October 3, 2019, respectively, which are hereby incorporated by référencé in their entirety.

FIELD

The disclosure provides for compounds that modulate the activity of cyclin-dependent kinase 9 (CDK9), pharmaceutical compositions comprising such compounds, and methods of using the compounds and/or pharmaceutical compositions for treatment, amelioration, and/or prévention of diseases arising from the activity of CDK9, e.g., hyperproli ferai ive diseases, virally înduced infections diseases, and cardîovascular diseases.

BACKGROUND

The cyclin-dependent kinase (CDK) family of proteins are key regulators of the cell cycle and gene transcription. The cell cycle is a regulatory cellular mechanism for the timing of cell growth and division. The cell cycle is a multipronged process that directs cellular prolifération through a sériés of checkpoints that correct for DNA damage, genetic dérangements, and other errors. Nonhuman Primates in Biomédical Research (Second Edition, 2012). Each stage is controlled by a combination of cyclîns and CDKs, where the CDKs phosphorylate a spécifie set of cyclîns to trigger entry into the next stage of the cell cycle. Cell Cycle Merri Lynn Casem B A, PhD, in Case Studies in Cell Biology, 2016. Accumulation of cyclin proteins through régulation of cyclin mRNA transcription fonction as “biologicai switches” to tum CDKs on and off and move the cell from one stage to the next. Id.

CDKs 1, 2, 3, 4 and 6 regulate time of the cell division cycle while CDK 7 and CDK 9 regulate the activity of transcription through régulation of RNA polymerase II via phosphorylation ofits carboxyterminal domain. Lucking, et al., ChemMedChem 2017, Î2, 1776- 1793.

CDK9 Controls the transcriptional activity of key oncogenic proteins such as AR, MYC, MCL-1, and BCL-2 and stimulâtes pro-inflammatory transcription factors such as NFkB and STAT3. Gregory et al., Leukemia. 2015 Jun; 29(6): 1437-1441; Krystofi et al., Curr Phann Des. 2012 Jul; 18(20): 2883-2890. CDK9 fonns a heterodimer with one of four cyclin partners (cyclin Tl, cyclin K, cyclin T2a, or cyclin T2b) called positive transcription élongation factor (P-TEFb). RNA polymerase II pauses mRNA transcription after 20-40 nucléotides along the DNA template due to interaction of négative élongation factors which serve as a major regulatory control mechanism for transcription of rapidly induced genes. P-TEFb overcomes pausing of RNA polymerase II by phosphorylation of the carboxy terminal domain of RNA polymerase II, and inactivation of négative élongation factors. Compounds targeting CDK9 and P-TEFb are currently undergoing clinical study. The enzymatic activity of CDK9 is important for stimulating transcription élongation of most protein coding genes. Krystof, et al., Curr Pharm Des. 2012 Jul; 18(20): 2883-2890.

A number of CDK inhibitors with heterocyclic core structures hâve been developed. For example, purine scaffolds hâve been the source of CDK inhibitors developed for treating cancer, including selicielib (Cyclacel Pharmaceuticals, Inc) and other purine dérivatives. S.C. Wilson et al., Bioorg & Med Chem 2011 Nov; 19(22): 6949-6965. Besides CDK9 these purine dérivatives also target CDK7 and CDK2, whereas CDK2 inhibition causes safety and toxicity concems. CDK9 inhibitors based on a triazîne core bave also been developed, e.g., Atuveciclîb. Lucking et al., ChemMedChem 2017, 12, 1776-1793, Unfortunately, treatment with CDK9 inhibitors remains relatively unsuccessful and involves many adverse effects. Morales et al., Cell Cycle 2016, vol. 15, no. 4, 519-527. Therefore, a need exists for new CDK9 inhibitors for treating diseases mediated by CDK9.

SUMMARY

In an embodiment, the disclosure provides for pyrazolo[l,5-cï]pyrimidines, and dérivatives thereof, that are inhibitors of CDK9.

In an embodiment, the disclosure provides for a compound of formula (I):

or a pharmaceutically acceptable sait thereof, wherein:

R is C1-C6 alkyl, C3-C6 cycloalkyl, tetrahydrofuranyl, or tetrahydropyranyl, optionally substituted at any position with one or more of D, halo, R⁷CO₂R⁸, CO?R⁸, CO?H

R⁷CO2H, NH2, NHR⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO₂NH₃, SO₂NHR⁸, or R¹ is NH2, NHR⁸, OH, OR⁸, NHCOR⁸, NHSO2R^s, SO2NH2, SO2NHR⁸, or R¹ and R² together form a fused C5-C6 cycloaryl, optionally substituted at any 5 position with one or more of D, halo, NH2, NHR⁸, NR⁷R⁸, OH, OR⁸, SH,

SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸;

R², R³, R⁴ and R⁵ are independently H, D, halo, or Cl-C5 alkyl or C3-C6 cycloalkyl optionally substituted at any position with one or more of D, halo, NH₂, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸, ^{î0 or} R³ and R⁴ together form a fused C5-C6 cycloaryl, optionally substituted at any position with one or more of D, halo, NH₂, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸, or R² and R⁵ together form a fused C5-C6 cycloaryl, optionally substituted at any position with one or more of D, halo, NH₂, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, 15 SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸;

R⁶ is H or D;

R is (CH₂)_n wherein n is an integer from 1 to 6; and

R is C1-C6 alkyl or C3-C6 cycloalkyl, optionally substituted at any position with one or moreofD, halo, OH, SH, orNH₂.

In another embodiment, in the compound or sait thereof of formula (I), R¹ is C1-C6 alkyl or C3-C6 cycloalkyl, optionally substituted at any position with NH₂, or R¹ is NHCOR⁸; R², R³, R⁴, and R⁶ are H; R⁵ is cyclobutyl, cyclopentyl, or cyclohexyl, optionally substituted at any position with D, NH₂, OH, NHR⁸, OR⁸, or combinations thereof; and R⁸ is C1-C4 alkyl.

In another embodiment, in the compound or sait thereof of formula (I), R^] is methyl, 25 ethyl, isopropyl, sec-butyl, 3-pentyI, cyclopropyl, cyclopentyl, or NHCOCH3; R², R³, R⁴, and R⁶ aie H, and R is cyclobutyl, cyclopentyl, or cyclohexyl, optionally substituted at any position with NH2.

The compound may be in the form of a composition including a pharmaceutically acceptable carrier.

In another embodiment, the disclosure provides for methods of treating, preventing, or ameliorating CDK9-mediated diseases such as hyperproliferative diseases (e.g., cancer), virally induced infeetious diseases, and cardiovascular diseases, by administering an effective amount of a pyrazolo[l,5-a]pyrimidine dérivative described herein to a subject în need thereof.

DETAILED DESCRIPTION

In an embodiment, a compound described herein capable of use in compositions or methods described herein comprises, consists of, or consists essentially of a compound of formula (I) or a sait thereof. In an aspect, the composition is formulated in a pharmaceutical composition or form.

Any of the compounds described via formula (I) or shown as spécifie compounds may be a single stereoisomer or a mixture of possible stereoisomers. For example, if a single chiral carbon is présent, the compound may be the (S) or (R) stereoisomer, with respect to the chiral carbon, or the compound may be a non-racemic mixture of (S) and (R) isomers, or the compound may be the (S) isomer alone or the (R) isomer alone. If the compound contains more than one chiral carbon, the compound may be a single diastereomer or a mixture of diastereomers.

By “sait” is meant a pharmaceuticaliy acceptable sait, e.g., a hydrochloride sait. A pharmaceuticaliy acceptable sait ’ is a sait that retains the activîty of the compound without significant adverse effects. Examples of pharmaceuticaliy acceptable salts include salts of organic or inorganic acids, e.g., hydrochloric acid, sulphuric acid, methanesulphonic acid, fumanc acid, maleic acid, succinic acid, acetic acid, benzoic acid, oxalic acid, citric acid, tartane acid, carbonic acid, phosphoric acid, trîfluoroacetic acid, and formic acid. The sait may contain one or more équivalents of acid per compound, i.e., the compound may be in the fonn of a dichloride sait.

The active compounds disclosed can also be in the form of their hydrates. The terni “hydrate” includes, e.g„ hemihydrate, monohydrate, dihydrate, trihydrate, and tetrahydrate.

The compounds of this disclosure may exhibit their natural isotopic abundances, or one or more of the atoms may be artificîally enriched in a particular isotope having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number predominantly found in nature. The présent disclosure includes ail suitable isotopic variations of the compounds described herein.

“Alkyl” means branched and straîght-chain saturated aliphatic hydrocarbons, and specifying the number of carbon atoms as in “C1-C6 alkyl” means ail isomers thereof having 1, 2, 3, 4, 5, or 6 carbons in a linear or branched arrangement. Thus, “C1-C6 alkyl” includes methyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, i-butyl, etc.

“Cycloalkyl” means cyclic saturated aliphatic hydrocarbons of the specified number of carbons.

“D” is deuterium.

“Halo” means a halogen substituent, e.g., F, Cl, or Br. Ex amples of compounds of formula (I) include:

139

145

204

206

208

An “effective amount” or “therapeutically effective amount” is an amount of the compound or composition that is sufficient to effect bénéficiai or desired results as described herein when administered to a subject. Effective dosage forms, modes of administration, and dosage amounts may be detennined empirically, and making such déterminations is within the ski 11 of the art. It is understood by those ski lied in the art that the dosage amount will vary with the route of administration, the rate of excrétion, the duration of the treatment, the identity of any other drugs being administered, the âge, size, and species of mammal, e.g., human patient, and like factors well known in the arts of medicine and veterinary medicine. In general, a suitable dose will be that amount of the compound that is the lowest dose effective to produce the desired effect with no or minimal side effects.

A suitable, non-lîmiting example of a dosage of the compounds according to the présent disclosure is from about 1 ng/kg to about 1000 mg/kg, such as from about 1 mg/kg to about 100 mg/kg, including from about 5 mg/kg to about 50 mg/kg. Other représentative dosages of a PI3K inhibitor include about 1 mg/kg, 5 mg/kg, 10 mg/kg, 15 mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg, 45 mg/kg, 50 mg/kg, 60 mg/kg, 70 mg/kg, 80 mg/kg, 90 mg/kg, 100 mg/kg, 125 mg/kg, 150 mg/kg, 175 mg/kg, 200 mg/kg, 250 mg/kg, 300 mg/kg, 400 mg/kg, 500 mg/kg, 600 mg/kg, 700 mg/kg, 800 mg/kg, 900 mg/kg, or 1000 mg/kg.

Yet another embodiment of the présent disclosure is a pharmaceutical composition for treating a CDK9-mediated disease. The CDK9-mediated disease may be a hyperproliferative diseases (e.g., cancer), virally induced infections diseases, or a cardiovascular disease. Examples include acute myelogenous leukemia, primary peritoneal carcinoma, chronic lymphocytic leukemia, relapsed multiple myeloma, non-Hodgkîn’s lymphoma, acute lymphoblastîc leukemia, acute byphenotypic leukemias, advanced breast cancer, non-small cell lung cancer, liver cancer such as hepato cellular carcinoma, and solid advanced tumors. In partîcular, the compounds may be used to treat a cancer caused by aberrant expression of MYCor MCL-1, a hématologie malignancy, or a solid tumor.

The pharmaceutical composition comprises a pharmaceutically acceptable carrier and an effective amount ofthe compounds described herein.

A pharmaceutical composition of the présent disclosure may be administered in any desired and effective manner: for oral ingestion, or as an ointment or drop for local administration to the eyes, or for parentéral or other administration in any appropriate manner such as intraperitoneal, subeutaneous, topical, intradermal, inhalation, intrapuimonary, rectal, vaginal, sublingual, intramuscular, intravenous, intraarterial, intrathecal, or intralymphatic. Further, a pharmaceutical composition of the présent disclosure may be administered in conjonction with other treatments. A pharmaceutical composition of the présent disclosure maybe encapsulated or otherwise protected against gastric or other sécrétions, if desired.

The pharmaceutical compositions of the disclosure are pharmaceutically acceptable and comprise one or more active ingrédients in admîxture with one or more pharmaceuticallyacceptable carriers and, optionally, one or more other compounds, drugs, ingrédients and/or materials. Regardless ofthe route of administration selected, the agents/compounds ofthe présent disclosure are formulated into pharmaceutically-acceptable dosage forms by conventional methods known to those of skill in the art using pharmaceutically acceptable carriers well-known in the art (see, e.g., Reinington, The Science and Practice of Pharmacy (21st Edition, Lippincott Williams and Wilkins, Philadelphia, Pa.) and The National Formulary (American Pharmaceutical Association, Washington, D.C.)) and include sugars (e.g., lactose, sucrose, mannitol, and sorbitol), starches, cellulose préparations, calcium phosphates (e.g., dicalcium phosphate, tri calcium phosphate and calcium hydrogen phosphate), sodium citrate, water, aqueous solutions (e.g., saline, sodium chloride injection, Ringer's injection, dextrose injection, dextrose and sodium chloride injection, lactated Ringer’s injection), alcohols (e.g., ethyl alcohol, propyl alcohol, and benzyl alcohol), polyols (e.g., glycerol, propylene glycol, and polyethylene glycol), organic esters (e.g., ethyl oleate and tryglycerides), biodégradable polymers (e.g., polylactide-polyglycolide, poly(orthoesters), and poly( anhydrides)), elastomeric matrices, liposomes, mîcrospheres, oils (e.g., corn, germ, olive, castor, sesame, cottonseed, and groundnut), cocoa butter, waxes (e.g., suppository waxes), paraffins, silicones, talc, silicylate, etc. Each pharmaceutically acceptable carrier used in a pharmaceutical composition of the disclosure is acceptable in the sense of being compatible with the other ingrédients of the fonnulation and not injurions to the subject. Carriers suitable for a selected dosage fonn and intended route of administration are well known in the art, and acceptable carriers for a chosen dosage form and method of administration can be determined using ordinary skill in the art.

The pharmaceutical compositions of the disclosure may, optionally, contain additional ingrédients and/or materials commonly used in such pharmaceutical compositions. These ingrédients and materials are well known in the art and include (1) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, and silicic acid; (2) binders, such as carboxymethylcellulose, alginates, gel afin, poïyvinyl pyrrolidone, hydroxypropylmethyl cellulose, sucrose and acacia; (3) humectants, such as glycerol; (4) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, sodium starch glycolate, cross-linked sodium carboxymethyl cellulose and sodium carbonate; (5) solution retarding agents, such as paraffin; (6) absorption accelerators, such as quatemary ammonium compounds; (7) wetting agents, such as cetyl alcohol and glycerol monostearate; (8) absorbents, such as kaolin and bentonite clay; (9) lubricants, such as talc, calcium stéarate, magnésium stéarate, solîd polyethylene glycols, and sodium lauryl sulfate; (10) suspending agents, such as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth; (11) buffering agents; (12) excipients, such as lactose, mîlk sugars, polyethylene glycols, animal and vegetable fats, oils, waxes, paraffins, cocoa butter, starches, tragacanth, cellulose dérivatives, polyethylene glycol, silicones, bentonites, silicic acid, talc, salicylate, zinc oxide, aluminum hydroxide, calcium silicates, and polyamide powder; (13) inert diluents, such as water or other solvents; (14) preservatives; (15) surface-active agents; (16) dispersing agents; (17) controlrelease or absorption-delaying agents, such as hydroxypropylmethyl cellulose, other polymer matrices, biodégradable polymers, liposomes, microspheres, aluminum monostearate, gelatin, and waxes; (18) opacifying agents; (19) adjuvants; (20) wetting agents; (21) emulsîfying and suspending agents; (22), solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan;

(23) propellants, such as chlorofluorohydro carbon s and volatile unsubstituted hydrocarbons, such as butane and propane; (24) antîoxidants; (25) agents which render the formulation isotonie with the blood of the intended récipient, such as sugars and sodium chloride; (26) thickening agents; (27) coating materials, such as lecithin; and (28) sweetening, flavorîng, coloring, perfuming and preservative agents. Each such ingrédient or material must be acceptable” in the sense of being compatible with the other ingrédients of the formulation and not injurious to the subject. Ingrédients and materials suitable for a selected dosage fonn and intended route of administration are well known in the art, and acceptable ingrédients and materials for a chosen dosage form and method of administration may be detennined using ordinary skill in the art.

Pharmaceutical compositions suitable for oral administration may be in the form of capsules, cachets, pills, tablets, powders, granules, a solution or a suspension in an aqueous or non-aqueous liquid, an oil-in-water or water-in-oil liquid émulsion, an élixir or syrup, a pastille, a bolus, an electuary or a paste. These formulations may be prepared by methods known in the art, e.g., by means of conventional pan-coating, mixing, granulation or lyophilization processes.

Solid dosage forms for oral administration (capsules, tablets, pills, dragees, powders, granules and the like) may be prepared, e.g., by mixing the active ingredient(s) with one or more pharmaceutical 1 y-acceptable carriers and, optionally, one or more fillers, extenders, bînders, humectants, disintegrating agents, solution retarding agents, absorption accelerators, wetting agents, absorbents, lubricants, and/or coloring agents. Solid compositions of a similar type maybe employed as fillers in soft and hard-filled gelatin capsules using a suitable excipient. A tablet may be made by compression or molding, optionally with one or more aceessory ingrédients. Compressed tablets may be prepared using a suitable binder, lubricant, inert diluent, preservative, disintegrant, surface-active or dispersing agent. Molded tablets may be made by molding in a suitable machine. The tablets, and other solid dosage forms, such as dragees, capsules, pills and granules, may optionally be scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical-formulating art. They may also be formulated so as to provide slow or controlled release of the active ingrédient therein. They may be sterilized by, for example, filtration through a bacteria-retaining filter. These compositions may also optionally contain opacifying agents and may be of a composition such that they release the active ingrédient only, or preferentially, in a certain portion of the gastrointestinal tract, optionally, in a delayed manner. The active ingrédient can also be in microencapsulated form.

Liquid dosage forms for oral administration include pharmaceutically-·acceptable émulsions, microemulsions, solutions, suspensions, syrups and élixirs. The liquid dosage fonns may contain suitable inert diluents commonly used in the art. Besides inert diluents, the oral compositions may also include adjuvants, such as wettîng agents, emulsifying and suspending agents, sweetening, flavoring, coloring, perfuming and preservative agents. Suspensions may contain suspending agents.

Pharmaceutical compositions for rectal or vaginal administration may be presented as a suppository, which may be prepared by mixing one or more active ingredient(s) with one or more suitable nonirritating carriers which are solid at room température, but liquid at body température and, therefore, will melt in the rectum or vaginal cavity and release the active compound. Pharmaceutical compositions which are suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams or spray formulations containing such pharmaceutically-acceptable carriers as are known in the art to be appropriate.

Dosage forms for the topical or transdennal administration include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, drops and inhalants. The active agent(s)/compound(s) may be mixed under stérile conditions with a suitable pharmaceuticallyacceptable carrier. The ointments, pastes, creams and gels may contain excipients. Powders and sprays may contain excipients and propellants.

Pharmaceutical compositions suitable for parentéral administrations comprise one or more agent(s)/compound(s) in combination with one or more pharmaceutically-acceptable stérile isotonie aqueous or non-aqueous solutions, dispersions, suspensions or émulsions, or stérile powders which may be reconstituted into stérile injectable solutions or dispersions just prior to use, which may contain suitable antioxidants, buffers, solutés which render the formulation isotonie with the blood of the intended récipient, or suspending or thickening agents. Proper fluidity can be maintained, for example, by the use of coating materials, by the maintenance of the required particle sîze in the case of dispersions, and by the use of surfactants. These compositions may also contain suitable adjuvants, such as wetting agents, emulsifying agents and dispersing agents. It may also be désirable to include isotonie agents. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by 5 the inclusion of agents which delay absorption.

In some cases, to prolong the effect of a drug (e.g., pharmaceutical formulation), it is désirable to slow its absorption from subeutaneous or intramuscular injection. This may be accomplished by use of a liquid suspension of crystalline or amorphous material having poor water solubility.

The rate of absorption of the active agent/drug then dépends upon its rate of dissolution which, in tum, may dépend upon crystal size and crystalline form. Alternative!y, delayed absorption of a parentérally-administered agent/drug may be accomplished by dissolving or suspending the active agent/drug in an oil vehicle. Injectable depot fonns may be made by fonning microencapsule matrices of the active ingrédient in biodégradable polymers.

Depending on the ratio of the active ingrédient to polymer, and the nature of the particular polymer employed, the rate of active ingrédient release can be controlled. Depot injectable formulations are also prepared by entrapping the drug in liposomes or micro émulsions which are compatible with body tissue. The injectable materials can be sterilized for example, by filtration through a bacterial-retaining fi lier.

The formulations may be presented in unît-dose or multi-dose sealed containers, for example, amputes and vials, and may be stored in a lyophilized condition requiring only the addition of the stérile liquid carrier, for example water for injection, immediately prier to use. Extemporaneous injection solutions and suspensions may be prepared from stérile powders, granules and tablets of the type described above.

As used herein, a “subject” is a mammal, preferably, a human. In addition to humans, categories ofmammals within the scope of the présent disclosure include, for example, agricultural animais, domestîc animais, laboratory animais, etc. Some examples of agricultural animais include cows, pigs, horses, goats, etc. Some examples of domestic animais include dogs, cats, etc. Some examples of laboratory animais include rats, mice, rabbits, guinea pigs, 30 etc.

As used herein, the tenus “treat,” “treating,” “treatment” and grammatical variations thereof mean subjecting an indîvidual subject to a protocol, regimen, process or remedy, in which it is desired to obtain a physiologie response or outcome in that subject, e.g., a patient. In particular, the methods and compositions of the présent disclosure may be used to slow the development of disease symptoms or delay the onset of the disease or condition, or hait the progression of disease development. However, because every treated subject may not respond to a particular treatment protocol, regimen, process or remedy, treating does not require that the desired physiologie response or outcome be achieved in each and every subject or subject, e.g., patient, population. Accordingly, a given subject or subject, e.g., patient, population may fail to respond or respond inadequately to treatment.

As used herein, the terms “ameliorate”, “amehorating” and grammatical variations thereof mean to decrease the severity of the symptoms of a disease in a subject.

As used herein, the terms “prevent”, “preventing” and grammatical variations thereof mean to administer a compound or composition of the présent disclosure to a subject who has not been diagnosed as having the disease or condition at the time of administration, but who could be expected to develop the disease or condition or be at increased risk for the disease or condition. Preventing also includes administration of at least one compound or a composition of the présent disclosure to those subjects thought to be predisposed to the disease or condition due to âge, familial history, genetic or chromosomal abnonnalities, due to the presence of one or more biological markers for the disease or condition and/or due to environmental factors.

The following examples serve to illustrate certain aspects of the disclosure and are not intended to limit the disclosure.

EXAMPLES

The following ex amples describe préparation and testing of représentative compounds.

Example 1: N-cvclopentyl-5-methyl-pvrazoloiL5-alpyrimidin-7-amine (1)

A stirred solution of 7-chloro-5-methyl-pyrazolo[l,5-a]pyrimidine (50.0 mg, 0.3000 mmol), cyclopentanamine (30.48 mg, 0.3600 mmol) and K₂CO₃ (82.34 mg, 0.6000 mmol) in MeCN (4 mL) was heated to reflux for 4h. The reaction mixture was filtered, concentrated under reduced pressure and purified by column chromatography, eluent 30% ethyl acetate in hexane, to give N-cyclopentyl-5-methyl-pyrazolo[l,5-a]pyrimidin-7-amine (51.58 mg, 0.2358 mmol, 79.05%yield) (1) as light yellow, amorphe us solid. The reaction mixture was monitored by TLC (40% ethyl acetate in hexanes; Product Rf=0.4, SM Rf=0.6).

Example 2: N-cvclopentvl-5-isopropyl-pyrazolori,5-a1pvrimidin-7-amine (2)

K2CO3

MeCN, reflux, 4h

To a stirred solution of 7-chloro-5-isopropyl-pyrazolo[i,5-a]pyrimidine (65.0 mg, 0.3300 mmol), cyclopentanamine (0.04 mL, 0.4000 mmol) and K₂CO₃ (91.69 mg, 0.6600 mmol) in MeCN (4 mL) were heated to reflux for 4h. The reaction mixture was filtered, concentrated under reduced pressure and purified by coluinn chromatography, eluent 15% ethyl acetate in hexane to give N-cyclopentyl-5-isopropyl-pyrazolo[l,5-a]pyrimidin-7-amine (48.02 mg, 0.1953 mmol, 58.79%yield) (2) as light yellow, amorphous solid. The reaction mixture was monitored by TLC (20% ethyl acetate in hexanes; Product Rf=0.3, SM Rf=0.6).

Example 3: N-cyciopentvl-5-sec-butvl-pyrazolo[l,5-a]pvrimidin-7-amine (3)

To a stirred solution of 7-chloro-5-sec-butyl-pyrazolo[l,5-a]pyrimidîne (50.mg, 0.2400mmol), cyclopentanamine (24.37 mg, 0.2900 mmol) and K₂CO₃ (82.28 mg, 0.6000 mmol) in MeCN (5 mL) were heated to reflux for 4 h. The reaction mixture was filtered, concentrated under reduced pressure and purified by column chromatography to get Ncyclopentyl-5-sec-butyl-pyrazoIo[l,5-a]pyrimidin-7-amine (21.27 mg, 0.0823 mmol, 34.52% yield) (3) as light yellow, amorphous solid. The reaction mixture was monitored by TLC (30% ethyl acetate in hexanes;, Product Rf=0.4, SM Rf=0.5).

Example 4: r44(5-methylpyrazolo|T.5-a1pyrimidin-4-ium-7yl)amino1cyclohexyl]ammonium dichloride (4)

HCI

Dioxane, 2h

To tert-butyl N-[4-[(5-methylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclohexyl]carbamate (60.0 mg, 03700 mmol), HCl in dioxane (2 mL, 0.1700 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [4-[(5-methylpyrazoïo[l,5-a]pyrimidin-4-ium-7-yl)amino]cyclohexyl]ammonium dichloride (39.S9 mg, 0.1247 mmol, 71.80% yield) (4) as a white solid. The reaction mixture was monitored by TLC ( 100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 5: N,5-dicyclopentvlpvrazolo[L5-alpvrimidin-7-amine (5)

A stirred solution of 7-chloro-5-cyclopentyl-pyrazolo[l,5-a]pyrimidine (50.0 mg, 0.2300 mmol), cyclopentanamine (23.05 mg, 0.2700 mmol) and K₂CO₃ (77.81 mg, 0.5600 mmol) in

MeCN (5 mL) was heated to reflux for 4 h. The reaction mixture was filtered, concentrated under reduced pressure and purified by column chromatography to get N,5dicyclopentylpyrazolo[l,5-a]pyrimidin-7-amine (33.17 mg, 0.1227 mmol, 54.40% yield) (5) as a light yellow solid. The reaction mixture was monitored by TLC (30% ethyl acetate in hexanes; Product Rf=0.4, SM Rf=0.5)

Example 6: r4-[(5-isopropvlpyrazolo[l ,5-a]pyrimidin-4-ium~7vDaminolcycIohexvl]ammonium dichloride (6)

HCl

Dioxane, 2b

To tert-butyl N-[4-[(5-isopropylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclohexyl]carbamate (80.0 mg, 0.2100 mmol), HCl in dioxane (2.mL, 0.2100 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [4-[(5-isopropylpyrazolo[l,5-a]pyrimidin-4-ium-75 yl)amino]cyclohexyl] ammonium dichloride (64.82 mg, 0.1796 mmol, 83.87% yield) (6) as an off-white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product RfHLl, SMRhO.8).

Example 7: r4-r(5-sec-butylpyrazolo|T.5-a]pvriinidin-4-ium-7vDaminolcyclohexy]] ammonium dichloride (7)

To tert-butyl N-[4-[(5-sec-butylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclohexyl]carbamate (65.0 mg, 0.1700 mmol), HCl in dioxane (2.mL, 0.1700mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give[4-[(5-sec-butylpyrazolo[ 1,5-a]pyrimidin-4-ium-7- yl) amino] cyclohexyl] ammonium dichloride (46.75 mg, 0.1297 mmol, 77.35% yield) (7) as an off-white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.78).

Example 8: [4-r(5-cvclopentvlpyrazolo|4.5-a]pyrimidin-4-ium-7yl)amino]cvclohexyllammonium dichloride |8)

To tert-butyl N-[4-[(5-cyclopentylpyrazolo[I,5-a]pyrimidin-7yl)amino]cyclohexyl]carbamate (72.17 mg, 0.1800 mmol), HCl in dioxane (2.mL, 0.1800 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated în vacuo to give[4-[(5-cyclopenty]pyrazolo[l,5-a]pyrimidin-4-ium-7yl)amino]cyclohexyl]ammonium dichloride (59.36 mg, 0.1594 mmol, 88.26% yield) (8) as an off white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1,SM Rf=0.8).

Example 9: [(lS,3S)-3-r(5-cyclopentvlpvrazolo[L5-afovrimidin-4-ium-7vl)amino1cyclopcntyl] ammonium dichloride (9)

To tert-butyl N-[(lS,3S)-3-[(5-cyclopentylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (105.mg, 0.2700mmol), HCl in dioxane (2.0 mL, 0.2700 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [(lS,3S)-3-[(5-cyclopentylpyrazolo[l,5-a]pyrimidin-4-ium-7yl)amino]cyclopentyl] ammonium dichloride (51.28 mg, 0.1431 mmol, 52.54% yield) (9) as a light-yellow solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 10: r(lS,3R)-3-ri5-methvlpvrazo]ofI,5-aÎpyrimidin-4-ium-7vDaminolcvclopentyllammonium dichloride (10)

HCl

Dioxane, 2h

To tert-butyl N-[(lS,3R)-3-[(5-methylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (100.0 mg, 0.3000 mmol), HCl in dioxane (2.mL, 0.3000 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [(lS,3R)-3-[(5-methylpyrazolo[l ,5-a]pyrimidin-4-ium-7yi)amino]cyclopentyl] ammonium dichloride (84.11 mg, 0.26S5 mmol, 88.98% yield) (10) as an off-white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 11: l3-I(5-methylpyrazolorL5-a1pyrimidin-4-iuin-7yl)am ino Icyclobutyl] ammonium dichloride (11)

HCl

Dioxane, 2h

To tert-butyl N-[3-[(5-methylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclobutyl]carbamate (90.mg, 0.2800mmol), HCl in dioxane (2.0 mL, 0.2800 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [3-[(5-methylpyrazolo[ 1,5-a]pyrimidin-4-ium-7-yl)amino]cyclobutyl]ammonium dichloride (65.42 mg, 0.2240 mmol, 79.01% yield) (11) as light brown solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 12: r(TR,3S)-3-r(5-methvlpyrazoloi L5-a]pvrimidin-4-ium-7yl)amino]cyclopentyl1 ammonium dichloride (12)

NH₂

To tert-butyl N-[(lR,3S)-3-[(5-methylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (80.0 mg, 0.2400 mmol), HCl in dioxane (2.0 mL, 0.2400 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo to give [(lR,3S)-3-[(5-methylpyrazolo[l,5-a]pyrimidin-4-ium-7yl)amino]cyclopentyl] ammonium dichloride (61.39 mg, 0.1993 mmol, 82.57% yield) (12) as an off-white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product RM1,SMRM8).

Example 13:(1 S,3R)-N3-(5-isopropvlpyrazolor 1,5-a'|pvrimidm-7-yl)cvclopentane-l .3diamine (13)

To tert-butyl N~[( 1 S,3R)-3-[(5-isopropylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (80.0 mg, 0.2200 mmol), HCl in dioxane (2.0 mL, 0.2200 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lS,3R)-N3-(5isopropylpyrazolo[l ,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (26.03 mg, 0.0978 mmol, 43.97% yield) (13) as a light yellow, amorphous soiid. The reaction mixture was monitored b y TLC (100% ethyl aeetate; Product Rf=0.1, SM Rf=0.8).

Example 14: (lS,3R)-N3-(5-sec-butylpvrazolorL5-a1pyrimidin-7-yl)cvclopentane-l,3diamine (14)

To tert-butyl N-[(lS,3R)-3-[(5-sec-butylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (lOO.mg, Û.2700mmol), HCI in dioxane (2.0 mL, 0.2700 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lS,3R)-N3-(5sec-butylpyrazolo[ 1,5-a]pyrimidin-7-yl)cyclopentane-1,3-diamine (34.04 mg, 0.1245 mmol, 46.51% yield) (14) as a light green, amorphous solid. The reaction mixture was monitored by TLC (100% ethyl aeetate; Product Rf=0.1, SM Rf^0.8).

Example 15: tert-butyl N-r(lS,3R)-3-((5-cyclopentyIpyrazoiori,5-a]pyrimidin-7yDaminolcyclopentylJcarbamate (15)

To tert-butyl N-[(lS,3R)-3-[(5-cyclopentylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (lOO.mg, 0.2600mmol), HCl in dioxane (2.0 mL, 0.2600 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lS,3R)-N3-(5cyclopentyIpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (3L91mg, 0.1118mmol, 43.10% yield) (15) as a light yellow, amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate, Product Rf=0.1, SM Rf=0.8).

Example 16: (lR,3S)-N3-(5-isopropylpyrazolo|T,5-a1pyrimidin-7-yl)cyclopentane-E3diamine (16)

HCl

Dioxane, 2h

To tert-butyl N-[( 1 R,3S)-3-[(5-isopropylpyrazolo[ 1,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (75.mg, 0.2100mmol), HCl in dioxane (2.0 mL, 0.2100 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lR,3S)-N3-(5isopropylpyrazolo[I,5-a]pyrimidin-7-yI)cycIopentane-l,3-diamine (30.78 mg, 0.1158 mmol, 55.49% yield) (16) as a light yellow, amorphous solid. The reaction mixture was monitored b y TLC (100% ethyl acetate; Product Rt=0.1, SM Rf=0.8).

Example 17: ( lS₁3S)-N3-(5-isoprop¥lpvrazoIorL5-a1pyrimidin-7-vl)cyclopentane-l₁3diamine (17)

HCl

Dioxane, 2h

To tert-butyl N-[(lS,3S)-3-[(5-isopropylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (80.0 mg, 0.2200 mmol), HCl in dioxane (2.0 mL, 0.2200 mmol) was added and stirred at room température for 2h. The réaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give(lS,3S)-N3-(5isopropylpyrazolo[l,5-a]pyrimidin-7-yI)cyclopentane-l,3-diamtne (31.02 mg, 0.1160 mmol, 52.14% yield) (17) as light yellow gummy. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rfo0.8).

Example 18: (lS,3S)-N3-i5-sec-butvlpvrazolo[l,5-a1pyrimidin-7-yI)cyclopentane-l,3diamine (18)

To tert-butyl N-[(lS,3S)-3-[(5-sec-butylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (105.0 mg, 0.2800 mmol), HCI in Dioxane (2.0 mL, 0.2800 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (IS,3S)-N3-(5sec-butylpyrazolo[l,5-a]pyrimidin-7-yI)cycIopentane-l,3-diamine (20.55 mg, 0.0752 mmol, 26,74% yield) (18) as a light yellow, amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 19: Nl-(5-sec-butvlpvrazolori,5-alpyrimidin-7-yl)cvclobutane-l,3-diamine (19)

To tert-butyl N-[3-[(5-sec-butylpyrazolo[l,5~a]pyrimidin-7yl)amino]cyclobutyl]carbamate (S5.0 mg, 0.2400 mmol), HCl in dioxane (2.0 mL, 0.2400 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to gîveNl-(5-secbutylpyrazolo[l,5-a]pyrimidin-7-yl)cyclobutane-l,3-diamine (30.61 mg, 0.1180 mmol, 49.91% yield) (19) as a light yellow, amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 20: Nl-(5-cvclopentylpyrazolorL5-a1pyrimidin-7-Yl)cvclobutane-L3-diamine (20)

Totert-butyl N-[3-[(5-cyclopentylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclobutyl]carbamate (80.0 mg, 0.2200mmol), HCl in dioxane (2.0 mL, 0.2200 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give N1 -(520 cyclopentylpyrazolo[l,5-a]pyrimidin-7-yl)cyclobutane-l,3-diamine (29.52 mg, 0.1088 mmol, 50.51% yield) (20) as a light-yellow gummy. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 21.; Nl-(5-isopropylpyrazolof L5-alpvrimidin-7-vl)cvclobutane-1.3-diamine (21)

HCl

Dioxane, 2h

To tert-butyl N-[3-[(5-isopropylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclobutyl]carbamate (75. 0 mg, 0.2200 mmol), HCl in dioxane (2.0 mL, 0.2200 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated în vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give Nl-(5isopropylpyrazolo[l,5-a]pyrimidin-7-yl)cyclobutane-l,3-diamine (32.01 mg, 0.1251 mmol, 57.62% yield) (21) as light yellow, amorphous solid. Tire reaction mixture was monitored b y TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

¹⁰ Example 22: ( 1 R,3S)-N3-(5-sec-butylpyrazolori,5-a1pyrimidin-7-vl)cvclopentane-1.3- diamine (22)

HCl

Dioxane, 2h

To tert-butyl N-[(lR,3S)-3-[(5-sec-butylpyrazolo[I,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (100.0 mg, 0.2700 mmol), HCl in dioxane (2.0 mL, 0.2700 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lR,3S)-N3-(5sec-butylpyrazolo[l,5-a]pyrimidin-7-yI)cyclopentane-l,3-diamine (28.39 mg, 0.1038 mmol,

38.79% yield) (22) as a light yellow, amorphous solid. The reaction mixture was monitored by

TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 23: ( 1 R,3S)-N3-(5-cYclopentylpyrazolo[ 1,5-alpvrimidin-7-yl)cyclopentane-1.3diamine (23)

HCl

Dioxane, 2h

To tert-butyl N-[( 1 R,3S)-3-[(5-cycIopentylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (100.0 mg, 0,2600 mmol), HCl in dioxane (2.0 mL, 0.2600 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo and neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge to give (lR,3S)-N3-(5cyclopentylpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (33.05 mg, 0.1158 mmol, 44.64% yield) (23) as a light yellow, amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=O.S).

¹⁰ Example 24: (lS,3S)-N3-i5-methvlpyrazolori,5-a1pyrimidin-7-vl)cyc]opentane-l.3-diamine (24)

HCI

Dioxane, 2h

¹⁵ To tert-butyl N-[(lS,3S)-3-[(5-methyIpyrazolo[l,5-a]pyrimidin-7- yl)amino]cyclopentyl]carbamate (80.0 mg, 0.2400 mmol), HCl in dioxane (2.0 mL, 0.2400 mmol) was added and stirred at room température for 2h. The reaction mixture was evaporated in vacuo, neutralized by PL-HCO3 MP SPE 200MG/6ML cartridge and purified by prep HPLC to give ( 1 S,3S)-N3-(5-methylpyrazolo[l ,5-a]pyrimidin-7-yl)cyclopentane-l ,3-diamine (21.15 mg, 0.0899 mmol, 37.26% yield) (24) as an off-white solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 25:(1 R,3R)-N3-(5-ethvlpyrazolor 1,5-a]pyrimidin-7-yl)cvc]opentane-l .,3-diamine

To a stirred solution of tert-butyl N-[(lR,3R)-3-[(5-ethylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (62.0 mg, 0.1800 mmol) in DCM (3.5896 mL) at 0 °C was added trifluoroacetic acid (0.34 mL, 4.49 mmol). The reaction was allowed to wann to room température and continuously stirred over 2 hours, whereupon LC-MS revealed reaction complété. The reaction was directly concentrated, washed with pentane and dried to yield (lR,3R)-N3-(5-ethylpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (35 mg, 0.1427 mmol, 79.49% yield) (25). The reaction mixture was monîtored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 26: (lR,3R)-N3-(5-propvlpyrazoIorL5-a1pyrimidin-7-vl)cyclopentane-l.3-diamine

To a stirred solution of tert-butyl N-[(IR,3R)-3-[(5-propylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (41.0 mg, 0.1100 mmol) in DCM (2.2811 mL) at 0 °C was added trifluoroacetic acid (0.22 mL, 2.85 mmol). The reaction was allowed to wann to room 20 température and continuously stirred over 2 hours, whereupon LC-MS revealed reaction complété. The reaction was directly concentrated, washed with pentane and dried to yield (IR,3R)-N3-(5-propylpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (16 mg, 0.0617 mmol, 54.09% yield) (26). The réaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8)

Example 27: ( 1 S,3S)-N3-(5-propyÎpyrazolor 1,5-a1pyrimidin-7-yDcyclopentane-l ,3-diamine 27)

To a stirred solution of tert-butyl N-[(l S,3S)-3-[(5-propylpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (47.0 mg, 0.1300 mmol) in DCM (2.615 mL) at 0 °C was added trifluoroacetic acid (0.25 mL, 3.27 mmol). The reaction was allowed to warm to room température and continuously stirred at room température over 2 hours, whereupon LC-MS revealed reaction complété. The reaction was directly concentrated, washed with pentane and dried to yield (lS,3S)-N3-(5-propylpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (33 mg, 0.1272 mmol, 97.32% yield) (27). The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8)

Example 28: (lS,3S)-N3-(5-ethylpyrazolo[L5-a]pvrimidin-7-vl)cyciopentane-L3-diamine

To a stirred solution of tert-butyl N-[(lS,3S)-3-[(5-ethyIpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (64.0 mg, 0.1900 mmol) in DCM (3.7054 mL) at 0 °C was added trifluoroacetic acid (0.35 mL, 4.63 mmol). The reaction was allowed to wann to room température and continuously stirred over 2 hours, whereupon LC-MS revealed reaction complété. The reaction was directly concentrated, washed with pentane and dried to yield (I S,3S)-N3-(5-ethyIpyrazolo[l,5-a]pyrimidin-7-yl)cyclopentane-l,3-diamine (17 mg.

0.0693 mmol, 37.40% yield) (28). The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=O.S).

Example 29: N-cyclohexyl-5-isopropyl-pvrazolorL5-a1pyrimidin-7-amine (29)

A stirred solution 7-chloro-5-isopropyl-pyrazolo[l,5-a]pyi-imidine (100.0 mg, 0.4800 mmol), cyclohexanamine (56.76 mg, 0.5700 mmol) and K.₂CO₃ (197.44 mg, 1.43 mmol) in MeCN (10 mL) was heated to reflux for 16 h. The reaction mixture was monitored by TLC (20% ethyl acetate in hexanes; Product Rf=0.3, SM Rf=0.6). Upon completion, the reaction mixture was concentrated under reduced pressure. Water was then added (50 ml) and extracted with ethyl acetate (20 ml x 2). The combined organic layers were combined, dried using anhydrous NajSCU, filtered, concentrated and purified via column chromatography (20% ethyl acetate in hexanes) to give N-cyclohexy]-5-isopropyl-pyrazolo[l,5-a]pyrimidin-7-amine (110 mg, 0.4216 mmol, 88.40% yield) (29) as an off-white solid.

Example 30: 5-isopropyl-N-f(3R)-tetrahvdrofuran-3-vl]pyrazolorL5-a1pyrimidin-7-amine (30)

Cl

K₂CO₃

MeCN, reflux, 16h

A stirred solution 7-chloro-5-isopropyl-pyrazolo[l ,5-a]pyrimidine (100.0 mg, 0.4800 mmol), (3R)~tetrahydrofuran-3-amine (49.86 mg, 0.5700 mmol) and K₂CO₃ (197.44 mg, 1.43 mmol) în MeCN ( 10 mL) was heated to reflux for 16 h. The reaction mixture was concentrated under reduced pressure, then water was added (50 ml) and extracted with ethyl acetate (20 ml x 2). The combined organic layers were dried under anhydrous Na₂SO₄ and concentrated. The crude was purified via column chromatography (20% ethyl acetate in hexane) to give 5 isopropyl-N-[(3R)~tetrahydrofuran-3-yl]pyrazolo[I,5-a]pyrimidin-7-amine (50 mg, 0.2016 mmol, 42.28% yield) (30) as a colourless, thick liquid.

Example 31: N-cyclobutyl-5-isopropvl-pyrazolori.5-a]pyrimidin-7-amine (31)

Cl

NH₂

K₂CO₃

MeCN, reflux, 16h

To a stirred solution 7-chloro-5-isopropyl-pyrazolo[l,5-a]pyrimidine (100.0 mg, 0.4800 mmol), cyclobutanamine (40.7 mg, 0.5700 mmol) and K₂CO₃ (197.44 mg, 1.43 mmol) in MeCN (10 mL) were heated to reflux for 16 h. The reaction mixture was concentrated under reduced pressure, then was added water (50 ml) and extracted with ethyl acetate (20 ml x 2). The combined organic layers were dried under anhydrous Na₂SO₄. The crude was purified via column chromatography (20% ethyl acetate in hexane) to give N-cyclobutyl-5-isopropylpyrazolo[l,5-a]pyrimidin-7-amine (50 mg, 0.2135mmol, 44.77% yield) (31) as an off-white solid

Example 32: iïlR.3R)-3-r(5-cycIopropylpyrazolo|T,5-alpyrimidin-7yDaminolcyclopentyUammonium; 2,2,2-trîfluoroacetate (32)

TFA

HFIP

To a stirred solution of tert-butyl N-[(lR,3R)-3-[(5-cyclopropylpyrazolo[l,5a]pyrimidin-7-yl)amino]cyclopentyl]carbamate (110.0 mg, 0.3100 mmol) in HFIP (51.71 mg, 0.3100 mmol), tert-butyl N-[(lR,3R)-3-[(5-cycIopropyIpyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (110.0 mg, 0.3100 mmol) and trifluoroacetic acid (0.12mL, 1.54mmol) were added and stirred at room température for 2h. The reaction mixture was concentrated and the résultant solid was triturated with ether to give [( 1 R,3R)-3-[(541 cyclopropylpyrazolo[l,5-a]pyrimidin-7-yl)amino]cyclopentyl]ammonium; 2,2,2-trifluoroacetate (58.37 mg, 0.1572 mmol, 51.08% yield) as light brown amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Représentative synthetic scheme for compounds 33 and 34:

NCS

MeCN, 2h (74% yield)

K₂CO₃

MeCN, reflux, 16h (79% yield)

Dioxane, 100^qC,10h (66% yield)

Pd₂(dba)₃ Xantphos GenS, Cs₂CO₃

TFA

HFIP * (70% yield)

Ex ample 33a

To a solution 5,7-dichloropyrazolo[ 1,5-a]pyrimidine (200.0 mg, 1.06 mmol) in MeCN (5 mL) was added N-chlorosuccinimide (149.15 mg, 1.12 mmol) at 0°C. The resulting mixture was stirred at room température for 2h. The reaction mixture was concentrated under reduced pressure, purified via column chromatography to give 3,5,7-trichloropyrazolo[l,5~a]pyrimidine (175 mg, 0.7867 mmol, 73.95% yield) as a light yellow solid. The reaction mixture was monitored by TLC (20% ethyl acetate in hexanes; Product Rf=0.6, SM Rf=0.5).

Ex ample 33b

Cl

Cl r/VRJ

K₂CO₃

MeCN, reflux, 16h

A stirred solution 3,5,7-trichloropyrazolo[ 1,5-a]pyrimidine (0.4 g, 1.8 mmol), tert-butyl N-[(lR,3R)-3-aminocyclopentyl]carbamate (0.4 g, 1.98 mmol) and K₂CO₃ (0.74 g, 5.39 mmol) in MeCN (20 mL) was heated to reflux for 16h. The reaction mixture was filtered, concentrated, and purified via column chromatography (30% ethyl acetate in hexanes) to give tert-butyl N-[( 1 R,3R)-3-[(3,5-dichloropyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]carbamate (0.5500 g, 1.4238 mmol, 79.187% yield) as an off-white solid. The réaction mixture was monitored by TLC (40% ethyl acetate in hexanes; Product Rf=0.5, SM Rf=0.8).

Example 33c

O

Dioxane, 100°C, 10h

NHBoc

Pd₂(dba)₃ Xantphos Gen3, Cs₂CO₃

A stirred solution tert-butyl N-[(lR,3R)-3-[(3,5-dichIûropyrazolo[l,5-a]pyrimidin-7yl) amino] cyclopenty]]carbamate (100.0 mg, 0.2600 mmol), acetamide (22.94 mg, 0.3900 mmol), Cs₂CO₃ (252.41 mg, 0.7800 mmol), Pd₂(dba)₃ (23.71 mg, 0.0300 mmol) and Xantphos (43.83 mg, 0.0500 mmol) in dioxane (5 mL) were heated at I Ü0°C in a sealed tube for 10 h. The reaction mixture was filtered through a pad of celite, concentrated under reduced pressure and purified by column chromatography to give tert-butyl N-[(lR,3R)-3-[(5-acetamido-3-chloropyrazolo[l,5-a]pyrimidin-7-yl)amino]cyclopentyl]carbamate (70 mg, 0.1712 mmol, 66.13% yield) as a brown liquid. The reaction mixture was monitored by TLC (30% ethyl acetate in hexanes; Product Rf^0.4, SM Rf=0.6).

Ex ample 33: r(lR.3R)-3-r(5-acetamido-3-chloro-pyrazoloFI,5-a]pyrimidin-7yl)amino] cyclopentyl] ammonium; 2,2,2-trifluoroacetate (33)

TFA

HFIP

To a stirred solution of tert-butyl N-[(lR,3R)-3-[(5-acetamido-3-chloro-pyrazoIo[l,5a]pyri midi n-7-yl)amino] cyclopentyl] carbamate (65.0 mg, 0.1600 mmol) in HFIP (1.1 mL, 01600 mmol), trifluoracetic acid (0.06 mL, 0.7900 mmol) was added and stirred at room température for 2h. The reaction mixture was concentrated and the résultant solid was triturâted with ether to give [(1 R,3R)-3-[(5-acetamido-3-chloro-pyrazolo[l,5-a]pyrimidin-7yl)amino]cyclopentyl]ammonium; 2,2,2-trifluoroacetate (47.21 mg, 0.1117 mmol, 70.24% yield) (33) as an off-white amorphous solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rfi=0.8).

Example 34: r(lR.3R)-3-[(5-acetamidopyrazolo[1.5-alnyrimidÎn-7yDamino] cyclopentyl] ammonium; 2,2,2-trifluoroacetate (34)

TFA

HFIP

To a stirred solution of tert-butyl N-[(lR,3R)-3-[(5-acetamidopyrazolo[l,5-a]pyrimidin7-yi)amino]cyclopentyl]carbamate (65.0 mg, 0.1700 mmol) in HFIP (29.17 mg, 0.1700 mmol), trifluoracetic acid (0.07 mL, 0.8700 mmol) were added and stirred at room température for 2h. The reaction mixture was concentrated and the résultant solid was triturated with ether to give[(IR,3R)-3-[(5-acetamidopyrazoIo[l,5-a]pyrimidin-7-yl)amino]cyclopentyI]ammonium;

2,2,2-trifluoroacetate (19.5 mg, 0.0502 mmol, 28.93% yield) as brown solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8).

Example 35: (lS,3S)-N3-[5-(l-ethylpropvl)pvrazoÎo[l,5-a1pvrimidin-7-vl1cYclopentane5 1,3-diamine (35)

1) CD), THF, rt, 1Sh

2) MgCI₂₁ THF, 50 ’C, 16h (35-45% yield)

POCI₃ neat, reflux, 2fi (58-77% yield)

K₂C0î r—\rs;

/NHBOC

AC N (67-00% yield)

HCl in dioxane rt, 4h (04-95% yield)

(1.84-1.09 equiv HCl) __(!!+ίΑ3ΐ_ rt, 4h (08% yield)

Step 1

1) CDI, THF, rt, 16h

2) MgC^, THF, 50 ’C, 16h (35-45% yield)

2-Ethylbutanoic acid (7.5 g, 64.57 mmol) was dissolved in THF (150 mL) and cooled to 0 °C. Within 20 min CDI (16.23 g, 100.08 mmol) was added portion-wise. (free-tase) The reaction warmed to room temp (rt) and the mixture was stirred at rt ovemight (Solution A). In another flask MgCl₂ (6.14 g, 64.57 mmol) and potassium 3-ethoxy-3-oxo-propanoate (17 g, 15 100.1 mmol) were mixed with THF (150 mL) and stirred under argon ovemight at 50 °C. The résultant white suspension was cooled to rt and solution A was added dropwise over 10 min and the reaction mixture (RM) was stirred for 16h at room température. After several minutes a sticky, amoiphous solid appeared whereupon after several hours the reaction mixture became homogenous in appearance. The RM was concentrated to about a third, taken up in half sat.

potassium bisulphate solution and extracted twice with ethyl acetate. The organic layers were subsequently washed with a sat. sodium bicarbonate solution, combined, dried over anhydrous sodium sulfate, filtered and evaporated. Purification by column chromatography gave ethyl 4ethyl-3-oxo-hexanoate (4.3 g, 23.087 mmol, 35.8% yield) as a transparent liquid. The RM was monitored by TLC (10% ethyl acetate in hexanes; Product Rfi=0.6, SM Rf=0.1).

Step 2

To a suspension of ethyl 4-ethyl-3-oxo-hexanoate (4.4 g, 23.62 mmol) in acetic acid (11 mL) at 70 °C was added IH-pyrazoL5-amine (4.71 g, 56.7 mmoi) in two portions (the second portion was added after 2 hours of stirring the first portion) over a 4 hour perîod. Upon consomption of SM as indicated by TLC, the reaction was cooled to rt and the solvent was evaporated in a rotary evaporator. The residue was treated with ethyl acetate and filtered to give 5-(l-ethylpropyl)-4H-pyrazolo[l,5-a]pyrimidin-7-one(3.7 g, 17.7 mmol, 74.9% yield) as an off-white solid. The reaction mixture was monitored by TLC (5% methanol in dichloromethane; Product Rf=0.3, SM Rf=0.8).

A stirred solution of 5-(l-ethyIpropyl)-4H-pyrazolo[l,5-a]pyrimidin-7-one (3.7 g, 18.03 mmol) in POC1₃ (33.7 mL, 360.52 mmol) was heated to reflux for 4 hours. The reaction mixture was cooled to room température, excess reagent was evaporated in a rotary evaporator, and the residue was treated with ice-water. The chlorinated product was extracted from aqueous mixture by DCM. The organic layer was separated, dried over anhydrous Na₂SO₄, filtered and purified by column chromatography to give 7-chloro-5-(l-ethylpropyI)pyrazolo[l,5-a]pyrimidine (3.1 g, 13.9 mmol, 76.9% yield) as a light yellow liquid. The reaction mixture was monitored by TLC (20% ethyl acetate in hexanes; Product RfoO.6, SM Rf=0.1).

Step 4

/NHBoc H₂N

K₂CO₃

AC N (67-80% yield)

To a stirred solution 7-chloro-5-(Dethylpropyl)pyrazolo[L5-a]pyrimidine (2.3 g, 10.28 mmol), tert-Butyl ((lS,3S)-3-aminocyclopentyl)carbamate (2.27 g, 11.31 mmol) and K2CO3 (4.26 g, 30.84 mmol) in MeCN (20 mL) were heated to reflux for 16 hours. The reaction mixture was filtered, concentrated under reduced pressure and purified by column chromatography, eluent 30% EA in hexane to give tert-butyl N-[(l S,3S)-3-[[5-(lethylpropyl)pyrazolo[l,5-a]pyrimidin-7-yl]amino]cyclopentyl]carbamate (4.5 g, 11.6 mmol, 112.8% yield) as an off-white solîd. The reaction mixture was monîtored by TLC (40% ethyl acetate in hexanes; Product Rf=0.5, SM Rf=0.7).

Step 5

tert-butyl N-[(lS,3S)-3-[[5-(l-ethylpropyl)pyrazolo[l,5-a]pyrimidin-7- i¹·⁸⁴'¹·^{39 eE}i^{uiv HCi}) yl]amino]cyclopentyl]carbamate (1.0 g, 2.58 mmol) in 1,4-Dioxane (0.2 mL), 4 M HCl in Dîoxane (3.22 mL, 12.9 mmol) was added and stirred at room température for 4 hours. The reaction mixture was evaporated in vacuo, triturated with pentane and lyophîlized from MeCN:H2O to give [(lS,3S)-3-[[5-(l-ethylpropyl)pyrazolo[l,5-a]pyrimidin-4-ium-7yl]amino]cyciopentyl]ammonium dichloride (0.9 g, 2.5 mmol, 96.8% yield) as a pale-yellow sticky solid. The reaction mixture was monîtored by TLC (100% ethyl acetate; Product Rf=0.1, SM Rf=0.8). 1H NMR (400 MHz, DMSO-d6) δ 15.00 (s, 1H), 9.93-9.86 (m, 1H), 8.51 (s, 3H), 8.30 (s, IH), 6.84 (s, 1H), 6.58 (s, 1H), 4.95 (q, J = 7.8 Hz, 1H), 3.77-3.66 (m, 1H), 2.84-2.71 (m, 1H), 2.29-2.05 (m, 4H), 1.94-1.63 (m, 6H), 0.81 (t, J = 7.4 Hz, 6H). LC-MS (m/z 287.21, found 288.0 [M+H+])

Step 6

ethyl propyl)pyrazoIo[l,5-a]pyrimidin-4-ium-7-yl]amîno]cyclopentyl]ammonium-dichloride (0.2 g, 0.5600 mmol) in aq. NH3 (4.0 mL, 0.56 mmol) was added and stirred at room température for 4 hours. The reaction mixture was evaporated in vacuo, triturated with pentane and lyophilized from MeCN:H2O to give (lS,3S)-N3-[5-(l-ethylpropyl)pyrazolo[l,5a]pyrimidin-7-yI]cyclopentane-l,3-diamine (140 mg, 0.49 mmol, 87.8% yield) as a pale-yellow sticky solid. The reaction mixture was monitored by TLC (100% ethyl acetate; Product Rf=0.1,

SM Rf=0.8). 1H NMR (400 MHz, DMSO-d6) δ 7.95 (d, J = 2.2 Hz, 1H), 6.86 (s, 1H), 6.29 (d, J = 2.2 Hz, 1 H), 5.95 (s, 1H), 4.3 W.19 (m, 1 H), 3.57-3.44 (m, 1 H), 2.52-2.44 (m, 1H), 2.362.22 (m, IH), 2.09-1.79 (m, 3H), 1.80-1.59 (m, 5H), 1.58-1.24 (m, 3H), 0.83 (t, J = 7.4 Hz, 6H). LC-MS (m/z 287.21, found 288.5 [M+H+]).

Table: Compound Data

Compound	MF	Mass	[M+X]	Mass Found
1	C12H16N4	216.1375	[M+H]	216.8
2	C14H20N4	244.1688	[M+H]	244.7
3	C15H22N4	258.1844	[M+H]	258.7
4	C13H19N5	245.164	[M+H]	246
5	C16H22N4	270.1844	[M+H]	270.7
6	C15H23N5	273.1953	[M+H]	274.2
7	C16H25N5	287.211	[M+H]	287.7
8	C17H25N5	299.211	[M+H]	299.8
9	C16H23N5	285.1953	[M+H]	285.8
10	C12H17N5	231.1484	[M+H]	232.2
11	C11H15N5	217.1327	[M+H]	218.3
12	C12H17N5	231.1484	[M+H]	232.3
13	C14H21N5	259.1797	[M+H]	259.8
14	C15H23N5	273.1953	[M+H]	273.9
15	C16H23N5	285.1953	[M+H]	286.4
16	C14H21N5	259.1797	[M+H]	259.8
17	C14H21N5	259.1797	[M+H]	260.2
18	C15H23N5	273.1953	[M+H]	273.8
19	C14H21N5	259.1797	[M+H]	260
20	C15H21N5	271.1797	[M+H]	372
21	C13H19N5	245.164	[M+H]	245.9
22	C15H23N5	273.1953	[M+H]	274,3
23	C16H23N5	285.1953	[M+H]	286.2
24	C12H17N5	231.1484	[M+H]	231.7
25	C13H19N5	245.164	[M+H]	246.2
26	C14H21N5	259.1797	[M+H]	260.2
27	C14H21N5	259.1797	[M+H]	260.2
28	C13H19N5	245.164	[M+H]	246.2
29	C15H22N4	258.1844	[M+H]	259
30	CI3H18N4O	246.1481	[M+H]	246.9
31	C13H18N4	230.1531	[M+H]	231

Compound	MF	Mass	[M+X]	Mass Found
32	C14H19N5	257.164	[M+H]	257.7
33	C13H17C1N6O	308.1152	[M+H]	308.9
34	C13H18N6O	274.1542	[M+H]	275.3
35	C16H25N5	287.211	[M+H]	288
36	C13H17C1N6O	308.1152	[M+H]	308.9
37	C13H18N6O	274.1542	[M+H]	275.3
38	C15H23N5	273.1953	[M+H]	274.4
39	C15H23N5	273.1953	[M+H]	273.7
40	C14H20C1N5	293.1407	[M+H]	293.8
41	C13H19N5	245.164	[M+H]	245.9
42	C15H2IN5	271.18	[M+H]	271.9
43	C14H19N5	257.164	[M+H]	257.7
44	C14H21N5	259.1797	[M+H]	260.1
45	C14H20C1N5	293.1407	[M+H]	293.8
46	C15H21N5O	287.1746	[M+H]	288.3
47	C13H19N5	245.164	[M+H]	246.4
48	C14H21N5	259.1797	[M+H]	260
49	C18H27N5O2	345.2165	[M+H]	346.3
50	C22H27N5O2	393.2165	[M+H]	394.5
51	C15H19N5O	285.159	[M+H]	285.8
52	C16H23N5O	301.1903	[M+H]	301.8
53	CI5H23N5	273.1953	[M+H]	273.9
54	C16H23N5O2	317.1852	[M+l-I]	318
55	C16H23N5O	301.1903	[M+H]	301.8
56	C16H24N6O	316.2012	[M+H]	317
57	C22H29N7O	407.2434	[M+H]	408
58	C21H28N6	364.2375	[M+H]	265.4
59	C17H27N5O	317.2216	[M+H]	318.1
60	C15H22N6O2	318.1804	[M+H]	319
61	C18HI9N5O2	337.1539	[M+H]	338
62	C20H33N5O	359.2685	[M+H]	360
63	C17H24N8	340.2124	[M+H]	340.9
64	C15H20N6	284.1749	[M+H]	285.2
65	C16H24N6	300.2062	[M+H]	301
66	C15H23N7	301.2015	[M+H]	302
67	C22H26C1N5O2	427.1775	[M+H]	428.2
68	CI8H26C1N5O2	379.1775	[M+H]	380.2
69	C21H21N5	343.1797	[M+H]	344.5
70	C19H23N5O	337.1903	[M+H]	338.3
71	C21H22N6	358.1906	[M+H]	359.3
72	C15H22N6O	302.1855	[M+H]	303.2
73	C18H2IN5	307.1797	[M+H]	308.3
74	C18H20C1N5O	357.1356	[M+H]	357.8
75	CI7H18C1N5	327.1251	[M+H]	327.8
76	C15H22C1N5	307.1564	[M+H]	307.8
77	C12H17N5O	247.1433	[M+H]	248.1

Compound	MF	Mass	[M+X]	Mass Found
78	C16H19N7O	325,1651	[M+H]	326.3
79	C15H24N6	288.2062	[M+H]	288.9
80	C21H22N6	358.1906	[M+H]	358.9
81	C21H21N5	343.1797	[M+H]	344.1
82	C17H17C12N5	361.0861	[M+H]	362.3
83	C15H19N7	297.1702	[M+H]	298.2
84	C22H25N5O2	391.2008	[M+H]	392
85	C19H29N5O2	359.2321	[M+H]	360.3
86	C15H24N6	288.2062	[M+H]	289.3
87	C23H29N5O2	407.2321	[M+H]	408.4
88	C14H22N6O	290.1855	[M+H]	291.2
89	C16H21N5O2	315.1695	[M+H]	316.3
90	C14H22N6O	290.1855	[M+H]	291.5
91	C13H19N5O	261.159	[M+H]	262.3
92	C14H22N6	274.1906	[M+H]	275.2
93	C16H25N5	287.211	[M+H]	288.3
94	C18H21N5O	323.1746	[M+H]	324
95	C18H21N5O	323.1746	[M+H]	324.2
96	CI8H21N5O	323.1746	[M+H]	324.4
97	C19H24N6	336.2062	[M+H]	337.4
98	C13H20N6	260.1749	[M+H]	261.3
99	C16H25N7	315.2171	[M+H]	316.4
100	C17H20N6	308.1749	[M+H]	309.1
101	C17H20N6O	324.1699	[M+H]	325.1
102	C16H23N7O	329.1964	[M+H]	330.3
103	C16H19N7	309.1702	[M+H]	310.3
104	C18H18C1N5O2	371.1149	[M+H]	372.3
105	C19H23N5	321.1953	[M+H]	322.4
106	C16H24C1N5	321.172	[M+H]	322.4
107	C17H18FN5	311.1546	[M+H]	312.1
108	C20H25N5	335.21 1	[M+H]	336.2
109	C17H23N5	297.1953	[M+H]	298.2
110	C16H21N5O	299.1746	[M+H]	300.2
111	C17HI8FN5	311.1546	[M+H]	312.1
112	C16H21N5O	299.1746	[M+H]	300.2
113	C21H24N6	360.2062	[M+H]	361.3
114	C20H23N5	333.1953	[M+H]	334.3
115	C17H25N5	299.211	[M+H]	300.1
116	C15H19N7	297.1702	[M+H]	298.2
117	C16H23N5	285.1953	[M+H]	286.2
118	C19H19N5	317.164	[M+H]	318
119	C19H25N5	323.211	[M+H]	324.1
120	C16H19N5	281.164	[M+H]	282.1
121	C20H25N5	335.211	[M+H]	336.1
122	C18H27N5	313.2266	[M+H]	314.1
123	C15H21N5	271.1797	[M+H] |	272.1

Compound	MF	Mass	[M+XJ	Mass Found
124	C15H21N5O	287.1746	[M+H]	288.3
125	C17H23F2N5	335.1922	[M+H]	336.1
126	C15H19N5O	285.159	[M+H]	286
127	C13H17C1N6O	308.1152	[M+H]	308.9
128	C13H18N6O	274.1542	[M+H]	275.3
129	C15H23N5	273.1953	[M+H]	274.4
130	C15H23N5	273.1953	[M+H]	273.7
131	C14H20CIN5	293.1407	[M+H]	293.8
132	C13H19N5	245.164	[M+H]	245.9
133	CI5H21N5	271.18	[M+H]	271.9
134	C14H19N5	257.164	[M+H]	257.7
135	C14H21N5	259.1797	[M+H]	260.1
136	C14H20C1N5	293.1407	[M+H]	293.8
137	C15H21N5O	287.1746	[M+H]	288.3
138	C13H19N5	245.164	[M+H]	246.4
139	C14H21N5	259.1797	[M+H]	260
140	C18H27N5O2	345.2165	[M+H]	346.3
141	C22H27N5O2	393.2165	[M+H]	394.5
142	C15H19N5O	285.159	[M+H]	285.8
143	C16H23N5O	301.1903	[M+H]	301.8
144	C15H23N5	273.1953	[M+H]	273.9
145	C16H23N5O2	317.1852	[M+H]	318
146	C16H23N5O	301.1903	[M+H]	301.8
147	C16H24N6O	316.2012	[M+HJ	317
148	C22H29N7O	407.2434	[M+H]	408
149	C21H28N6	364.2375	[M+H]	265.4
150	C17H27N5O	317.2216	[M+H]	318.1
151	C15H22N6O2	318.1804	[M+H]	319
152	C18H19N5O2	337.1539	[M+H]	338
153	C20H33N5O	359.2685	[M+H]	360
154	C17H24N8	340.2124	[M+H]	340.9
155	C15H20N6	284.1749	[M+H]	285.2
156	C16H24N6	300.2062	[M+H]	301
157	C15H23N7	301.2015	[M+H]	302
158	C22H26C1N5O2	427.1775	[M+H]	428.2
159	C18H26C1N5O2	379.1775	[M+H]	380.2
160	C21H21N5	343.1797	[M+H]	344.5
161	C19H23N5O	337.1903	[M+H]	338.3
162	C21H22N6	358.1906	[M+H]	359.3
163	C15H22N6O	302.1855	[M+H]	303.2
164	C18H21N5	307.1797	[M+H]	308.3
165	C18H20C1N5O	357.1356	[M+H]	357.8
166	C17H18C1N5	327.1251	[M+H]	327.8
167	C15H22C1N5	307.1564	[M+H]	307.8
168	C12H17N5O	247.1433	[M+H]	248.1
169	CI6H19N7O	325.1651	[M+H]	326.3

Compound	MF	Mass	[M+X]	Mass Found
170	C15H24N6	288.2062	[M+H]	288.9
171	C21H22N6	358.1906	[M+H]	358.9
172	C21H21N5	343.1797	[M+H]	344.1
173	C17H17CI2N5	361.0861	[M+H]	362.3
174	C15H19N7	297.1702	[M+H]	298.2
175	C22H25N5O2	391.2008	[M+H]	392
176	C19H29N5O2	359.2321	[M+H]	360.3
177	C15H24N6	288.2062	[M+H]	289.3
178	C23H29N5O2	407.2321	[M+H]	408.4
179	C14H22N6O	290.1855	[M+H]	291.2
180	C16H21N5O2	315.1695	[M+H]	316.3
181	C14H22N6O	290.1855	[M+H]	291.5
182	C13H19N5O	261.159	[M+H]	262.3
183	C14H22N6	274.1906	[M+H]	275.2
184	C16H25N5	287.211	[M+H]	288.3
185	C18H21N5O	323.1746	[M+H]	324
186	C18H21N5O	323.1746	[M+H]	324.2
187	C18H21N5O	323.1746	[M+H]	324.4
188	C19H24N6	336.2062	[M+H]	337.4
189	C13H20N6	260.1749	[M+H]	261.3
190	C16H25N7	315.2171	[M+H]	316.4
191	C17H20N6	308.1749	[M+H]	309.1
192	C17H20N6O	324.1699	[M+H]	325.1
193	C16H23N7O	329.1964	[M+H]	330.3
194	C16H19N7	309.1702	[M+H]	310.3
195	C18H18C1N5O2	371.1149	[M+H]	372.3
196	C19H23N5	321.1953	[M+H]	322.4
197	C16H24C1N5	321.172	[M+H]	322.4
198	C17H18FN5	311.1546	[M+H]	312.1
199	C20H25N5	335.211	[M+H]	336.2
200	C17H23N5	297.1953	[M+H]	298.2
201	CI6H21N5O	299.1746	[M+H]	300.2
202	C17H18FN5	311.1546	[M+H]	312.1
203	C16H21N5O	299.1746	[M+H]	300.2
204	C2IH24N6	360.2062	[M+H]	361.3
205	C20H23N5	333.1953	[M+H]	334.3
206	C17H25N5	299.211	[M+H]	300.1
207	C15H19N7	297.1702	[M+H]	298.2
208	C16H23N5	285.1953	[M+H]	286.2

Example 36: CDK9/cyclin Tl inhibition

Exemplary compounds of the invention (1-34) were tested for inhibition of CDK9/cyclin Tl. Using a radiometric assay (reaction time 60 minutes) the compounds were tested in IO-dose

IC50 duplicate mode with a 3-fold serial dilution starting at 10 μΜ. A control compound (Staurosporine) was tested in 10-dose IC50 mode with 3-fold serial dilution starting at 10 μΜ. Reactions were carried out at 10 μΜ ATP. Results for the tested compounds are shown in the Table below. Data was normalized to positive and négative Controls and curve fits were executed with GraphPad software and were performed where the enzyme activîties at the highest concentration of compounds were less than 65%.

Exemplary compounds of the invention (35-126) were tested for inhibition of CDK9/cyclin Tl kinase. Using a LANCE assay (reaction time 60 minutes), the compounds were tested in 10-dose IC50 duplicate mode with a 3-fold serial dilution starting at 4.3 μΜ. A 10 control compound (SNS-032) was tested in 10-dose IC50 mode with 3-fold serial dilution starting at 10 μΜ. Reactions were carried out at 10 μΜ ATP. Results for the tested compounds are shown in the Table below. Data was normalized to positive and négative Controls and curve fits were analyzed with XLFIT5 as % inhibition vs. log [compound concentration] using a 4parameter logistic model205. Fit = (A+((B-A)/(l+((C/x)^AD)))); Res = (y-fit).

Table: Results of cyclin TI inhibition

Potency grade: A = 1—500 nM; B = 501 —1000 nM; C > 1001 nM

Compound	CDK9/cyclin Tl Potency Grade
1	C
2	A
3	A
4	B
5	A
6	A
7	A
8	A
9	A
10	C
11	B
12	B
13	A
14	A
15	A
16	A
17	A
18	A
19	A
20	A
21	A
22	A
23	A
24	-___B_____

Compound	CDK9/cyclin Tl Potency Grade
25	B
26	N/A
27	N/A
28	A
29	A
30	C
31	A
32	A
33	A
34	C
35	A
36	A
37	C
38	N/A
39	A
40	A
41	A
42	A
43	A
44	C
45	A
46	A
47	A
48	A
49	A
50	A
51	A
52	A
53	A
54	A
55	A
56	A
57	A
58	A
59	A
60	A
61	A
62	A
63	A
64	A
65	A
66	A
67	A
68	B
69	B
70	A
71	A
72	__________A___________

Compound	CDK9/cyclin Tl Potency Grade
73	A
74	A
75	A
76	A
77	B
78	B
79	A
80	A
81	A
82	A
83	A
84	C
85	A
86	A
87	A
88	B
89	C
90	A
91	B
92	C
93	A
94	A
95	A
96	A
97	A
98	A
99	C
100	A
101	A
102	A
103	B
104	A
105	A
106	A
107	A
108	A
109	A
110	B
111	A
112	A
113	A
114	A
115	A
116	A
117	A
118	A
119	A
120	__________A__________

Compound	CDK9/cyclîn Tl Potency Grade
121	A
122	A
123	A
124	B
125	A
126	B
127	A
128	C
129	N/A
130	A
131	A
132	A
133	A
134	A
135	C
136	A
137	A
138	A
139	A
140	A
141	A
142	A
143	A
144	A
145	A
146	A
147	A
148	A
149	A
150	A
151	A
152	A
153	A
154	A
155	A
156	A
157	A
158	A
159	B
160	B
161	A
162	A
163	A
164	A
165	A
166	A
167	A
168	__B____________

Compound	CDK9/cyclin Tl Potency Grade
169	B
170	A
171	A
172	A
173	A
174	A
175	C
176	A
177	A
178	A
179	B
180	C
181	A
182	B
183	C
184	A
185	A
186	A
187	A
188	A
189	A
190	C
191	A
192	A
193	A
194	B
195	A
196	A
197	A
198	A
199	A
200	A
201	B
202	A
203	A
204	A
205	A
206	A
207	A
208	A
Staurosporine	A

The test data shows that the compounds disclosed herein are effective as CDK.9 inhibitors and would be suitable candidates for therapy relating to CDK9-mediated disorders.

The contents of ail référencés, pending patent applications and published patents, cited throughout this application are hereby expressly încorporated by référencé.

The foregoing embodiments are presented by way of example only. A person of ordinary skill in the relevant field would understand that various modifications may be made without deviating from the spirit and scope of the présent invention.

Claims (11)

1. A compound of formula (I)

or a pharmaceutically acceptable sait thereof, wherein:

R¹ is C1-C6 alkyl, C3-C6 cycloalkyl, tetrahydrofuranyl, or tetrahydropyranyl, optionally substituted at any position with one or more of D, halo, R⁷CO2R⁸, CO2R⁸, CO?H, R⁷CO2H, NH2, NHR⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH2, SO2NHR⁸, or R¹ is NH2, NHR⁸, OH, OR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, SO₂NHR⁸, or R and R together form a fused C5-C6 cycloaryl, optionally substituted at any position with one or more of D, halo, NH2, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO3R⁸, SO₂NH₂, or SO₂NHR⁸;

R², R³, R⁴ and R⁵ are independently H, D, halo, or C1-C5 alkyl or C3-C6 cycloalkyl optionally substituted at any position with one or more of D, halo, NH2, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH2, or SO2NHR⁸, or R³ and R⁴ together form a fused C5-C6 cycloaryl, optionally substituted at any position with one or more of D, halo, NH2, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸, or R and R together form a fused C5-C6 cycloaryl, optionally substituted at any position with one or more of D, halo, NH₂, NHR⁸, NR⁷R⁸, OH, OR⁸, SH, SR⁸, NHCOR⁸, NHSO2R⁸, SO2NH₂, or SO₂NHR⁸;

R⁶ is H or D;

R is (CH₂)_n wherein n is an integer from 1 to 6; and

R is C1-C6 alkyl or C3-C6 cycloalkyl, optionally substituted at any position with one or more of D, halo, OH, SH, or NH₂.

2. The compound or sait thereof of claim 1, wherein

R¹ is C1-C6 alkyl or C3-C6 cycloalkyl, optionally substituted at any position with NH₂, or

R¹ is NHCOR⁸;

R², R³, R⁴, and R⁶ are H;

R⁵ is cyclobutyl, cyclopentyl, or cyclohexyl, optionally substituted at any position with D, NH2, OH, NHR⁸, OR⁸, or combinations thereof; and

R⁸ isCl-C4 alkyl.

3. The compound or sait thereof of daim 1, wherein

R¹ is methyl, ethyl, isopropyl, sec-butyl, 3-pentyl, cyclopropyl, cyclopentyl, or NHCOCH3;

R², R³, R⁴, and R⁶ are H; and

R⁵ is cyclobutyl, cyclopentyl, or cyclohexyl, optionally substituted at any position with NH₂. 10

4. A compound of one of the following structures or a pharmaceuticaliy acceptable sait thereof:

25 26 27

28 29 30

5. The compound or sait of any one of claims 1 to 4, wherein the compound or sait is a racemic or non-racemic mixture of optically active stereoisomers.

5

6. The compound or sait of any one of claims 1 to 4, wherein the compound or sait is a substantially pure single enantiomer or diastereomer.

7. The compound or sait of any one of claims 1 to 6 as a dichloride sait.

10

8. A composition comprising a compound or sait of any one of claims 1 to 7 and a pharmaceutically acceptable carrier.

9. A method of treating a CDK9-mediated disease, comprising administering to a subject in need thereof a compound according to any of claims 1 to 7 or a composition according to claim 8.

10. The method of claim 9, wherein the disease is a cancer caused by aberrant expression of MYCor MCL-1, a hématologie malignancy, or a solid tumor.

11. The method of claim 9, wherein the disease is acute myelogenous leukemia, primary peritoneal carcinoma, chronic lymphocytic leukemia, relapsed multiple myeloma, non-Hodgkin’s lymphoma, acute lymphoblastic leukemia, acute byphenotypîc leukemia, advanced breast cancer, non-small cell 5 lung cancer, or liver cancer.