WO2008113095A1

WO2008113095A1 - Compositions and therapies comprising tranilast compounds and angiotensin-converting enzyme (ace) inhibitors and/or angiotensin receptor blockers (arb)

Info

Publication number: WO2008113095A1
Application number: PCT/AU2007/000343
Authority: WO
Inventors: Darren James Kelly; Richard Ernest Gilbert
Original assignee: Fibrotech Therapeutics Pty Ltd
Priority date: 2007-03-20
Filing date: 2007-03-20
Publication date: 2008-09-25

Abstract

The invention relates generally to compositions, methods and uses of at least one tranilast compound and at least one angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker.

Description

COMPOSITIONS AND THERAPIES COMPRISING TRANILAST

COMPOUNDS AND ANGIOTENSIN-CONVERTING ENZYME (ACE)

INHIBITORS AND/OR ANGIOTENSIN RECEPTOR BLOCKERS (ARB)

Field of the invention

The invention relates generally to combinations, compositions, conjugates, and methods including at least one tranilast compound and at least one angiotensin- converting enzyme (ACE) inhibitor and/or angiotensin receptor blocker (ARB).

Background of the invention

Fibrosis is a condition which is characterized by excessive scarring which is thought to be the result of the normal wound healing response gone awry. The causes of fibrosis are diverse, and include trauma, surgery, infection, and exposure to toxins such as environmental pollutants and alcohol. Fibrotic conditions share the common characteristic of excessive collagen accumulation and an associated loss of function with the replacement or displacement of normal tissue by fibrotic tissue. Organs that are commonly affected by fibrosis include the liver, kidney and lung. Fibrosis is associated with a number of disease states such as diabetes, obesity, hepatic cirrhosis, pulmonary interstitial fibrosis, kidney disease (eg glomerulonephritis, diabetic nephropathy, progressive kidney disease), cardiovascular disease (eg heart failure due to ischaemic heart disease, valvular heart disease, hypertensive heart disease), scleroderma, excessive scar tissue post surgery or device insertion, and tumor progression and metastasis.

Studies conducted over more than a decade have consistently indicated a major role for the prosclerotic growth factor, transforming growth factor-β, in organ fibrosis. The beneficial effects of tranilast (n-[3,4-dimethoxycinnamoyl] anthranilic acid) as an inhibitor of transforming growth factor-β (TGF-β) has been reported in a range of diseases that are characterized by fibrosis.

Kidney disease is a common and increasing public health problem in the US, Europe, and in most countries where reliable epidemiological data are available. For instance, in the US, approximately eight million people have impaired renal function (calculated glomerular filtration rate <60 ml/min/1.73m²⁾, with a further 300,000 individuals in end- stage renal failure.

Almost regardless of the insult, an initial critical nephron loss is frequently followed by progressive proteinuria and declining glomerular filtration rate, with an inexorable progression to end-stage renal disease. Current renoprotective therapy, aimed at slowing this progression, centres on blood pressure control and blockade of the renin- angiotensin system, for which data from controlled clinical trials provide a strong evidence base for treatment. Treatment regimes typically include administration of angiotensin-converting enzyme (ACE) inhibitors and/or angiotensin receptor blockers (ARBs). However, despite these therapies, a considerable proportion of patients with chronic kidney disease continue to progress and new therapies are needed.

Summary of the invention

The present invention provides a pharmaceutical composition including at least one tranilast compound and at least one ACE inhibitor and/or ARB. The invention also provides a method of using the above composition for the prevention and/or treatment of a disease disclosed herein. The invention further provides a combination therapy for prevention and/or treatment of a disease disclosed herein including administering to a subject in need thereof a therapeutically effective amount of at least one tranilast compound and at least one ACE inhibitor and/or ARB. In aspects of the invention, a composition, method or combination therapy of the invention is used in the treatment of a kidney disease and related diseases, disorders, or conditions as well as to prevent and/or treat complications associated with kidney disease, and related diseases, disorders, or conditions.

The compositions of this invention, as well as the combination of compounds used in the methods of this invention, may demonstrate a different pharmacologic, therapeutic and/or metabolic profile than each compound alone. Therefore, compositions and methods of this invention and the compounds used therein may offer advantages as therapeutics for a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease. These advantages may include increased overall therapeutic benefit and reduction in deleterious side effects. Other advantages may include reduced frequency of administration of a tranilast compound and/or an ACE inhibitor and/or an ARB.

In a first aspect the present invention provides a pharmaceutical composition including at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB. Preferably, the pharmaceutical composition includes N-[3,4-dimethoxycinnamoyl]anthranilic acid or a pharmaceutically acceptable salt or solvate thereof and perindopril or a pharmaceutically acceptable salt or solvate thereof.

The pharmaceutical composition is useful for the treatment of a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease or a cardiovascular disease.

In an embodiment, a pharmaceutical composition of the invention includes additive amounts or synergistic amounts, of at least one tranilast compound and at least one ACE inhibitor and/or ARB. Such a composition includes sufficient amounts of each component to achieve a desired result that is greater than the result achieved with each component on its own. In a particular embodiment of the invention, a pharmaceutical composition is provided including a combination of at least one tranilast compound and at least one ACE inhibitor and/or ARB effective to exert a synergistic effect in preventing and/or treating a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease.

In an embodiment, a pharmaceutical composition is provided including a therapeutically effective suboptimal dosage of at least one tranilast compound and at least one ACE inhibitor and/or ARB that is more effective at slowing, reducing or preventing fibrosis following treatment compared with a dosage of any one of the compounds alone.

In another embodiment, a pharmaceutical composition is provided including therapeutically effective suboptimal amounts of at least one tranilast compound and at least one ACE inhibitor and/or ARB in a form for chronic or acute therapy of a disease. In particular embodiments, the above described compositions include at least one tranilast compound and at least one ACE inhibitor and/or ARB, one or more of which are in doses that are at least 1 to 10 fold, 2 to 10 fold, or 5 to 10 fold lower than the respective doses of each component required to treat a disease disclosed herein when administered individually, in particular a fibrotic condition, more particularly a kidney disease.

In a further aspect, the present invention provides a method of treating a disease as disclosed herein including administering to a patient a therapeutically effective amount of a tranilast compound or a pharmaceutically acceptable salt or solvate thereof and a therapeutically effective amount of an ACE inhibitor and/or ARB. In particular, the disease is a fibrotic condition, and more particularly a kidney disease or a cardiovascular disease.

The tranilast compound and the ACE inhibitor and/or ARB may be administered simultaneously or sequentially.

The method may be performed by administering a composition as described above. Further, the therapeutically effective dose of the ACE inhibitor and/or ARB may be less than the therapeutic dose range of the ACE inhibitor and/or ARB when administered alone.

In a further embodiment, the present invention provides a method of treating a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease, including administering at least one tranilast compound at a frequency of: more than once daily, daily, more than once weekly, weekly, or more than once monthly and monthly, and administering at least one ACE inhibitor and/or ARB at a frequency of: more than once daily, daily, more than once weekly, weekly, more than once monthly, or monthly.

The invention provides in some aspects methods for the potentiation of a tranilast compound in the treatment of a disease in a subject, in particular a fibrotic condition and related diseases, disorders, or conditions, including co-administering at least one ACE inhibitor and/or ARB to the subject.

In a further aspect, the invention provides a method of treating a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease, in a patient that has had a sub-optimal response to treatment with an ACE inhibitor and/or ARB, including administering either simultaneously or sequentially to the patient an effective amount of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB, to provide an enhanced or optimal response.

The invention contemplates a method of retreatment using a therapeutically effective amount of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB for patients suffering from a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease, who fail to respond to therapy with the ACE inhibitor and/or ARB, or who, following cessation of such therapy, suffer a relapse or who relapse while on therapy.

The invention includes combination treatments providing synergistic activity or delivering synergistically effective amounts of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB.

In particular aspects of the invention, the disease is a progressive kidney disease. Thus, the invention provides a method for treating a progressive kidney disease in a patient including the step of administrating to the patient any of the compositions and combinations described above.

In a further aspect, the present invention provides a method for improving the efficacy of an ACE inhibitor and/or ARB including administering, either sequentially or simultaneously with the administration of the ACE inhibitor and/or ARB, a tranilast compound or a pharmaceutically acceptable salt or solvate thereof. In a further aspect the present invention provides the use of the composition described above for treating a disease as disclosed herein, particularly a fibrotic condition, and more particularly a kidney disease or a cardiovascular disease.

In yet another aspect the present invention provides the use of a tranilast compound or a pharmaceutically acceptable salt or solvate thereof and an ACE inhibitor and/or ARB for treating a disease as disclosed herein, particularly a fibrotic condition, and more particularly a kidney disease or a cardiovascular disease.

In yet another aspect the present invention provides the use of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB in the preparation of a medicament for the treatment of a disease as disclosed herein, particularly a fibrotic condition and more particularly a kidney disease, especially a progressive kidney disease, or a cardiovascular disease.

In an embodiment, the invention relates to synergistically effective amounts of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one ACE inhibitor and/or ARB in the preparation of a medicament for preventing and/or treating a disease disclosed herein, in particular a fibrotic condition, and more particularly a kidney disease or a cardiovascular disease.

In a further aspect, the present invention provides a kit for use in treating a fibrotic condition including at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof; at least one ACE inhibitor and/or ARB; and instructions and/or labelling for use in treating a fibrotic condition.

Encompassed within embodiments of this invention are any of the above treatments and compositions wherein the dose of tranilast compound is between about 0.1 to 1000 mg/kg/day or between about 10 and 1000 mg/kg/day, in particular between about 10 and 500 mg/kg/day, more particularly between about 50 to 500 mg/kg/day, between about 100 to 400 mg/kg/day or between about 100 to 300 mg/kg/day.

These and other aspects, features, and advantages of the present invention should be apparent to those skilled in the art from the following detailed description.

Brief description of the drawings

The invention will be better understood with reference to the drawings in which:

Figure 1. Glomerulosclerotic Index (upper panel) and tubulointerstitial fibrosis (lower panel) in sham and STNx rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. Data are presented as mean±s.e.m. *P<0.01 compared with shams, ^tP<0.01 versus untreated STNx rat kidneys. *P<0.05 versus STNx + tranilast. *P<0.05 versus monotherapy.

Figure 2. Representative photomicrographs of periodic acid Schiff-stained sections from Sham, STNX rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. In (a) sham rats there is no glomerulosclerosis, while (b) STNx is associated with a dramatic increase in glomerulosclerosis. Treatment of STNx rats with (c) tranilast and (d) perindopril was associated with a reduction in extent of glomerulosclerosis, (e)

Combination therapy with tranilast and perindopril was associated with less glomerulosclerosis than monotherapy. Original magnification x360.

Figure 3. Representative trichrome-stained sections showing tubulointerstitial fibrosis and atrophy in Sham, STNx rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. In (a) sham, there is no cortical tubular fibrosis or atrophy, while (b) STNx is associated with an increase in interstitial fibrosis (blue) and atrophic tubules (arrows). Treatment of STNx rats with (c) tranilast, (d) perindopril, and (e) combination therapy was associated with a reduction in both tubular fibrosis and atrophy. Original magnification x380.

Figure 4. Quantitation of atrophic tubules in rat kidneys from sham, STNx rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. Data are presented as mean±s.e.m. *P<0.01 compared with shams, ^tP<0.01 versus untreated STNx rat kidneys. *P<0.05 versus STNx +tranilast. ^#P<0.05 versus monotherapy.

Figure 5. Quantitation of tubulointerstitial (upper panel) and glomerular phospho-Smad 2 (lower panel) immunostaining in rat kidneys from sham, STNx rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. *P<0.01 compared with shams, ^tP<0.01 versus untreated STNx rat kidneys. *P<0.05 versus STNx +Tran. ^#P<0.05 versus monotherapy.

Figure 6. Representative photomicrographs of (a-e) glomeruli and (f-j) tubulointerstitium immunostained for phospho-Smad 2 in Sham, STNx rats treated with tranilast, perindopril, or a combination of tranilast and perindopril. In (a, f) sham there is little evidence of phospho-Smad 2, while (b, g) STNx is associated with an increase immunostaining for phospho-Smad 2. (c, h) Treatment of STNx rats with tranilast, (d, i) perindopril, and (e, j) combination therapy was associated with an incremental reduction in phospho-Smad 2 immunostaining in the tubular epithelium. Original magnification x380.

Figure 7. In vitro effects of tranilast on TGF-β induced 3H-proline incorporation in cultured rat mesangial cells. Values are expressed as mean7s.e.m. *P<0.01 versus cells grown in control medium, ^fP<0.01 versus TGF-β-treated cells.

Detailed description of the embodiments

As used throughout the disclosure, the following terms, unless otherwise indicated, shall be understood to have the following meanings:

Numerical ranges recited herein by endpoints include all numbers and fractions subsumed within that range (e.g. 1 to 5 includes 1 , 1.5, 2, 2.75, 3, 3.90, 4, and 5). It is also to be understood that all numbers and fractions thereof are presumed to be modified by the term "about." The term "about" means plus or minus 0.1 to 50%, 5- 50%, or 10-40%, preferably 10-20%, more preferably 10% or 15%, of the number to which reference is being made.

Further, it is to be understood that "a," "an," and "the" include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to a composition containing "a tranilast compound" includes a mixture of two or more compounds.

A "tranilast compound" includes tranilast (N-(3,4-dimethoxycinnamoyl)anthranilic acid) and derivatives thereof, such as those disclosed in US patent no 3,940,422. In particular, a tranilast compound is a compound of the formula

where Ri and R₂ are the same or different and are selected from the group consisting of optionally substituted alkyl, in particular Ci to C₁₀ alkyl, optionally substituted alkynyl, in particular C₂ to Ci₀ alkynyl, and a chain including an optionally substituted triazole; Xi and X₂ are the same or different and are selected from O and S; and R3 and R₄ are the same or different and are selected from the group consisting of OH, hydrogen, halogen, optionally substituted alkyl, in particular Ci to C₁₀ alkyl, and optionally substituted alkynyl, in particular a C₂ to C₁₀ alkynyl.

In certain embodiment of the invention, a tranilast compound is a compound of the formula I wherein one of R₁ and R₂ is optionally substituted methyl or ethyl and the other of R₁ and R₂ is optionally substituted alkynyl, in particular a C₂ to C₁O alkynyl, or a chain including an optionally substituted triazole. The alkynyl may be a terminal or nonterminal alkynyl. In one embodiment, the triazole is a 1 ,4-disubstituted 1 ,2,3-triazole.

In certain embodiments of the invention, a tranilast compound is a compound of the formula Il

wherein R₅ is optionally substituted methyl, ethyl, propyl, hydroxyl or halogen; and R₆ is an optionally substituted alkynyl, in particular a C₂ to C₁₀ alkynyl or a chain containing an optionally substituted triazole. In a more particular embodiment, R₆ is propargyl or a

1 ,4-disubstituted 1 ,2,3-triazole. In a preferred embodiment, R₅ is methyl and R₆ is propargyl.

In a further embodiment, a tranilast compound has the formula

wherein n is an integer between 1 and 10, preferably 1 and 6; and R is hydrogen or Ci to Cio alkyl.

In a further embodiment, a tranilast compound has the formula IV

wherein n is an integer between 1 and 10, preferably 1 and 6; and R is hydrogen or optionally substituted Ci to C-io alkyl.

In a further embodiment, a tranilast compound has the formula V

wherein n is an integer between 1 and 10, preferably 1 and 6; and R is hydrogen or any one of the following optionally substituted groups, alkyl, in particular Ci to C-io alkyl, aryl, arylalkyl, alkenyl, alkynyl, haloalkyl, haloalkenyl, heteroalkyl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, heteroaryl, cycloalkylalkyl, heterocycloalkylalkyl, arylalkyl, heteroarylalkyl, arylalkenyl, cycloalkylheteroalkyl, arylheteroalkyl, heterocycloalkylheteroalkyl, heteroarylheteroalkyl, alkylamido, aminoalkyl, acylamino and arylamido.

In a further embodiment, a tranilast compound has the formula Vl

wherein n is an integer between 1 and 10, preferably 1 and 6; and R is hydrogen, or any one of the following optionally substituted groups, alkyl, in particular Ci to Cio alkyl, aryl, arylalkyl, alkenyl, alkynyl, haloalkyl, haloalkenyl, heteroalkyl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl, heteroaryl, cycloalkylalkyl, heterocycloalkylalkyl, arylalkyl, heteroarylalkyl, arylalkenyl, cycloalkylheteroalkyl, arylheteroalkyl, heterocycloalkylheteroalkyl, heteroarylheteroalkyl, alkylamido, aminoalkyl, acylamino and arylamido.

A tranilast compound also includes a prodrug of the compounds described above. As used herein, the term "prodrug" refers to a tranilast compound including structural modifications thereto, such that in vivo the prodrug is converted, for example, by hydrolytic, oxidative, reductive, or enzymatic cleavage into a parent compound (e.g., active compound or active derivative or analogue thereof). The term includes bioreversible derivatives of drug molecules used to overcome some barriers to the utility of the parent drug molecule. Examples of barriers include, without limitation, solubility, permeability, stability, presystemic metabolism and targeting limitations (J. Stella, "Prodrugs as therapeutics", Expert Opin. Ther. Patents, 14(3), 277-280, 2004). Prodrugs may be, for example, metabolically labile mono- or di-ester derivatives of a parent compound having a carboxylic acid group.

A tranilast compound also includes a derivative or analog of the compounds described above. "Derivatives" include functional derivatives, chemical derivatives, or variants. A "functional derivative" refers to a compound that possesses an activity (either functional or structural) that is substantially similar to the activity of a described tranilast compound. The term "chemical derivative" describes a molecule that contains additional chemical moieties which are not normally a part of the base molecule. The term "variant" is meant to refer to a molecule substantially similar in structure and/or biological activity to a compound or parts thereof. The term "analog" includes a compound substantially similar in function to a compound described above. An "analog" can include a chemical compound that is structurally similar to another but differs slightly in composition. Differences include without limitation the replacement of an atom or functional group with an atom or functional group of a different element.

Further, a tranilast compound encompasses all possible enantiomers, stereoisomers including the stereoisomerically pure form (e.g., geometrically pure, enantiomerically pure or diastereomerically pure) and enantiomeric and stereoisomeric mixtures and tautomers of the compounds described above.

A tranilast compound also includes an isotopically labelled compound described above, for example, a compound incorporating at least one ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, or ¹⁷O, atom.

The present invention also encompasses the use of a pharmaceutically acceptable salt or solvate of a tranilast compound. Suitable salts include salts that may be formed where acidic protons in the compounds are capable of reacting with inorganic or organic bases. Suitable inorganic salts include those formed with alkali metals, e.g. sodium and potassium, magnesium, calcium, and aluminum. Suitable organic salts include those formed with organic bases such as the amine bases, e.g. ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like. Suitable salts include acid addition salts formed with inorganic acids (e.g. hydrochloric and hydrobromic acids) and organic acids (e.g. acetic acid, citric acid, maleic acid, and the alkane- and arene-sulfonic acids such as methanesulfonic acid and benezenesulfonic acid). When there are two acidic groups present, a pharmaceutically acceptable salt may be a mono-acid-mono-salt or a di-salt; and similarly where there are more than two acidic groups present, some or all of such groups can be salified. Examples of pharmaceutically acceptable salts include salts of pharmaceutically acceptable cations such as sodium, potassium, lithium, calcium, magnesium, ammonium and alkylammonium; acid addition salts of pharmaceutically acceptable inorganic acids such as hydrochloric, orthophosphoric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic and hydrobromic acids; or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, trihalomethanesulphonic, toluenesulphonic, benzenesulphonic, salicylic, sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids.

As used herein "alkyl", either alone or within other terms such as "arylalkyl", means a monovalent, saturated hydrocarbon radical which may be a straight chain (i.e. linear) or a branched chain. In certain aspects of the invention, an alkyl radical includes from about 1 to 24 or 1 to 20 carbon atoms, preferably from about 1 to 10, 1 to 8, 3 to 8, 1 to 6, or 1 to 3 carbon atoms. Examples of alkyl radicals include methyl, ethyl, n-propyl, n- butyl, n-pentyl, n-hexyl, isopropyl, isobutyl, isopentyl, amyl, sec-butyl, tert-butyl, tert- pentyl, n-heptyl, n-octyl, n-nonyl, n-decyl, undecyl, n-dodecyl, n-tetradecyl, pentadecyl, n-hexadecyl, heptadecyl, n-octadecyl, nonadecyl, eicosyl, dosyl, n-tetracosyl, and the like, along with branched variations thereof.

In certain embodiments of the invention an alkyl radical is a C-i-Cβ lower alkyl including or selected from the group consisting of methyl, ethyl, n-propyl, n-butyl, n-pentyl, n- hexyl, isopropyl, isobutyl, isopentyl, amyl, tributyl, sec-butyl, tert-butyl, tert-pentyl, and n- hexyl. Where an alkyl radical is defined as being optionally substituted, the alkyl radical may be substituted with substituents at positions that do not significantly interfere with the preparation of tranilast compounds and that do not significantly reduce the efficacy of the compounds. The alkyl radicals may be substituted with one to five substituents including halo, lower alkoxy, hydroxyl, cyano, nitro, thio, alkenyl, alkynyl, amino, substituted amino, carboxyl, sulfonyl, sulfenyl, sulfinyl, sulfate, sulfoxide, substituted carboxyl, halogenated lower alkyl (e.g. CF3), halogenated lower alkoxy, hydroxycarbonyl, lower alkoxycarbonyl, lower alkylcarbonyloxy, lower alkylcarbonylamino, aryl (e.g., phenylmethyl), heteroaryl (e.g., pyridyl), heterocyclyl (e.g., piperidinyl, morpholinyl) and by groups as described below. The term "alkenyl" refers to an unsaturated, acyclic branched or straight-chain hydrocarbon radical including at least one double bond. Alkenyl radicals may contain from about 2 to 24 or 2 to 10 carbon atoms, preferably from about 3 to 8 carbon atoms and more preferably about 3 to 6 or 2 to 6 carbon atoms. Examples of suitable alkenyl radicals include ethenyl, propenyl such as prop-1-en-1-yl, prop-1-en-2-yl, prop-2-en-1-yl (allyl), prop-2-en-2-yl, buten-1-yl, but-1-en-2-yl, 2-methyl-prop-1-en-1-yl, but-2-en-1-yl, but-2-en-2-yl, buta-1 ,3-dien-1-yl, buta-1 ,3-dien-2-yl, hexen-1-yl, 3-hydroxyhexen-1-yl, hepten-1-yl, and octen-1-yl, and the like. Where an alkenyl radical is defined as being optionally substituted, the alkenyl radical may be substituted with those substituents listed above in relation to the term alkyl or by groups as described below.

The term "alkynyl" refers to an unsaturated, branched or straight-chain hydrocarbon radical including one or more triple bonds. Alkynyl radicals may contain about 1 to 20, 1 to 15, or 2 to 10 carbon atoms, preferably about 3 to 8 carbon atoms and more preferably about 3 to 6 carbon atoms. Examples of suitable alkynyl radicals include ethynyl, such as prop-1-yn-1-yl, prop-2-yn-1-yl (propargyl), butynyls such as but-1-yn-1- yl, but-1-yn-3-yl, but-3-yn-1-yl, pentynyls such as pentyn-1-yl, pentyn-2-yl, 4- methoxypentyn-2-yl, 3-methylbutyn-1-yI, hexynyls such as hexyn-1-yl, hexyn-2-yl, hexyn-3-yl, and 3,3-dimethylbutyn-1-yl radicals and the like. Where an alkynyl radical is defined as being optionally substituted, the alkynyl radical may be substituted with those substituents listed above in relation to the term alkyl or by groups as described below. The term "cycloalkynyl" refers to cyclic alkynyl groups.

The term "alkoxy" refers to a linear or branched oxy-containing radical having an alkyl portion of one to about ten carbon atoms, such as a methoxy radical. Particular alkoxy radicals are "lower alkoxy" radicals having about 1 to 6, 1 to 4 or 1 to 3 carbon atoms. An alkoxy having about 1-6 carbon atoms includes a CrC₆ alkyl-O- radical wherein Cr Cβ alkyl has the meaning set out herein. Illustrative examples of alkoxy radicals include without limitation methoxy, ethoxy, propoxy, butoxy, isopropoxy and tert-butoxy. When defined as being optionally substituted, an "alkoxy" radical may be further substituted with one or more substituents including alkyl atoms (in particular lower alkyl) to provide "alkylalkoxy" radicals; halo, such as fluoro, chloro or bromo, to provide "haloalkoxy" radicals (e.g. fluoromethoxy, chloromethoxy, trifluoromethoxy, difluoromethoxy, trifluoroethoxy, fluoroethoxy, tetrafluoroethoxy, pentafluoroethoxy, and fluoropropoxy) and "haloalkoxyalkyl" radicals (e.g. fluoromethoxymethyl, chloromethoxyethyl, trifluoromethoxymethyl, difluoromethoxyethyl, and trifluoroethoxymethyl).

The term "acyl", alone or in combination, means a carbonyl or thiocarbonyl group bonded to a radical selected from, for example, optionally substituted, hydrido, alkyl

(e.g. haloalkyl), alkenyl, alkynyl, alkoxy ("acyloxy" including acetyloxy, butyryloxy, iso- valeryloxy, phenylacetyloxy, benzoyloxy, p-methoxybenzoyloxy, and substituted acyloxy such as alkoxyalkyl and haloalkoxy), aryl, halo, heterocyclyl, heteroaryl, sulfinyl (e.g. alkylsulfinylalkyl), sulfonyl (e.g. alkylsu lfony lalkyl) , cycloalkyl, cycloalkenyl, thioalkyl, thioaryl, amino (e.g alkylamino or dialkylamino), and aralkoxy. Illustrative examples of

"acyl" radicals are formyl, acetyl, 2-chloroacetyl, 2-bromacetyl, benzoyl, trifluoroacetyl, phthaloyl, malonyl, nicotinyl, and the like.

The term "cycloalkyl" refers to radicals having from about 3 to 16 or 3 to 15 carbon atoms and containing one, two, three, or four rings wherein such rings may be attached in a pendant manner or may be fused, in particular cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, adamantyl, and the like. In certain aspects of the invention the cycloalkyl radicals are "lower cycloalkyl" radicals having from about 3 to 10, 3 to 8, 3 to 6, or 3 to 4 carbon atoms, in particular cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl. The term "cycloalkyl" also embraces radicals where cycloalkyl radicals are fused with aryl radicals or heterocyclyl radicals. Where a cycloalkyl radical is defined as being optionally substituted, the cycloalkyl radical may be substituted with groups as described below.

The term "cycloalkenyl" refers to a radical including about 2 to 16, 4 to 16, 2 to 15, 2 to 10, 4 to 10, 3 to 8, 3 to 6, or 4 to 6 carbon atoms, one or more carbon-carbon double bonds, and one, two, three, or four rings wherein such rings may be attached in a pendant manner or may be fused. In certain aspects of the invention the cycloalkenyl radicals are "lower cycloalkenyl" radicals having three to seven carbon atoms, in particular cyclobutenyl, cyclopentenyl, cyclohexenyl and cycloheptenyl. Where a cycloalkenyl radical is defined as being optionally substituted, the cycloalkyl radical may be substituted with groups as described below. The term "aryl", alone or in combination, refers to a carbocyclic aromatic system containing one, two or three rings wherein such rings may be attached together in a pendant manner or may be fused. The term "fused" means that a second ring is present (i.e., attached or formed) by having two adjacent atoms in common or shared with the first ring. In aspects of the invention an aryl radical has 4 to 24 carbon atoms, in particular 4 to 10, 4 to 8, or 4 to 6 carbon atoms. The term "aryl" includes without limitation aromatic radicals such as phenyl, naphthyl, indenyl, benzocyclooctenyl, benzocycloheptenyl, pentalenyl, azulenyl, tetrahydronaphthyl, indanyl, biphenyl, acephthylenyl, fluorenyl, phenalenyl, phenanthrenyl, and anthracenyl, preferably phenyl. An aryl radical may be optionally substituted with groups as disclosed herein, in particular hydroxyl, alkyl, carbonyl, carboxyl, thiol, amino, and/or halo.

The term "aralkyl" used herein refers to an alkyl group with an aryl substituent, the term "aralkenyl" used herein refers to an alkenyl group with an aryl substituent, and the term "aralkynyl" used herein refers to an alkynyl group with an aryl substituent. Examples of substituted aryl radicals include benzyl, chlorobenyzl, and amino benzyl.

The term "heteroatom" refers to an atom other than carbon, e.g., a nitrogen, oxygen, sulfur or phosphorus atom. A heteroatom-containing radical refers to a molecule or molecular fragment in which one or more carbon atoms is replaced with a heteroatom. Accordingly, the term "heteroalkyl" refers to an alkyl substituent that is heteroatom- containing, the term "heterocyclic" refers to a cyclic substituent that is heteroatom- containing (see below), the term "heteroaryl" refers to an aryl substituent that is heteroatom-containing (see below), and the like.

The term "heteroaryl" refers to fully unsaturated heteroatom-containing ring-shaped aromatic radicals having from 3 to 15, 3 to 10, 5 to 15, 5 to 10, or 5 to 8 ring members selected from carbon, nitrogen, sulfur and oxygen, wherein at least one ring atom is a heteroatom. A heteroaryl radical may contain one, two or three rings and the rings may be attached in a pendant manner or may be fused. Examples of "heteroaryl" radicals, include without limitation, an unsaturated 5 to 6 membered heteromonocyclyl group containing 1 to 4 nitrogen atoms, in particular, pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, triazolyl, tetrazolyl and the like; an unsaturated condensed heterocyclic group containing 1 to 5 nitrogen atoms, in particular, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, indazolyl, benzotriazolyl, tetrazolopyridazinyl and the like; an unsaturated 3 to 6-membered heteromonocyclic group containing an oxygen atom, in particular, 2-furyl, 3-furyl, and the like; an unsaturated 5 to 6-membered heteromonocyclic group containing a sulfur atom, in particular, 2-thienyl, 3-thienyl, and the like; unsaturated 5 to 6-membered heteromonocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, in particular, oxazolyl, isoxazolyl, and oxadiazolyl; an unsaturated condensed heterocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, in particular benzoxazolyl, benzoxadiazolyl and the like; an unsaturated 5 to 6-membered heteromonocyclic group containing 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms, for example, thiazolyl, thiadiazolyl and the like; an unsaturated condensed heterocyclic group containing 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms such as benzothiazolyl, benzothiadiazolyl and the like. The term also includes radicals where heterocyclic radicals are fused with aryl radicals, in particular bicyclic radicals such as benzofuran, benzothiophene, and the like. When defined as being optionally substituted, a heteroaryl radical may be optionally substituted with groups as described below.

The term "heterocyclic" refers to saturated and partially saturated heteroatom- containing ring-shaped radicals having from about 3 to 15, 3 to 10, 5 to 15, 5 to 10, or 3 to 8 ring members selected from carbon, nitrogen, sulfur and oxygen, wherein at least one ring atom is a heteroatom. A heterocylic radical may contain one, two or three rings wherein such rings may be attached in a pendant manner or may be fused. Examples of saturated heterocyclic radicals include without limitation a saturated 3 to 6-membered heteromonocylic group containing 1 to 4 nitrogen atoms [e.g. pyrrolidinyl, imidazolidinyl, piperidinyl, and piperazinyl]; a saturated 3 to 6-membered heteromonocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms [e.g. morpholinyl]; and, a saturated 3 to 6-membered heteromonocyclic group containing 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms [e.g., thiazolidinyl] etc. Examples of partially saturated heterocyclyl radicals include without limitation dihydrothiophene, dihydropyran, dihydrofuran and dihydrothiazole. Illustrative heterocyclic radicals include without limitation 2-pyrrolinyl, 3-pyrrolinyl, pyrrolindinyl, 1 ,3-dioxolanyl, 2H-pyranyl, 4H-pyranyl, piperidinyl, 1 ,4-dioxanyl, morpholinyl, 1 ,4-dithianyl, thiomorpholinyl, and the like. When defined as being optionally substituted, a heterocyclic radical may be optionally substituted with groups as described below.

As used herein, "halo" or "halogen" refers to fluoro, chloro, bromo and iodo, especially fluoro or chloro. The terms "haloalkyl," "haloalkenyl" or "haloalkynyl" refer to an alkyl, alkenyl, or alkynyl group, respectively, in which at least one of the hydrogen atoms in the group has been replaced with a halogen atom.

The term "amino", alone or in combination, refers to a radical where a nitrogen atom (N) is bonded to three substituents being any combination of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkynyl, aryl or silyl with the general chemical formula -NRi₀Rn where R-io and Rn can be any combination of hydrogen, hydroxyl, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, silyl, heteroaryl, or heterocyclic which may or may not be substituted. Optionally one substituent on the nitrogen atom may be a hydroxyl group (- OH) to provide an amine known as a hydroxylamine. Illustrative examples of amino groups are amino (-NH₂), alkylamino, acylamino, cycloamino, acycloalkylamino, arylamino, arylalkylamino, and lower alkylsilylamino, in particular methylamino, ethylamino, dimethylamino, 2-propylamino, butylamino, isobutylamino, cyclopropylamino, benzylamino, allylamino, hydroxylamino, cyclohexylamino, piperidine, benzylamino, diphenylmethylamino, tritylamino, trimethylsilylamino, and dimethyl-tert- butylsilylamino.

"Amido" refers to a functional group containing a carbon atom double-bonded to an oxygen atom and additionally singly bonded to a nitrogen atom [-C(O)-N]. The term includes a primary amide i.e., an unsubstituted amide group [-C(O)-NH₂]; a secondary amide and a tertiary amide, i.e., amides in which nitrogen is substituted with one and two non-hydrogen groups respectively.

When a radical in a tranilast compound may be optionally substituted, one or more substituents apparent to a person skilled in the art may be used, including without limitation alkyl, alkenyl, alkynyl, alkanoyl, alkylene, alkenylene, hydroxyalkyl, haloalkyl, haloalkylene, haloalkenyl, alkoxy, alkenyloxy, alkenyloxyalkyl, alkoxyalkyl, aryl, alkylaryl, haloalkoxy, haloalkenyloxy, heterocyclic, heteroaryl, sulfonyl, sulfenyl, alkylsulfonyl, sulfinyl, alkylsulfinyl, aralkyl, heteroaralkyl, cycloalkyl, cycloalkenyl, cycloalkoxy, cycloalkenyloxy, amino, oxy, halo, azido, thio, cyano, hydroxyl, phosphonato, phosphinato, thioalkyl, alkylamino, arylamino, arylsulfonyl, alkylcarbonyl, arylcarbonyl, heteroarylcarbonyl, heteroarylsulfinyl, heteroarylsulfonyl, heteroarylamino, heteroaryloxy, heteroaryloxylalkyl, arylacetamidoyl, aryloxy, aroyl, aralkanoyl, aralkoxy, aryloxyalkyl, haloaryloxyalkyl, heteroaroyl, heteroaralkanoyl, heteroaralkoxy, heteroaralkoxyalkyl, thioaryl, arylthioalkyl, alkoxyalkyl, and acyl groups.

A tranilast compound may be prepared using reactions and methods generally known to the person of ordinary skill in the art, having regard to that knowledge and the disclosure of this application. For example, tranilast compounds have been synthesized and tested within the context of the treatment of allergies, as described in US 3,940,422. Reactions for producing tranilast compounds are generally performed in a solvent appropriate to the reagents and materials used and suitable for the reactions being effected. It will be understood by those skilled in the art of organic synthesis that the functionality present on the compounds should be consistent with the proposed reaction steps. This will sometimes require modification of the order of the synthetic steps or selection of one particular process scheme over another in order to obtain a desired tranilast compound. It will be recognized that another major consideration in the development of a synthetic route is the selection of the protecting group used for protection of the reactive functional groups present in the compounds described in this invention. An authoritative account describing the many alternatives to the skilled artisan is Greene and Wuts (Protective Groups In Organic Synthesis, Wiley and Sons, 1991).

The starting materials and reagents used in preparing tranilast compounds are either available from commercial suppliers such as the Aldrich Chemical Company (Milwaukee, Wϊs.), Bachem (Torrance, Calif.), Sigma (St. Louis, Mo.), or Lancaster Synthesis Inc. (Windham, N. H.) or are prepared by methods well known to a person of ordinary skill in the art, following procedures described in such references as Fieser and Fieser's Reagents for Organic Synthesis, vols. 1-17, John Wiley and Sons, New York, N.Y., 1991 ; Rodd's Chemistry of Carbon Compounds, vols. 1-5 and supps., Elsevier Science Publishers, 1989; Organic Reactions, vols. 1-40, John Wiley and Sons, New York, N.Y., 1991 ; March J.: Advanced Organic Chemistry, 4th e<±, John Wiley and Sons, New York, N. Y.; and Larock: Comprehensive Organic Transformations, VCH Publishers, New York, 1989.

The starting materials, intermediates, and tranilast compounds may be isolated and purified using conventional techniques, such as precipitation, filtration, distillation, crystallization, chromatography, and the like. Tranilast compounds may be characterized using conventional methods, including physical constants and spectroscopic methods, in particular HPLC.

Tranilast compounds which are basic in nature can form a wide variety of different salts with various inorganic and organic acids. In practice is it desirable to first isolate a tranilast compound from the reaction mixture as a pharmaceutically unacceptable salt and then convert the latter to the free base compound by treatment with an alkaline reagent and subsequently convert the free base to a pharmaceutically acceptable acid addition salt. The acid addition salts of base tranilast compounds are readily prepared by treating the base compound with a substantially equivalent amount of the chosen mineral or organic acid in an aqueous solvent medium or in a suitable organic solvent such as methanol or ethanol. Upon careful evaporation of the solvent, the desired solid salt is obtained.

Tranilast compounds which are acidic in nature are capable of forming base salts with various pharmacologically acceptable cations. These salts may be prepared by conventional techniques by treating the corresponding acidic compounds with an aqueous solution containing the desired pharmacologically acceptable cations and then evaporating the resulting solution to dryness, preferably under reduced pressure. Alternatively, they may be prepared by mixing lower alkanolic solutions of the acidic compounds and the desired alkali metal alkoxide together and then evaporating the resulting solution to dryness in the same manner as before. In either case, stoichiometric quantities of reagents are typically employed to ensure completeness of reaction and maximum product yields. The term "ACE inhibitor" refers to a molecule which decreases the amount or biological activity of angiotensin converting enzyme. In particular an ACE inhibitor includes a molecule that is capable of inhibiting, either partially or completely, the enzyme involved in the conversion of the decapeptide angiotensin I to the active angiotensin Il in the renin-angiotensin system.

Suitable ACE inhibitors include: pharmaceutical compounds that inhibit the amount and/or biological activity of ACE, ACE antisense nucleic acid molecules (such as antisense mRNA, antisense DNA or antisense oligonucleotides), ACE ribozymes, molecules that inhibit the biological activity of ACE (such as anti-ACE antibodies, or a blocking peptide which interacts with the active site of ACE) and molecules that decrease the amount of ACE carried into the aortic valve by plasma low density lipoproteins (LDLs).

Preferably, the ACE inhibitor is a pharmaceutical compound that inhibits ACE and is effective to treat or prevent kidney disease. Suitable pharmaceutical compounds include, without limitation, alacepril, alatriopril, altiopril calcium, ancovenin, benazepril, benazepril hydrochloride, benazeprilat, benzazepril, benzoylcaptopril, captopril, captopril-cysteine, captopril-glutathione, ceranapril, ceranopril, ceronapril, cilazapril, cilazaprilat, converstatin, delapril, deiapril-diacid, enalapril, enalaprilat, enalkiren, enapril, epicaptopril, foroxymithine, fosfenopril, fosenopril, fosenopril sodium, fosinopril, fosinopril sodium, fosinoprilat, fosinoprilic acid, glycopril, hemorphin-4, idapril, imidapril, indolapril, indolaprilat, libenzapril, lisinopril, lyciumin A, lyciumin B, mixanpril, moexipril, moexiprilat, moveltipril, muracein A, muracein B, muracein C, pentopril, perindopril, perindoprilat, pivalopril, pivopril, quinapril, quinapril hydrochloride, quinaprilat, ramipril, ramiprilat, spirapril, spirapril hydrochloride, spiraprilat, spiropril, spiropril hydrochloride, temocapril, temocapril hydrochloride, teprotide, trandolapril, trandolaprilat, utibapril, zabicipril, zabiciprilat, zofenopril and zofenoprilat (See also US Patent Nos. 5,238,924, 4,258,027, 4,374,829, 4,472,380, 4,264,611 , 4,743,450 and 4,344,949, 4,587,258 and 5,061 ,722). Presently, preferred ACE inhibitors that are approved for use in humans and are commercially available include, without limitation, perindopril, ramipril, quinapril, captopril, lisinopril, benazepril, enalapril and fosinopril.

The use of derivatives of known ACE inhibitors is also encompassed by the compositions and methods of the present invention.

An "angiotensin receptor blocker (ARB)" refers to a substance that blocks or reduces the action of angiotensin II. An ARB prevents and/or reverses the effects of angiotensin II, including, without limitation: (1) rapid pressor responses; (2) slow pressor responses; (3) stimulatory effects on the peripheral sympathetic nervous system; (4) CNS effects; (5) release of adrenal catecholamines; (6) secretion of aldosterone; (7) direct and indirect effects of angiotensin Il on the kidneys; (8) growth promoting actions and (9) contraction of vascular smooth muscle [Goodman & Gilman's The Pharmacological Basis of Therapeutics, p. 752 (9^th ed. 1996)]. ARBs including candesartan [e.g., ATACAND® (AstraZeneca)], eprosartan [e.g., TEVETAN® and TEVETAN SB®, (Biovail)], irbesartan [AVAPRO® (Bristol-Myers Squibb)], losartan [(COZAAR® (Merck)], olmesartan [BENICAR® (Sankyo Pharmaceuticals, Inc.)], telmisartan [MICARDIS® (Boehringer Ingelheim)] and valsartan [DIOVAN® (Novartis)], have been approved by the U.S. Food and Drug Administration, and are indicated for the treatment of hypertension [see, for example, Physician's Desk Reference, p. 2068 (56^th ed., 2002)].

ARBs that can be used in the invention include, but are not limited to, candesartan, eprosartan, telmisartan, irbesartan, pratosartan (e.g. from Kotobuki), valsartan, olmesartan, iosartan, tasosartan, embusartan, GA-0113, KRH-594 and UR-7247, or pharmaceutically acceptable prodrugs, salts, or solvates thereof. An ARB can be racemic or enantiomerically pure. The ARB activity of as yet untested compounds can also be readily identified using any methods well-known in the art, including, but not limited to, pressor responses attenuation assays and selective binding assays using an angiotensin receptor (e.g., AT₁ receptor).

In an embodiment of the invention, the ARB is losartan. In another embodiment, the ARB is eprosartan, in particular, TEVATAN® or TEVATAN SB®. In another embodiment, the ARB is valsartan. In another embodiment, the ARB is irbesartan. In another embodiment, the ARB is telmisartan.

"Antiproliferative agent" means a member of a class of compounds for treating proliferative diseases. Any prophylactic or therapeutic agent which is known to be useful, has been used, or is currently being used for the prevention, treatment, management, or amelioration of one or more symptoms associated with a proliferative disorder, such as cancer, can be used in compositions and method of the invention. The compounds include compounds in research, in development and compounds marketed and sold. Examples of anti-proliferative agents include altretamine (hexamethylmelamine, Hexalen), anastrozole (Arimidex), Exemestane (Aromasin), bicalutamide (Casodex), busulfan (Myleran), capecitabine (Xeloda), chlorambucil (Leukeran), cyclophosphamide (Cytoxan), diethylstilbestrol diphosphate (Stilphostrol), estramustine (Emcyt), etoposide (VP-16, Vepesid), flutamide (Eulexin), hydroxyurea (Droxia), Hydrea, Mylocel, letozole (Femara), leucovorin calcium (Leucovorin), levamisole (Ergamisol), lomustine (CCNU, CeeNU), megestrol (Megace), melphalan (Alkeran), mercaptopurine (6-MP, Purinethol), methotrexate (Methotrexate, Rheumatrex), mitotane (Lysodren), nilutamide (Nilandron), procarbazine (Matulane), tamoxifen (Nolvadex), testolactone (Teslac), thioguanine, tretinoin (Vesanoid), mechlorethamine, 5-fluorouracil, cytarabine, gemcitabine, vinblastine, vincristine, vinorelbine, paclitaxel, etoposide, irinotecan, topotecan; leuprolide, flutamide, doxorubicin, bleomycin, epirubicin, mitomycin, interferon compounds, carmustine, lomustine, cisplatin, mitoxantrone (Novantrone). The singular form, "anti-proliferative compound", may mean any one or more compounds from the class of anti-proliferative compounds.

In certain aspects the anti-proliferative agent is an immunomodulatory agent such as a chemotherapeutic agent. In other aspects, the antiproliferative agent is not an immunomodulatory agent. In specific aspects, the anti-proliferative agent is an anti- angiogenic agent. In other aspects, the anti-proiferative agent is not an anti-angiogenic agent. In embodiments of the invention the antiproliferative agent is an anti-cancer agent incuding without limitation acivicin; aclarubicin; acodazole hydrochloride; acronine; adozelesin; aldesleukin; altretamine; ambomycin; ametantrone acetate; aminoglutethimide; amsacrine; anastrozole; anthramycin; asparaginase; asperlin; azacitidine; azetepa; azotomycin; batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bisnafide dimesylate; bisphosphonates (e.g., pamidronate (Aredria), sodium clondronate (Bonefos), zoledronic acid (Zometa), alendronate (Fosamax), etidronate, ibandomate, cimadronate, risedromate, and tiludromate); bizelesin; bleomycin sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin; calusterone; caracemide; carbetimer; carboplatin; carmustine; carubicin hydrochloride; carzelesin; cedefingol; chlorambucil; cirolemycin; cisplatin; cladribine; crisnatol mesylate; cyclophosphamide; cytarabine; dacarbazine; dactinomycin; daunorubicin hydrochloride; decitabine; dexormaplatin; dezaguanine; dezaguanine mesylate; diaziquone; docetaxel; doxorubicin; doxorubicin hydrochloride; droloxifene; droloxifene citrate; dromostanolone propionate; duazomycin; edatrexate; eflomithine hydrochloride; elsamitrucin; enloplatin; enpromate; epipropidine; epirubicin hydrochloride; erbulozole; esorubicin hydrochloride; estramustine; estramustine phosphate sodium; etanidazole; etoposide; etoposide phosphate; etoprine; fadrozole hydrochloride; fazarabine; fenretinide; floxuridine; fludarabine phosphate; fluorouracil; flurocitabine; fosquidone; fostriecin sodium; gemcitabine; gemcitabine hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide; ilmofosine; interleukin-2 (including recombinant interleukin 2, or rlL2), interferon alpha-2a; interferon alpha-2b; interferon alpha-nl; interferon alpha-n3; interferon beta-l a; interferon gamma-l b; iproplatin; irinotecan hydrochloride; lanreotide acetate; letrozole; leuprolide acetate; liarozole hydrochloride; lometrexol sodium; lomustine; losoxantrone hydrochloride; masoprocol; maytansine; mechlorethamine hydrochloride; anti-CD2 antibodies; megestrol acetate; melengestrol acetate; melphalan; menogaril; mercaptopurine; methotrexate; methotrexate sodium; metoprine; meturedepa; mitindomide; mitocarcin; mitocromin; mitogillin; mitomalcin; mitomycin; mitosper; mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole; nogalamycin; ormaplatin; oxisuran; paclitaxel; pegaspargase; peliomycin; pentamustine; peplomycin sulfate; perfosfamide; pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin; plomestane; porfimer sodium; porfiromycin; prednimustine; procarbazine hydrochloride; puromycin; puromycin hydrochloride; pyrazofurin; riboprine; rogletimide; safingol; safingol hydrochloride; semustine; simtrazene; sparfosate sodium; sparsomycin; spirogermanium hydrochloride; spiromustine; spiroplatin; streptonigrin; streptozocin; sulofenur; talisomycin; tecogalan sodium; tegafur; teloxantrone hydrochloride; temoporfin; teniposide; teroxirone; testolactone; thiamiprine; thioguanine; thiotepa; tiazofurin; tirapazamine; toremifene citrate; trestolone acetate; triciribine phosphate; trimetrexate; trimetrexate glucuronate; triptorelin; tubulozole hydrochloride; uracil mustard; uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine sulfate; vindesine; vindesine sulfate; vinepidine sulfate; vinglycinate sulfate; vinleurosine sulfate; vinorelbine tartrate; vinrosidine sulfate; vinzolidine sulfate; vorozole; zeniplatin; zinostatin; and zorubicin hydrochloride.

Other anti-cancer agents include, without limitation: 20-epi-1 ,25 dihydroxyvitamin D3;

5-ethynyluracil; abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox; amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G; antarelix; anti-dorsalizing morphogenetic protein-i; antiandrogen, prostatic carcinoma; antiestrogen; antineoplaston; antisense oligonucleotides; aphidicolin glycinate; apoptosis gene modulators; apoptosis regulators; apurinic acid; ara-CDP-DL-PTBA; arginine deaminase; asulacrine; atamestane; atrimustine; axinastatin 1 ; axinastatin 2; axinastatin

3; azasetron; azatoxin; azatyrosine; baccatin III derivatives; balanol; batimastat;

BCR/ABL antagonists; benzochlorins; benzoylstaurosporine; beta lactam derivatives; beta-aiethine; betaclamycin B; betulinic acid; bFGF inhibitor; bicalutamide; bisantrene; bisaziridinylspermine; bisnafide; bistratene A; bizelesin; breflate; bropirimine; budotitane; buthionine sulfoximine; calcipotriol; calphostin C; camptothecin derivatives; canarypox IL-2; capecitabine; carboxamide-amino-triazole; carboxyamidotriazole;

CaRest M3; CARN 700; cartilage derived inhibitor; carzelesin; casein kinase inhibitors

(ICOS); castanospermine; cecropin B; cetrorelix; chlorlns; chloroquinoxaline sulfonamide; cicaprost; cis-porphyrin; cladribine; clomifene analogues; clotrimazole; collismycin A; collismycin B; combretastatin A4; combretastatin analogue; conagenin; crambescidin 816; crisnatol; cryptophycin; cryptophycin A derivatives; curacin A; cyclopentanthraquinones; cycloplatam; cypemycin; cytarabine ocfosfate; cytolytic factor; cytostatin; dacliximab; decitabine; dehydrodidemnin B; deslorelin; dexamethasone; dexifosfamide; dexrazoxane; dexverapamil; diaziquone; didemnin B; didox; diethylnorspermine; dihydro-5-azacytidine; dihydrotaxol; dioxamycin; diphenyl spiromustine; docetaxel; docosanol; dolasetron; doxifluridine; droloxifene; dronabinol; duocarmycin SA; ebselen; ecomustine; edelfosine; edrecolomab; eflomithine; elemene; emitefur; epirubicin; epothilone A; epothilone B; epristeride; estramustine analogue; estrogen agonists; estrogen antagonists; etanidazole; etoposide phosphate; exemestane; fadrozole; fazarabine; fenretinide; filgrastim; finasteride; flavopiridol; flezelastine; fluasterone; fludarabine; fluorodaunorunicin hydrochloride; forfenimex; formestane; fostriecin; fotemustine; gadolinium texaphyrin; gallium nitrate; galocitabine; ganirelix; gelatinase inhibitors; gemcitabine; glutathione inhibitors; HMG CoA reductase inhibitors (e.g., atorvastatin, cerivastatin, fluvastatin, lescol, lupitor, lovastatin, rosuvastatin, and simvastatin); hepsulfam; heregulin; hexamethylene bisacetamide; hypericin; ibandronic acid; idarubicin; idoxifene; idramantone; ilmofosine; ilomastat; imidazoacridones; imiquimod; immunostimulant peptides; insulin-like growth factor-1 receptor inhibitor; interferon agonists; interferons; interleukins; iobenguane; iododoxorubicin; ipomeanol; iroplact; irsogladine; isobengazole; isohomohalicondrin B; itasetron; jasplakinolide; kahalalide F; lamellarin-N triacetate; lanreotide; leinamycin; lenograstim; lentinan sulfate; leptolstatin; letrozole; leukemia inhibiting factor; leukocyte alpha interferon; leuprolide+estrogen+progesterone; leuprorelin; levamisole; LFA-3TIP

(Biogen, Cambridge, Mass.; U.S. Pat. No. 6,162,432); liarozole; linear polyamine analogue; lipophilic disaccharide peptide; lipophilic platinum compounds; lissoclinamide; lobaplatin; lombricine; lometrexol; lonidamine; losoxantrone; lovastatin; loxoribine; lurtotecan; lutetium texaphyrin; lysofylline; lytic peptides; maitansine; mannostatin A; marimastat; masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinase inhibitors; menogaril; merbarone; meterelin; methioninase; metoclopramide; MIF inhibitor; mifepristone; miltefosine; mirimostim; mismatched double stranded RNA; mitoguazone; mitolactol; mitomycin analogues; mitonafide; mitotoxin fibroblast growth factor-saporin; mitoxantrone; mofarotene; molgramostim; monoclonal antibody, human chorionic gonadotrophin; monophosphoryl lipid A+myobacterium cell wall sk; mopidamol; multiple drug resistance gene inhibitor; multiple tumor suppressor 1 -based therapy; mustard anticancer agent; mycaperoxide B; mycobacterial cell wall extract; myriaporone; N-acetyldinaline; N-substituted benzamides; nafarelin; nagrestip; naloxone+pentazocine; napavin; naphterpin; nartograstim; nedaplatin; nemorubicin; neridronic acid; neutral endopeptidase; nilutamide; nisamycin; nitric oxide modulators; nitroxide antioxidant; nitrullyn; 06-benzylguanine; octreotide; okicenone; oligonucleotides; onapristone; ondansetron; ondansetron; oracin; oral cytokine inducer; ormaplatin; osaterone; oxaliplatin; oxaunomycin; paclitaxel; paclitaxel analogues; paclitaxel derivatives; palauamine; palmitoylrhizoxin; pamidronic acid; panaxytriol; panomifene; parabactin; pazelliptine; pegaspargase; peldesine; pentosan polysulfate sodium; pentostatin; pentrozole; perflubron; perfosfamide; perillyl alcohol; phenazinomycin; phenylacetate; phosphatase inhibitors; picibanil; pilocarpine hydrochloride; pirarubicin; piritrexim; placetin A; piacetin B; plasminogen activator inhibitor; platinum complex; platinum compounds; platinum-triamine complex; porfimer sodium; porfiromycin; prednisone; propyl bis-acridone; prostaglandin J2; proteasome inhibitors; protein A-based immune modulator; protein kinase C inhibitor; protein kinase C inhibitors, microalgal; protein tyrosine phosphatase inhibitors; purine nucleoside phosphorylase inhibitors; purpurins; pyrazoloacridine; pyridoxylated hemoglobin polyoxyethylene conjugate; raf antagonists; raltitrexed; ramosetron; ras famesyl protein transferase inhibitors; ras inhibitors; ras-GAP inhibitor; retelliptine demethylated; rhenium Re 186 etidronate; rhizoxin; ribozymes; RII retinamide; rogletimide; rohitukine; romurtide; roquinimex; rubiginone B1 ; ruboxyl; safingol; saintopin; SarCNU; sarcophytol A; sargramostim; Sdi 1 mimetics; semustine; senescence derived inhibitor 1 ; sense oligonucleotides; signal transduction inhibitors; signal transduction modulators; single chain antigen binding protein; sizofiran; sobuzoxane; sodium borocaptate; sodium phenylacetate; solverol; somatomedin binding protein; sonermin; sparfosic acid; spicamycin D; spiromustine; splenopentin; spongistatin 1 ; squalamine; stem cell inhibitor; stem-cell division inhibitors; stipiamide; stromelysin inhibitors; sulfinosine; superactive vasoactive intestinal peptide antagonist; suradista; suramin; swainsonine; synthetic glycosaminoglycans; tallimustine; 5-fluorouracil; leucovorin; tamoxifen methiodide; tauromustine; tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomerase inhibitors; temoporfin; temozolomide; teniposide; tetrachlorodecaoxide; tetrazomine; thaliblastine; thiocoraline; thrombopoietin; thrombopoietin mimetic; thymalfasin; thymopoietin receptor agonist; thymotrinan; thyroid stimulating hormone; tin ethyl etiopurpurin; tirapazamine; titanocene bichloride; topsentin; toremifene; totipotent stem cell factor; translation inhibitors; tretinoin; triacetyluridine; triciribine; trimetrexate; triptorelin; tropisetron; turosteride; tyrosine kinase inhibitors; tyrphostins; UBC inhibitors; ubenimex; urogenital sinus-derived growth inhibitory factor; urokinase receptor antagonists; vapreotide; variolin B; vector system, erythrocyte gene therapy; thalidomide; velaresol; veramine; verdins; verteporfin; vinorelbine; vinxaltine; vorozole; zanoterone; zeniplatin; zilascorb; and zinostatin stimalamer.

In particular aspects of the invention, the proliferative agent is a paclitaxel compound including without limitation paclitaxel, TAXOTERE®, TAXOL®, Docetaxel, and derivatives, analogues and prodrugs thereof.

An "anti-osteolysis agent" refers to a member of a class of compounds for treating osteolysis. Any prophylactic or therapeutic agent which is known to be useful, has been used, or is currently being used for the prevention, treatment, management, or amelioration of a condition associated with osteolysis can be used in compositions and methods of the invention. The compounds include compounds in research, in development and compounds marketed and sold. Exemplary anti-osteolysis agents include bisphosphonates such as Zoledronic acid (Zometa^®) (Novartis) and Osteoprotegerin.

The term "treating" refers to reversing, alleviating, or inhibiting the progress of a disease, or one or more symptoms of such disease, to which such term applies. Treating includes the management and care of a subject at diagnosis or later. A treatment may be either performed in an acute or chronic way. Depending on the condition of the subject, the term refers to preventing a disease, and includes preventing the onset of a disease, or preventing the symptoms associated with a disease. The term refers to reducing the severity of a disease or symptoms associated with such disease prior to affliction with the disease. "Preventing" refers to preventing the recurrence of a disease or of one or more symptoms associated with such disease. An objective of treatment is to combat the disease and includes administration of the active compounds to prevent or delay the onset of the symptoms or complications, or alleviate the symptoms or complications, or eliminate or partially eliminate the disease. The terms "treatment" and "therapeutically," refer to the act of treating, as "treating" is defined above.

"Combination therapy", "co-administration" and "administering in combination" are used interchangeably herein and mean that the active ingredients are administered concurrently to a patient being treated. When administered in combination each component may be administered at the same time, or sequentially in any order at different points in time. Therefore, each component may be administered separately, but sufficiently close in time to provide the desired effect, in particular a beneficial, complementary, additive, or synergistic effect. The first compound may be administered in a regimen which additionally includes treatment with a second and third compound. In certain embodiments, the term refers to administration of one or more tranilast compounds, and one or more ACE inhibitor and/or ARB to a patient, including separate administration of medicaments each containing one of the compounds as well as simultaneous administration whether or not the compounds are combined in one formulation or whether they are separate formulations.

The terms "subject", "individual", or "patient" are used interchangeably herein and refer to an animal including a warm-blooded animal such as a mammal. Mammal includes without limitation any members of the Mammalia. Suitable mammals include members of the Orders Primates, Rodentia, Lagomorpha, Cetacea, Carnivora, Perissodactyla and Artiodactyla. Members of the Orders Perissodactyla and Artiodactyla are particularly preferred because of their similar biology and economic importance. In general, the terms refer to a human. The terms include domestic animals bred for food or as pets, including horses, cows, sheep, poultry, fish, pigs, cats, dogs, and zoo animals, goats, apes (e.g. gorilla or chimpanzee), and rodents such as rats and mice.

Typical subjects for treatment include persons afflicted with or suspected of having or being pre-disposed to a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease, or persons susceptible to, suffering from or have suffered from such a disease. A subject may or may not have a genetic predisposition for a disease. In particular aspects a subject has symptoms of a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease. In embodiments of the invention, the subjects are susceptible to, or suffer from a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease.

As utilized herein, the term "healthy subject" means a subject, in particular a mammal, having no diagnosed or symptoms of a disease disclosed herein, in particular a fibrotic condition, more particularly a kidney disease.

"Therapeutically effective amount" relates to the amount or dose of a tranilast compound, an ACE inhibitor and/or ARB, or a composition including a tranilast compound and ACE inhibitor and/or ARB₁ that will lead to one or more desired effects, in particular, one or more beneficial effects. A therapeutically effective amount of a substance can vary according to factors such as the disease state, age, sex, and weight of the subject, and the ability of the substance to elicit a desired response in the subject. A dosage regimen may be adjusted to provide the optimum therapeutic response (e.g. sustained beneficial effects). For example, several divided doses may be administered daily or the dose may be proportionally reduced as indicated by the exigencies of the therapeutic situation. A therapeutically effective amount includes a prophylactically effective amount and a synergistically effective amount.

The term "prophylactically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result. Typically, since a prophylactic dose is used in subjects prior to or at an earlier stage of disease, the prophylactically effective amount will be less than an amount which would be used at a later stage in the treatment of the disease.

"Synergistically effective amount" relates to the amount of dose of active compounds or compositions that will provide a synergistic effect, in particular a synergistic beneficial effect.

"Suboptimal dose" or "suboptimal dosage" refers to a dose or dosage of one or more active compounds which is less than the optimal dose or dosage for that compound when used in monotherapy. "Enhanced", "enhancing" or "enhances", used in the context of a composition or method of the invention, means a beneficial effect, an enhanced therapeutic effect, and includes an additive or synergistic effect.

A "beneficial effect" refers to an effect of a combination of compounds or composition described herein including favorable pharmacological and/or therapeutic effects, and improved pharmacokinetic properties and biological activity. The beneficial effect may be a statistically significant effect in terms of statistical analysis of an effect of the two compounds versus the effects of each of the compounds. "Statistically significant" or

"significantly different" effects or levels with two compounds compared with each compound alone may represent levels that are higher or lower than a standard. In embodiments of the invention, the difference may be 1.5, 2, 3, 4, 5, or 6 times higher or lower compared with the effect obtained with each compound alone.

In particular aspects, beneficial effects in the context of a kidney disease refer to enhanced renoprotective effects on the structural and functional manifestations of renal injury, reduction or inhibition of glomerulosclerosis, tubulointerstitial fibrosis and/or tubular atrophy, reduction or inhibition of nuclear phosph-Smad2 in glomerular and/or tubulointerstotium, inhibition or blockage of the renin-angiotensin system, enhanced glomerular filtration rate, reduction or inhibition of TGF-β activity, reduction or inhibition of TGF-β induced collagen production, reduction or elimination of proteinuria, and/or reduction or elimination of hypertension.

In aspects of the invention, the beneficial effect is a "sustained beneficial effect" where the beneficial effect is sustained for a prolonged period of time after termination of treatment. A treatment can be sustained over several years thereby having a major beneficial impact on the severity of the disease and its complications. A beneficial effect may be sustained for a prolonged period of at least about 2 to 4 weeks, 2 to 5 weeks, 3 to 5 weeks, 2 to 6 weeks, 2 to 8 weeks, 2 to 10 weeks, 2 to 12 weeks, 2 to 14 weeks, 2 to 16 weeks, 2 to 20 weeks, 2 to 24 weeks, 2 weeks to 12 months, 2 weeks to 18 months, 2 weeks to 24 months, or several years following treatment. The period of time a beneficial effect is sustained may correlate with the duration and timing of the treatment. A subject may be treated continuously for about or at least about 2 to 4 weeks, 2 to 6 weeks, 2 to 8 weeks, 2 to 10 weeks, 2 to 12 weeks, 2 to 14 weeks, 2 to 16 weeks, 2 weeks to 6 months, 2 weeks to 12 months, 2 weeks to 18 months, or several years, periodically or continuously.

A beneficial effect may be an additive, complementary or synergistic effect.

The terms "additive' or "additive effect" are used to describe an effect that is equal to the sum of the effects of the individual compounds.

The expression "complementary effect" refers to the pharmacological action of one, two, or more different compounds making it possible to act on the same pathology via different pharmacological mechanisms.

The terms "synergistic" or "synergistic effect" are used to describe an effect that is greater than the additive effect which results from the sum of the effects of the individual compounds. A synergistic effect can work through similar or different mechanisms or pathways of action. One potential advantage of a combination therapy with a synergistic effect is that standard dosages may be used for a greater therapeutic effect than expected from the addition of the effect of either compound administered alone; or alternatively lower dosages or reduced frequency of administration of the therapeutic compound(s) may be used to achieve a better therapeutic effect.

A synergistic effect in some fields refers to an effect which is more than additive (e.g., 1+1=3) or less than additive (1+1=1.6). For example, if each of two drugs administered individually slows the growth of cancer cells by 50%, it would not be expected that the two drugs would be combined to completely stop the growth of cancer cells. A synergistic effect may be obtained if the two drugs slow tumor growth by more than either drug alone (preferably decrease cancer cell growth by 50% or more compared to either drug alone) while not causing an unacceptable increase in adverse side effects e.g., increased toxicity.

The term "potentiation" refers to an increase of a corresponding pharmacological activity or therapeutic effect. Potentiation of one component of a combination or composition of the present invention by co-administration of the other components according to the present invention means that an effect is being achieved that is greater than that achieved with one component alone.

A "disease(s)" refers to one or more pathological symptoms or syndromes for which a tranilast compound, ACE inhibitor and/or ARB provide a beneficial effect or therapeutic effect. In aspects of the invention, the disease requires a slowing, reduction or inhibition of fibrosis. In aspects of the invention the disease is a fibrotic condition. In other aspects of the invention the disease is a proliferative disease. In particular aspects of the invention the disease is a kidney disease. In other particular aspects of the invention, the disease is a cardiovascular disease.

A "fibrotic condition" includes a condition or disease involving fibrosis, i.e., excessive collagen accumulation and an associated loss of function with the replacement or displacement of normal tissue by fibrotic tissue. A fibrotic condition includes conditions involving traumatic fibrosis, organ fibrosis, fibrosis due to radiation damage, and fibrosis due to exposure to chemotherapeutics. The term "organ fibrosis" includes without limitation liver fibrosis, fibrosis of the kidneys, pulmonary fibrosis, cardiac fibrosis, and fibrosis of ocular structures. "Traumatic fibrosis" includes without limitation fibrosis secondary to surgery (surgical scarring), accidental physical trauma, burns, and hypertrophic scarring.

Examples of fibrotic conditions include without limitation diabetes, keloids, obesity, hepatic cirrhosis, pulmonary interstitial fibrosis, kidney disease (glomerulonephritis, diabetic nephropathy, progressive kidney disease), cardiovascular disease (e.g. heart failure - ischaemic and non-ischaemic; valvular heart disease; hypertensive heart disease, diabetic cardiomyopathy, hypertension), hypertrophic scars, scleroderma, and excessive scar tissue post surgery or device insertion.

"Kidney disease" generally refers to a disorder of at least one kidney in a subject that compromises the function of the kidney. The kidney disease may result from a primary pathology of the kidney (e.g., injury to the glomerulus or tubule), or another organ (e.g., pancreas) which adversely affects the ability of the kidney to perform biological functions. A kidney disease in a human can be the direct or indirect effect of disease. Examples of a kidney disease as a result or consequence of an indirect effect on the kidneys is kidney disease as a consequence of diabetes or systemic lupus. A kidney disease may be the result or a consequence of any change, damage, or trauma to the glomerulus, tubules or interstitial tissue in either the renal cortex or renal medulla of the kidney.

In aspects of the invention, the kidney disease is a progressive kidney disease that over time (e.g., days, weeks, months, years) leads to a loss of renal function. "Renal function" generally refers to a physiological property of the kidney, such as the ability to retain protein thereby preventing proteinuria. Renal function can be assessed using methods known in the art such as determining one or more of glomerular filtration rate (e.g., creatinine clearance), excretion of protein in urine, blood urea nitrogen, and serum or plasma creatinine.

In particular aspects of the invention, a progressive kidney disease treated by the compositions and methods described herein includes any kidney disease that can, ultimately, lead to end-stage renal disease. A progressive kidney disease that can be treated by the compositions and methods of the invention can be, for example, associated with endogenous iron deposit in the kidney (e.g., glomerulus, tubules).

In an aspect, the kidney disease is a progressive glomerular kidney disease including without limitation diabetic nephropathy (e.g., as a consequence of Type I or Type Il diabetes or systemic lupus), primary glomerulonephritis (e.g., membranous nephropathy, focal segmental glomerulosclerosis, membranoproliferative glomerulonephritis, diffuse proliferative glomerulonephritis, membranous focal segmental glomerulosclerosis) or secondary glomerulonephritis (e.g., diabetic nephropathy, ischemic nephropathy).

"Proliferative disease" refers to class of diverse disorders and diseases characterized by a lack of control or poorly controlled cell division or proliferation. Proliferative diseases include disorders associated with an overgrowth of connective tissues, such as various fibrotic conditions, including scleroderma, arthritis, juvenile arthritis, gouty arthritis, and liver cirrhosis, and conditions such as restenosis, arteriosclerosis, and proliferative diabetic retinopathy. Proliferative disorders also refers to neoplastic disorders including without limitation anal cancer, bile duct cancer, colon cancer, esophageal cancer, gallbladder cancer, pancreatic cancer, small intestine cancer, stomach cancer, osteosarcoma, ovarian epithelial cancer, gestational trophoblastic tumor, uterine sarcoma, vaginal cancer, vulvar cancer, ovarian germ cell tumor, soft tissue sarcoma, hematopoietic malignancies including acute lymphoblastic leukemia, acute myeloid leukemia, and chronic myelogenous leukemia, lung cancer, small cell lung cancer, malignant mesothelioma, malignant thymoma, hypopharyngeal cancer, laryngeal cancer, nasopharyngeal cancer, oropharyngeal cancer, parathyroid cancer, salivary gland cancer, brain tumor, glioma, cerebellar astrocytoma, cerebral astrocytoma, ependymoma, medulloblastoma, adrenocortical carcinoma, pituitary tumor, islet cell carcinoma, bladder cancer, kidney cancer, penile cancer, Wilm's tumor, AIDS-related lymphoma, cutaneous T-cell lymphoma, Hodgkin's lymphoma, Ewing's sarcoma, skin cancer, hemangiomas of infancy and childhood, mycosis funoides, hairy cell leukemia, Kaposi's sarcoma, non-hodgkin's lymphoma, multiple myeloma, basal cell carcinoma, malignant melanoma, colorectal cancer, non-small cell lung carcinoma, bladder cancer, pancreatic carcinoma, renal cell carcinoma, neuroblastoma, bladder cancer, breast cancer, cervical cancer, liver cancer, sarcomas, thyroid cancer, endometrial cancer, uterine cancer, multiple myeloma, testicular cancer, retinoblastoma, colorectal cancer, oral cancer, rectal cancer, and prostate cancer. The singular form "proliferative disease" includes any one or more diseases selected from the class of proliferative diseases, and includes any compound or complex disease state wherein a component of the disease state includes a disease selected from the class of proliferative diseases. The term also includes proliferative disorders refractory to treatment with other chemotherapeutics or that is refractory to treatment with other chemotherapeutics due to multidrug resistance.

"Cardiovascular disease" includes without limitation vasoconstriction, atherosclerosis, abnormal angiogenesis, thrombosis, stroke, chronic heart failure, myocardial infarction, pulmonary embolism, deep-vein thrombosis, transplant-associated vasculopathy, stenosis (e.g., vein graft stenosis or peri-anastomatic prosthetic graft stenosis), restenosis (e.g., restenosis after angioplasty or stent placement, and the like), atheroma, and vasculitis. Cardiovascular disease also includes vascular conditions that develop after surgical treatments, such as venous bypass surgery, balloon angioplasty, post-angioplasty of atherosclerotic plaques of both coronary and peripheral arteries, and allo- and xenograft rejection. Alternatively, cardiovascular disease refers to the disease of a subject that has suffered ischemia, reperfusion injury, mechanical injury, immunologic injury, pharmacologic injury of a vessel, or coronary trauma.

In aspects of the invention, the cardiovascular disease is diabetic cardiomyopathy. The term "diabetic cardiomyopathy" refers to any one or more cardiac pathology and/or dysfunction which is a complication of diabetes. The diabetes may be symptomatic or asymptomatic. Cardiac pathology which is characteristic of diabetic cardiomyopathy includes myocellular hypertrophy, myocardial fibrosis, and in some cases left ventricular hypertrophy. The pathologies which are contemplated arise independently from complications arising from coronary artery disease, although both diabetic complications and coronary artery complications may be present in the same subject. Diastolic dysfunction, such as an impairment in early diastolic filling, a prolongation of isovolumetric relaxation and increased atrial filling is also characteristic of diabetic cardiomyopathy, and may be identified using Doppler methods such as Doppler 2- dimensional echocardiography (for example Redford MM et al.,JAMA (2003) 289:194- 203) or radionuclide imaging for early or mild dysfunction and by standard echocardiograph testing for more severe dysfunction.

Compositions

The present invention provides compositions for enhancing effects of an ACE inhibitor, ARB and/or a tranilast compound, and compositions for prevention and/or treatment of a fibrotic disease, in particular a kidney disease. A composition of the invention includes at least one tranilast compound and at least one ACE inhibitor and/or ARB.

The compositions of the invention preferably contain a pharmaceutically acceptable carrier or excipient suitable for rendering the compounds administrable orally, intranasally, parenterally, intravenously, intradermally, intramuscularly, subcutaneously, rectally, via inhalation or via buccal administration, or transdermally. The active ingredients may be admixed or compounded with any conventional, pharmaceutically acceptable carrier or excipient. It will be understood by those skilled in the art that any mode of administration, vehicle or carrier conventionally employed and which is inert with respect to the active agents may be utilized for preparing and administering the pharmaceutical compositions of the present invention. Illustrative of such methods, vehicles and carriers are those described, for example, in Remington: The Science and Practice of Pharmacy (21^st Edition, university of the sciences in Philadelphia (Editor), Mack Publishing Company). Those skilled in the art, having been exposed to the principles of the invention, will experience no difficulty in determining suitable and appropriate vehicles, excipients and carriers or in compounding the active ingredients therewith to form the pharmaceutical compositions of the invention.

Compositions of the invention can be formulated as neutral or pharmaceutically acceptable salt forms.

The compositions of the invention may also be conjugated to transport molecules, monoclonal antibodies or transport modalities such as vesicles and micelles that preferentially target recipient cells.

A composition of the invention may be formulated as a unit dosage of at least one tranilast compound and at least one ACE inhibitor and/or ARB to provide beneficial effects. A "unit dosage" refers to a unitary i.e. a single dose which is capable of being administered to a patient, and which may be readily handled and packed, remaining as a physically and chemically stable unit dose including either the active agents as such or a mixture with one or more solid or liquid pharmaceutical excipients or carriers.

In another aspect, an improved composition is provided including therapeutically effective suboptimal doses of at least one tranilast compound and at least one ACE inhibitor and/or ARB in a form for chronic or acute therapy of a disease disclosed herein, in particular kidney disease.

A composition of the invention may be sterilized by, for example, filtration through a bacteria retaining filter, addition of sterilizing agents to the composition, irradiation of the composition, or heating the composition. Alternatively, the compounds or compositions of the present invention may be provided as sterile solid preparations e.g. lyophilized powder, which are readily dissolved in sterile solvent immediately prior to use.

In addition to the formulations described herein, the compositions can also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example, subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the fractions may be formulated with suitable polymeric or hydrophobic materials (for example, as an emulsion in an acceptable oil), or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.

After compositions have been prepared, they can be placed in an appropriate container and labelled for treatment of an indicated condition. For administration of a composition of the invention, such labelling would include amount, frequency, and method of administration.

According to another aspect of the invention, a kit is provided. In an aspect, the kit includes compounds or compositions described herein. The kit may be a package that houses a container which contains a composition of the invention and also houses instructions for administering the composition to a subject.

In embodiments of the invention, a pharmaceutical pack or kit is provided including one or more containers filled with one or more of the ingredients of a composition of the invention to provide a beneficial effect. Associated with such container(s) can be various written materials such as instructions for use, or a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use, or sale for human administration. Administration

Tranilast compounds, ACE inhibitors, ARBs and compositions herein can be administered by any means that produce contact of the active agent(s) with the agent's sites of action in the body of a subject or patient to produce a beneficial effect. The active ingredients can be administered simultaneously or sequentially and in any order at different points in time, to provide the desired beneficial effects. Compounds and compositions disclosed herein can be formulated for sustained release, for delivery locally or systemically. It lies within the capability of a skilled physician or veterinarian to select a form and route of administration that optimizes the effects of the compositions and treatments of the present invention to provide beneficial effects, in particular sustained beneficial effects.

Compounds and compositions can be administered, using any mode of administration that is medically acceptable, meaning any mode that produces therapeutic levels of the active components without causing clinically unacceptable adverse effects. Such modes of administration include, for example, oral, rectal, topical, nasal, intranasal, inhalation, transdermal, parenteral (e.g. subcutaneous, intramuscular, and intravenous), intraocular, intravitreal, or sublingual. Other routes include intrathecal administration directly into spinal fluid, direct introduction such as by catheter and balloon angioplasty devices and intraparenchymal injection into targeted areas.

In particular, the compounds or compositions may be administered in oral dosage forms such as tablets, troches, capsules (each of which includes sustained release or timed release formulations), chachets, lozenges, pills, powders, granules, elixirs, tinctures, suspensions, syrups, chewing gum, wafers, and emulsions. They may also be administered in intravenous (bolus or infusion), intraperitoneal, subcutaneous, or intramuscular forms, all utilizing dosage forms well known to those of ordinary skill in the pharmaceutical arts. The compounds or compositions of the invention may be administered by intranasal route via topical use of suitable intranasal vehicles, or via a transdermal route, for example using conventional transdermal skin patches. A dosage protocol for administration using a transdermal delivery system may be continuous rather than intermittent throughout the dosage regimen. The dosage regimen of the invention will vary depending upon known factors such as the pharmacodynamic characteristics of the agents and their mode and route of administration; the species, age, sex, health, medical condition, and weight of the patient, the nature and extent of the symptoms, the kind of concurrent treatment, the frequency of treatment, the route of administration, the renal and hepatic function of the patient, and the desired effect.

An amount of compound(s) or a composition of the invention which will be therapeutically effective in the treatment of a particular disease to provide effects, in particular beneficial effects, will depend on the nature of the disease, and can be determined by standard clinical techniques. The precise dose to be employed in the formulation will also depend on the route of administration, and the seriousness of the disease, and should be decided according to the judgment of the practitioner and each patient's circumstances.

The dosages used for each compound can be similar to those dosages known to those skilled in the art and used in pre-clinical and clinical studies and in commercial use. The concentrations may be lower than the currently used dosages as the combination of the compounds can increase efficacy of one or more of the compounds. For example, tranilast compounds may be combined with at least one ACE inhibitor and/or ARB with the objective to reduce the dosages and/or reduce frequency of administration of the ACE inhibitor and/or ARB in order to achieve both effective treatment and to lessen any negative effects of the agent(s).

A particular dosage of a tranilast compound for the present invention is the maximum a patient requires to provide an optimal enhancing effect (e.g. synergistic effect), such maximum being tempered by the absolute upper limit of tranilast dosage being the maximum that a patient can tolerate and not develop any serious complications.

A dosage of a tranilast compound for use in the present invention may be within the range of about 0.1 mg to about 2 g per kilogram body weight (assuming a 70 kg subject) day. In aspects of the invention the dosage range may be about 1 mg to about 200 mg per kg per day about 1 mg to about 200 mg per kg per day, about 1 mg to about 100 mg per kg per day, about 5 to 100 mg per kg per day, about 1 mg to about 50 mg per kg per day, about 2 to about 50 mg per kg per day, about 2 mg to about 40 mg per kg, about 2 mg to 30 mg per kg per day, about 2 to 20 mg per kg per day, about 2 to about 25 mg per kg per day, about 2 to 20 mg per kg per day, about 3 to 25 mg per kg per day, about 3 to 20 mg per kg per day, or about 3 to 15 mg per kg per day. In aspects of the invention, a pharmaceutical composition may include about 1 mg to about 1000 mg, about 50 mg to about 1000 mg, about 50 mg to about 900 mg, about 75 mg to about 900 mg, about 100 mg to about 900 mg, about 200 mg to about 900 mg, about 300 mg to about 900 mg, or about 100 mg to about 500 mg, about 100 mg to about 400 mg, or about 100 mg to about 300 mg, of a tranilast compound.

In an aspect of the invention a dosage level of a tranilast compound, in particular a compound of the formula I, II, III, IV, V or Vl is of the order of about 4 mg to about 13 mg per kilogram body weight (assuming a 70 kg subject) per day. The amount of the compound of the formula I, II, III, IV, V or Vl which may be combined with the carrier materials to produce a single dosage will vary, depending upon the host treated and the particular mode of administration. For example, a formulation intended for oral administration to humans may contain about 300 mg to 900mg of the compound with an appropriate and convenient amount of carrier material which may vary from about 5 to 95 percent of the total composition. Unit dosages will generally contain between from about 100 mg to 300 mg of the compound of the formula I, II, III, IV, V or Vl.

In aspects of the invention, the daily doses of a tranilast compound may be from about 0.01 mg/kg per day to 1000 mg/kg per day. Small doses (0.01-1 mg) may be administered initially, followed by increasing doses up to about 1000 mg/kg per day. In the event that the response in a subject is insufficient at such doses, even higher doses (or effective higher doses by a different, more localized delivery route) may be employed to the extent patient tolerance permits.

A subject may be treated with a compound of the formula I, II, III, IV, V or Vl or medicament or formulation thereof on substantially any desired schedule. Multiple doses per day are contemplated to achieve appropriate systemic levels of compounds. For example, a daily dose may be administered in one, two, three or four doses per day. Thus, a medicament of the invention may be administered one or more times per day, in particular 1 or 2 times per day, once per week, once a month or continuously. However, a subject may be treated less frequently, such as every other day or once a week, or more frequently. A compound, medicament or formulation of the invention may be administered to a subject for about or at least about 1 week, 2 weeks to 4 weeks, 2 weeks to 6 weeks, 2 weeks to 8 weeks, 2 weeks to 10 weeks, 2 weeks to 12 weeks, 2 weeks to 14 weeks, 2 weeks to 16 weeks, 2 weeks to 6 months, 2 weeks to 12 months, 2 weeks to 18 months, or 2 weeks to 24 months, periodically or continuously.

In aspects of the invention, the dosage ranges of a tranilast compound may be administered once twice, three times or more daily, especially once or twice daily. In a particular aspect, a tranilast compound of the formula I, II, III, IV, V or Vl is administered in a divided dose schedule, such that there are at least two administrations in total in the schedule. Administrations are given preferably at least every two hours for up to four hours or longer; for example the compound may be administered every hour or every half hour. In an embodiment, the divided-dose regimen includes a second administration of the compound after an interval from the first administration sufficiently long that the level of the compound in the blood has decreased to approximately from 5-

30% of the maximum plasma level reached after the first administration, so as to maintain an effective content of the compound in the blood. Optionally one or more subsequent administrations may be given at a corresponding interval from each preceding administration, preferably when the plasma level has decreased to approximately 10-50% of the immediately-preceding maximum.

In an aspect, the required dose of a tranilast compound administered twice daily is about 1 to 50 mg/kg/day, 1 to 40 mg/kg/day, 3 to 40 mg/kg/day, 3 to 30 mg/kg/day, 3 to 25 mg/kg/day, most preferably 3 to 20 mg/kg/day.

The dosages for ACE inhibitors are known in the art. In aspects of the invention, an ACE inhibitor and/or ARB may generally be administered at dosages in the range of about 0.05 to about 500 mg/day, more preferably about 0.1 to about 250 mg/day and most preferably about 0.2 to about 100 mg/day. For example, doses for perindopril may be within the range of about 0.01 mg to about 100 mg per kilogram body weight (assuming a 70 kg subject) day, and within that range about 0.1 mg to about 75 mg per kg per day, about 0.5 mg to about 50 mg per kg per day, about 0.5 mg to about 30 mg per kg per day, about 1 to 25 mg per kg per day, about 1 mg to about 20 mg per kg per day, about 1 to about 10 mg per kg per day, about 2 mg to about 10 mg per kg, or about 2 mg to 6 mg per kg per day.

Methods and Uses

The present invention provides methods to enhance or potentiate the effects of an ACE inhibitor and/or ARB and/or tranilast compound, and methods of treating a disease disclosed herein, in particular a kidney disease, in patients by administering a therapeutically effective amount of at least one tranilast compound in combination with at least one ACE inhibitor and/or ARB, or alternatively a composition of the invention.

Tranilast compounds can be administered simultaneously, separately or in combination with at least one ACE inhibitor and/or ARB, under different dose and route regimens, to enhance the efficacy of the ACE inhibitor and/or ARB in the treatment of a disease disclosed herein, in particular a kidney disease, in patients compared to when such compounds are administered alone. Greater efficacy and potency of a treatment of the invention may improve the therapeutic ratio of treatment, reducing untoward side effects and toxicity. The methods of the invention may also enhance utility improving longstanding treatment of disease.

The invention additionally provides uses of at least one tranilast compound and at least one ACE inhibitor and/or ARB, or a pharmaceutical composition of the invention in the preparation of medicaments for the prevention and/or treatment of a disease disclosed herein, in particular a kidney disease.

Therapeutic efficacy and toxicity of compositions and methods or the invention may be determined by standard pharmaceutical procedures in cell cultures or with experimental animals such as by calculating a statistical parameter such as the ED₅₀ (the dose that is therapeutically effective in 50% of the population) or LD₅₀ (the dose lethal to 50% of the population) statistics. The therapeutic index is the dose ratio of therapeutic to toxic effects and it can be expressed as the ED₅o/LD₅₀ ratio. Compositions which exhibit large therapeutic indices are preferred.

A method of treatment of the invention may involve administration of a composition including at least one tranilast compound and at least one ACE inhibitor and/or ARB. An alternate method of treatment includes the step of the administration of a composition including at least one tranilast compound followed by the step of the administration of a second pharmaceutical composition including at least one ACE inhibitor and/or ARB.

Alternatively, the administration of the tranilast compound can follow administration of the ACE inhibitor and/or ARB. The administration of the pharmaceutical compositions can occur separately or simultaneously.

The invention will be described in greater detail by way of a specific example. The following example is offered for illustrative purposes, and is not intended to limit the invention in any manner.

Example The renoprotective effects of combining tranilast with ACE inhibition was examined using the 5/6 nephrectomy model, a well characterized model of non-inflammatory proteinuric renal disease in which critical nephron loss leads to progressive renal dysfunction. In this model, TGF-β expression is increased but unaffected by tranilast. The effects of tranilast on TGF-β-dependent collagen production were also examined.

Summary:

Subtotally (5/6) nephrectomized rats were randomized to receive vehicle, the ACE inhibitor, perindopril (6mg/l), tranilast (400mg/kg/day), or their combination for 12 weeks. When compared with sham-nephrectomized animals, subtotally nephrectomized animals had reduced creatinine clearance, proteinuria, glomerulosclerosis, interstitial fibrosis, tubular atrophy, and evidence of TGF-β activity, as indicated by the abundant nuclear staining of phosphorylated Smad2. These manifestations of injury and TGF-β activation were all attenuated by treatment with either tranilast or perindopril, with the latter also attenuating the animals' hypertension. When compared with single-agent treatment, the combination of tranilast and perindopril provided additional, incremental improvements in creatinine clearance, proteinuria, and glomerulosclerosis, and a reduction in nuclear phsopho-Smad2 beyond single-agent treatment.

These findings indicate that the combination of tranilast and perindopril is superior to single-agent treatment on kidney structure and function in the remnant kidney model, and suggests the potential for such dual therapy in kidney disease that continues to progress despite blockade of the renin-angiotensin system.

Materials and methods

Animals

A total of 80 male Sprague-Dawley rats weighing 200-250 g were randomized to eight groups of 10 animals each. Anaesthesia was achieved by the intraperitoneal administration of pentobarbital (6 mg/100 g body weight, Boehringer Ingelheim, Artarmon, NSW, Australia). A total of 60 rats underwent STNx performed by right subcapsular nephrectomy and infarction of approximately two thirds of the left kidney by selective ligation of two of 3-4 extrarenal branches of the left renal artery. Animals were then randomly assigned to the following groups: STNx and vehicle or STNx with either tranilast (400 mg/kg/day by twice daily gavage, Pharm Chemical, Shanghai Lansheng Corporation, China), perindopril (6 mg/kg/day, drinking water) or a combination of tranilast and perindopril. The control group underwent sham surgery consisting of laparotomy and manipulation of both kidneys before wound closure. Rats were housed in a temperature (22°C)-controlled room with ad libitum access to commercial standard rat chow (Norco Co-Operative Ltd, Lismore, NSW, Australia) and water during the entire study. Rats from each group were killed at 12 weeks post-surgery. On killing, the remnant (left) kidney was excised, decapsulated, then sliced sagitally with one-half immersion fixed in 10% neutral buffered formalin and embedded in paraffin for histology, and the other half frozen in liquid nitrogen. All experiments adhered to the guidelines of the Animal Welfare and Ethics Committee of the St Vincent's Hospital and the National Health and Medical Research Foundation of Australia. Renal function

Body weight was measured weekly. Plasma creatinine was measured by autoanalyzer (Beckman Instrumentals, Palo Alto, CA, USA) at the beginning and end of the study. Systolic blood pressure was measured in conscious rats using an occlusive tail-cuff plethysmo-graph attached to a pneumatic pulse transducer (Narco Bio-system Inc., Houston, TX, USA). Before killing, rats were housed in metabolic cages for 24 h for subsequent measurement of urinary creatinine and albumin excretion using a rat- specific radioimmune assay.

Tissue preparation

Rats were anesthetized (Nembutal 60 mg/kg body weight intraperitoneal^, Boehringer- Ingelheim, Australia) and the abdominal aorta cannulated with an 18-G needle. Perfusion-exsanguination commenced at SBP (180-220mm Hg) via the abdominal aorta with 0.1 M phosphate-buffered saline, pH 7.4 (20-50 ml) to remove circulating blood, and the inferior vena cava adjacent to the renal vein was simultaneously severed, allowing free flow of the perfusate. After clearance of circulating blood, 10% buffered formalin was perfused for a further 5 min (100-200 ml) to fix the tissues.

Histopatholoαy

Changes in kidney structure were assessed in a masked protocol in at least 25 randomly selected tissue sections from each group studied. Sections were stained with Mayer's hematoxylin and eosin, periodic acid Schiff's stain, or Masson's modified trichrome to demonstrate collagen matrix.

Glomerulosclerosis

The extent of glomerulosclerosis was determined in 3 mm kidney sections stained with periodic acid Schiff's stain. In brief, 50-80 glomeruli from each rat were examined in a masked protocol. The degree of sclerosis in each glomerulus was graded on a scale of

0-4 as described previously with Grade 0, normal; Grade 1 , sclerotic area up to 25% (minimal); Grade 2, sclerotic area 25-50% (moderate); Grade 3, sclerotic area 50-75% (moderate to severe), and Grade 4, sclerotic area 75-100% (severe). A glomerulosclerotic index (GSI) was then calculated using the formula:

GSI = ∑Fi(/)

i = 0

where Fi is the fraction of glomeruli in the rat with a given score (/).

Tubular atrophy

Tubular atrophy was assessed in kidney sections stained with Masson's trichrome. At x40 magnification (Olympus BX-50 light microscope, Olympus, Tokyo, Japan), six random and non-overlapping fields from each slide (two slides analyzed for N = rats/group) were selected. Atrophic tubules were identified with results expressed as the number of atrophied tubules per field of kidney cortex.

TGF-β receptor activation

Activation of the TGF-β receptor was assessed by quantifying the nuclear expression of phosphorylated Smad2 using a rabbit antiphospho-Smad2 antibody (Cell Signalling Technology, Boston, MA, USA) that detects endogenous Smad2 only when dually phosphorylated at Ser463 and Ser465. Sections were immunostained according to the manufacturer's instructions. Sections incubated with 1 :10 NGS, instead of the primary antiserum, served as the negative control.

Quantitation of matrix deposition and phospho-Smad2 expression

The extent of phospho-Smad2 immunostaining was also quantified using computer- assisted image analysis. For glomerular phospho-Smad2, 10 glomeruli were examined from each rat and the number of glomerular nuclei that showed phospho-Smad2 immunolabelling were counted and expressed as positive nuclei/per glomerulus. For the tubulointerstitium, five random non-overlapping fields from six rats per group were captured and digitized using a BX50 microscope attached to a Fujix HC5000 digital camera. Digital images were then loaded onto a Pentium III IBM computer. The accumulation of matrix within the tubulointerstitium was similarly assessed on Masson's trichrome-stained sections using computer-assisted image analysis. An area of blue on trichrome-stained sections (for matrix) or brown on immunostained sections (for phospho-Smad2) were selected for their color ranges and the proportional area of tissue with their respective ranges of color was then quantified. Calculation of the proportional area stained blue (matrix) or brown (phospho-Smad2) was then determined using image analysis (AIS, Analytical imaging Station Version 6.0, Ontario, Canada).

Cell culture studies

To determine the effects of tranilast on glomerular collagen production in vitro, a well- characterized cloned rat mesangial cell line was studied, which was cultured in

Dulbecco's modified Eagle's medium (GibcoTM; Invitrogen, Grand Island, NY, USA) with fetal bovine serum, 100 U/ml penicillin, and 100 mg/ml streptomycin in humidified

5% CO₂ atmosphere at 37°C. Cells were plated into 24-well dishes at low density and allowed to attach overnight. The subconfluent cells were then serum starved overnight in 0.1% fetal bovine serum before treatment with or without tranilast. Tranilast, 100 and

300 mM, was then added to the wells, followed 4 h later by ³H-proline (1 mCi/well) and

TGF-β 5 ng/ml (R&D systems, Minneapolis, MN, USA). Cells were harvested 48 h poststimulation, washed twice with ice-cold phosphate-buffered saline, and twice with

10% trichloroacetic acid, 500 ml 1M NaOH, and then neutralized with 500 ml 1M HCI. Incorporation of exogenous ³H-proline (L-[2,3,4,5-3H]-proline; Amersham Biosciences,

Piscataway, NJ, USA) was then measured using a liquid scintillation counter (Wallac

1410; Amersham Biosciences). Cell viability was assessed by trypan blue exclusion.

Statistics Data are expressed as means±s.e.m. unless otherwise stated. Statistical significance was determined by a two-way analysis of variance with a Fishers post-hoc comparison. Owing to its skew distribution, data on albuminuria were log transformed prior to analysis and expressed as geometric mean x/tolerance factor. All analyses were performed using Statview Il + Graphics package (Abacus Concepts, Berkeley, CA, USA) on an Apple Macintosh G4 computer (Apple Computer Inc., Cupertino, CA, USA). A P-value <0.05 was regarded as statistically significant.

Results

Animal characteristics

All rats that underwent subtotal nephrectomy were hypertensive and developed substantial proteinuria and reduced creatinine clearance. Treatment of subtotal (5/6) nephrectomy (STNx) rats with perindopril attenuated all of these changes (Table 1). Tranilast also reduced proteinuria and improved creatinine clearance, albeit to a lesser extent than perindopril, but without affecting blood pressure. Without additional blood pressure lowering, the addition of tranilast to perindopril led to a further reduction in proteinuria and incremental improvement in creatinine clearance when compared with either drug as monotherapy (Table 1).

Table 1 - Animal characteristics

SPB, systolic blood pressure; STNx, subtotal (5/6) nephrectomy. Data are expressed as mean + s.e.m. *P<0.05, ^fP<0.05, *P<0.01 versus sham, *P<0.05, ¹¹POOI versus STNx, ^§P<0.05 versus STNx+perindopril.

Histopatholoqy

Subtotal nephrectomy led to significant glomerulosclerosis and tubulointerstitial fibrosis (Figures 1-3), with the proportional area of kidney cortex occupied by collagenous matrix increased more than 10-fold (P<0.01 ; Figure 1) compared with shams. Considerable inter-nephron heterogeneity in the extent of fibrosis and atrophy was also noted. When used as monotherapy, perindopril, and tranilast each reduced the extent of glomerulosclerosis in STNx rats, although the magnitude of this reduction was greater with perindopril than tranilast. The combination of the two agents resulted in a still greater reduction in glomerulosclerosis when compared with either single-agent treatment.

Tubular atrophy, absent in sham kidneys, was also a prominent feature of STNx kidneys (Figures 3, 4) that was reduced by perindopril and to a lesser extent by tranilast also. However, when used together, the perindopril and tranilast combination led to a greater diminution in the extent of tubular atrophy when compared with either agent as monotherapy. All treatments substantially reduced the degree of interstitial fibrosis to a similar extent (Figures 1-4).

Phosphorylated Smad2

In contrast to its minimal immunostaining in sham animals, abundant phosphorylated Smad2 was noted in STNx kidneys (Figures 5, 6), where it was confined to the nuclei and noted to be particularly prominent in areas of fibrosis and atrophy (Figure 6). In contrast, phosphorylated Smad2 immunostaining was substantially lower in STNx rats treated with perindopril and with tranilast. However, animals treated with the combination of perindopril and tranilast showed less phosphorylated Smad2 immunostaining than did rats treated with either agent as monotherapy. Glomerular and tubulointerstitial regions showed similar patterns of phosphorylated Smad2 immunostaining.

In vitro studies

In cultured rat mesangial cells, TGF-β led to a significant increase in extracellular matrix production, as measured by ³H-proline incorporation. This was attenuated by tranilast at 100 mM, although 3H-proline incorporation remained higher than in controls. However, at 300 mM tranilast, 3H-proline incorporation was similar to that in control cells (Figure 7).

Discussion

The present experiment demonstrates that therapy designed to antagonize the effects of TGF-β, another pathogenetic factor in renal disease in addition to blood pressure, resulted in an incremental renoprotective effect on the structural and functional manifestations of renal injury in the subtotally nephrectomized rat, when added to background ACE inhibitor and/or ARB therapy.

TGF-β is a multi-functional cytokine that has long been associated with the pathogenesis of progressive kidney disease, implicated in particular, with the development of fibrosis and atrophy. It is synthesized as a 391 amino-acid precursor molecule with little biological activity, requiring cleavage of its N-terminal latency- associated peptide to give rise to its active form. In addition, the biological effects of TGF-β may also be modified by the presence of the proteoglycan decorin and the scavenging protein alpha 2-macroglobulin. Given these factors that may modulate TGF- β activity, the biological effects were assessed by examining one of its specific intracellular actions, the phosphorylation and nuclear translocation of the TGF-β receptor-activated protein, Smad2. Although only minimal phospho-Smad2 immunostaining was detected in the kidney of sham rats, those from subtotally nephrectomized animals showed prominent nuclear staining of phosphorylated Smad2. In contrast to untreated animals, those receiving either tranilast or perindopril as single agents showed substantial reductions in nuclear phospho-Smad2. Moreover, the combination of the two agents resulted in further incremental reductions in nuclear phospho-Smad2 in both glomerular and the tubulointerstitium that were greater than seen when either agent was used on its own.

Blood pressure reduction and blockade of the renin-angiotensin system are key elements in attenuating the progression of chronic renal disease. Therefore a dose of perindopril was used that in previous dose-finding studies had been shown to effectively reduce blood pressure and provide renal protection with higher doses of the compound failing to provide further benefit. As previously described in the 5/6 nephrectomy model treatment with either tranilast or ACE inhibitor led to improved glomerular filtration rate and diminution in proteinuria. However, in contrast to perindopril, tranilast exerted its renoprotective effects without affecting hypertension. Moreover, despite its lack of effect on systemic pressure, 'add-on' therapy with tranilast led to further incremental benefits on glomerular filtration rate and proteinuria beyond that with perindopril alone.

Histopathologically, kidney disease is characterized by glomerulosclerosis, tubulointerstitial fibrosis, and atrophy. When used as monotherapy, tranilast and perindopril each reduced the extent of glomerulosclerosis and tubulointerstitial fibrosis in subtotally nephrectomized rats. Moreover, when both drugs were used in combination, a further reduction in glomerulosclerosis, beyond that seen with single- agent treatment, was also noted. In addition to fibrosis, tubular atrophy has also been long recognized as an indicator of renal disease severity and progression where its development is thought to reflect apoptosis, rather than necrosis of epithelial cells. It was found that both ACE inhibition and tranilast treatment reduced tubular atrophy. However, the combination of these two agents showed a further beneficial effect on the extent of tubular atrophy, beyond that found with either single-agent treatment.

Overexpression of TGF-β is a characteristic feature of many human kidney diseases and of the 5/6 nephrectomy model in rats. Although ACE inhibitors and ARBs attenuate this overexpression, tranilast does not. However, tranilast effectively reduced TGF-β induced collagen production. These separate, but related mechanisms of RAS blockade and tranilast in inhibiting TGF-β are consistent with their additive effects of these agents when used together in the setting of progressive renal disease.

In clinical studies, additional renal protection can be achieved with increasing the dose of ARB36 or by combining ARBs and ACE inhibitors together. The dose-response relationship between renal protection and perindopril dose has been previously explored demonstrating a maximal perindopril effect at 2 mg/kg/day and also showing that higher doses of perindopril such as the 6 mg/kg/day have similar efficacy to combining perindopril with the ARB, valsartan. Accordingly, the 6 mg/kg/day dose was used to determine whether an alternative non-RAS-dependent treatment may provide additional efficacy.

In summary, treatment with tranilast in addition to ACE inhibitor therapy resulted in an incremental attenuation in the structural and functional manifestations of injury in association with evidence of reduced activation of the TGF-β signalling pathway.

The present invention is not to be limited in scope by the specific embodiments described herein, since such embodiments are intended as but single illustrations of one aspect of the invention and any functionally equivalent embodiments are within the scope of this invention. Indeed, various modifications of the invention in addition to those shown and described herein will become apparent to those skilled in the art from the foregoing description. Such modifications are intended to fall within the scope of the appended claims.

All publications, patents and patent applications referred to herein are incorporated by reference in their entirety to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference in its entirety. The citation of any reference herein is not an admission that such reference is available as prior art to the instant invention.

Claims

WHAT IS CLAIMED IS:

A pharmaceutical composition including at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one angiotensin- converting enzyme inhibitor and/or angiotensin receptor blocker.

2 A pharmaceutical composition according to claim 1 wherein the tranilast compound is a compound of formula I

Formula I

wherein

Ri and R₂ are the same or different and are selected from the group consisting of optionally substituted alkyl, optionally substituted alkynyl and a chain including an optionally substituted triazole;

Xi and X₂ are the same or different and are selected from O and S; and

R₃ and R₄ are the same or different and are selected from the group consisting of OH, hydrogen, halogen, optionally substituted alkyl, optionally substituted alkynyl;

or a pharmaceutically acceptable salt or solvate thereof.

A pharmaceutical composition according to claim 2 wherein Ri and R₂ are the same or different and are selected from the group consisting of optionally substituted Ci to Ci₀ alkyl, optionally substituted C₂ to C₁₀ alkynyl and a chain including an optionally substituted triazole. A pharmaceutical composition according to claim 2 wherein R₃ and R₄ are the same or different and are selected from the group consisting of OH, hydrogen, halogen, optionally substituted Ci to C₁₀ alkyl and optionally substituted C₂ to Ci₀ alkynyl.

A pharmaceutical composition according to claim 2 wherein one of Ri and R₂ is methyl and the other of Ri and R₂ is propargyl or a chain containing a 1 ,4- disubstituted 1 ,2,3-triazole.

A pharmaceutical composition according to claim 1 wherein the tranilast compound is N-[3,4-dimethoxycinnamoyl]anthranilic acid or a pharmaceutically acceptable salt or solvate thereof.

A pharmaceutical composition according to claim 1 wherein the angiotensin- converting enzyme inhibitor is selected from one or more of the group consisting of alacepril, alatriopril, altiopril calcium, ancovenin, benazepril, benazepril hydrochloride, benazeprilat, benzazepril, benzoylcaptopril, captopril, captopril- cysteine, captopril-glutathione, ceranapril, ceranopril, ceronapril, cilazapril, cilazaprilat, converstatin, delapril, delapril-diacid, enalapril, enalaprilat, enalkiren, enapril, epicaptopril, foroxymithine, fosfenopril, fosenopril, fosenopril sodium, fosinopril, fosinopril sodium, fosinoprilat, fosinoprilic acid, glycopril, hemorphin-4, idapril, imidapril, indolapril, indolaprilat, libenzapril, lisinopril, lyciumin A, lyciumin B, mixanpril, moexipril, moexiprilat, moveltipril, muracein A, muracein B, muracein C, pentopril, perindopril, perindoprilat, pivalopril, pivopril, quinapril, quinapril hydrochloride, quinaprilat, ramipril, ramiprilat, spirapril, spirapril hydrochloride, spiraprilat, spiropril, spiropril hydrochloride, temocapril, temocapril hydrochloride, teprotide, trandolapril, trandolaprilat, utibapril, zabicipril, zabiciprilat, zofenopril, zofenoprilat and pharmaceutically acceptable salts and solvates thereof.

A pharmaceutical composition according to claim 1 wherein the angiotensin- converting enzyme inhibitor is perindopril or a pharmaceutically acceptable salt or solvate thereof. 9 A pharmaceutical composition according to claim 1 including N-[3,4- dimethoxycinnamoyl]anthranilic acid or a pharmaceutically acceptable salt or solvate thereof and perindopril or a pharmaceutically acceptable salt or solvate thereof.

10 A pharmaceutical composition according to claim 1 wherein the tranilast compound and/or the angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker are present in a dose that is 1 to 10 fold lower than the respective dose of each component required to treat a disease when administered individually.

11 A pharmaceutical composition according to claim 1 wherein the tranilast compound is present in a synergistically effective amount.

12 A method of treating a disease including administering to a patient a therapeutically effective amount of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and a therapeutically effective amount of at least one angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker.

13 A method according to claim 12, wherein the disease is a fibrotic condition.

14 A method according to claim 12, wherein the disease is a kidney disease or a cardiovascular disease.

15 A method according to claim 12 wherein the tranilast compound and the angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker are administered simultaneously or sequentially.

16 A method according to claim 12 wherein a composition according to claim 1 is administered to the patient. A method according to claim 12 wherein the tranilast compound is a compound of formula I

Formula I

wherein

R₁ and R₂ are the same or different and are selected from the group consisting of optionally substituted alkyl, optionally substituted alkynyl and a chain including an optionally substituted triazole;

Xi and X₂ are the same or different and are selected from O and S; and

R₃ and R₄ are the same or different and are selected from the group consisting of

OH, hydrogen, halogen, optionally substituted alkyl, optionally substituted alkynyl;

or a pharmaceutically acceptable salt or solvate thereof.

A method according to claim 12 wherein the tranilast compound is N-[3,4- dimethoxycinnamoyl]anthranilic acid or a pharmaceutically acceptable salt or solvate thereof.

A method according to claim 12 wherein the angiotensin-converting enzyme inhibitor is perindopril or a pharmaceutically acceptable salt or solvate thereof.

A method according to claim 12 wherein the tranilast compound is N-[3,4- dimethoxycinnamoyl]anthranilic acid or a pharmaceutically acceptable salt or solvate thereof and the angiotensin-converting enzyme inhibitor is perindopril or a pharmaceutically acceptable salt or solvate thereof.

A method according to claim 12 wherein the therapeutically effective dose of the angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker is less than the therapeutic dose range of the angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker when administered alone.

A method for improving the efficacy of an angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker including administering, either sequentially or simultaneously with the administration of the angiotensin- converting enzyme inhibitor and/or angiotensin receptor blocker, at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof.

Use of a composition of claim 1 for treating a fibrotic condition.

Use according to claim 23, wherein the disease is a kidney disease or a cardiovascular disease.

Use of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker for treating a fibrotic condition.

Use of at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof and at least one angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker in the preparation of a medicament for the treatment of a fibrotic condition.

A kit for use in treating a fibrotic condition including at least one tranilast compound or a pharmaceutically acceptable salt or solvate thereof; at least one angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker; and instructions and/or labelling for use in treating a fibrotic condition.