HK1148196B - Use of synthetic triterpenoids in the manufacture of medicament - Google Patents

Description

Pharmaceutical use of synthetic triterpenoids

This application claims U.S. provisional application: priority of both 61/020,624, filed on 11/2008, and 61/109,114, filed on 28/2008, each of which is incorporated by reference in its entirety.

Technical Field

The present invention relates generally to the fields of biology and medicine, and more particularly to compositions and methods for treating and/or preventing renal or renal disease (RKD), insulin resistance, diabetes, endothelial dysfunction, fatty liver disease, and cardiovascular disease (CVD).

Background

Renal failure, which results in insufficient removal of metabolic wastes from the blood and leads to abnormal electrolyte concentrations in the blood, is a significant medical problem throughout the world, especially in developed countries; diabetes and hypertension are among the most important causes of chronic renal failure, also known as Chronic Kidney Disease (CKD), but CKD is also associated with other diseases such as lupus or systemic cardiovascular diseases. These diseases often cause vascular endothelial dysfunction, which is considered to be a major cause of the development of chronic kidney disease. Acute renal failure can result from the use of certain drugs (e.g., acetaminophen) or exposure to toxic chemicals; ischemia-reperfusion injury associated with shock or surgical procedures (such as transplantation) may also lead to acute renal failure, which may ultimately lead to CKD; CKD progresses to end-stage renal disease (ESRD) in many patients, requiring kidney transplantation or regular dialysis to continue life. Both procedures are highly invasive and involve significant side effects and quality of life issues. Although there are some effective therapies for complications such as hyperthyroidism and hyperphosphatemia, there are no therapies that have proven to stop or reverse the progression of the underlying renal failure. Thus, the presence of drugs that improve impaired renal function would indicate an important advance in the treatment of renal failure.

Triterpenoids biosynthesized in plants by squalene cyclization are used for pharmaceutical purposes in many Asian countries, some of which are known to be anti-inflammatory and anti-cancer, such as ursolic acid and oleanolic acid (Huang et al, 1994; Nishino et al, 1988). However, the biological activity of these natural molecules is weak and synthesis of new analogues has been undertaken to enhance efficacy (Honda et al, 1997; Honda et al, 1998). Through continuing efforts to improve the anti-inflammatory and anti-proliferative activities of oleanolic acid and ursolic acid analogs, 2-cyano-3, 12-dioxooleanane-1, 9(11) -diene-28-oic acid (CDDO) and related compounds have been discovered (Honda et al, 1997, 1998, 1999, 2000a, 2000b, 2002; Suh et al, 1998; 1999; 2003; Place et al, 2003; Liby et al, 2005). Several potent derivatives of oleanolic acid have been identified, including methyl-2-cyano-3, 12-dioxooleanane-1, 9-diene-28-oic acid (CDDO-Me; RTA 402); RTA402 inhibits the induction of several important inflammatory mediators such as iNOS, COX-2, TNF α, and IFN γ in activated macrophages. RTA402 has been reported to also activate the Keap1/Nrf2/ARE signaling pathway, thereby producing several anti-inflammatory and antioxidant proteins, such as heme oxygenase-1 (HO-1). These properties have made RTA402 a candidate for the treatment of neoplastic diseases and proliferative diseases, such as the treatment of cancer. However, the ability of such compounds and related molecules to treat and/or prevent renal and cardiovascular diseases has not been tested.

Disclosure of Invention

The present invention provides novel methods for the treatment and/or prevention of renal or renal disease (RKD), insulin resistance, diabetes, endothelial dysfunction, fatty liver disease, cardiovascular disease (CVD) and related disorders. The present application refers to compounds encompassed by or specifically named for the following general or specific chemical formula as "inventive compounds", "compounds of the invention" or "synthetic triterpenoids".

One aspect of the present invention provides a method of treating or preventing renal or renal disease (RKD), insulin resistance, diabetes, endothelial dysfunction, fatty liver disease, or cardiovascular disease (CVD) in a subject, comprising administering to the subject a pharmaceutically effective amount of a compound having the structure:

wherein R1 is: -CN, C₁-C₁₅-acyl group or C₁-C₁₅-alkyl, wherein these groups may or may not be heteroatom-substituted; or a pharmaceutically acceptable salt, hydrate or solvate thereof.

In some embodiments, methods of treating RKD are provided. In some variations, RKD is Diabetic Nephropathy (DN). In some variations, RKD results from toxic damage, for example, poisoning by imaging agents or drugs therein. For example, the drug may be a chemotherapeutic agent. In another variation, RKD is caused by ischemia/reperfusion injury. In yet another variation, RKD is caused by diabetes or hypertension. In other variations, RKD is caused by autoimmune diseases. In other variations, the RKD is chronic RKD. In still other variations, the RKD is acute RKD.

In some embodiments, the subject has received or is receiving dialysis. In some embodiments, the subject has received or is prepared to receive a kidney transplant. In some embodiments, the subject has RKD and is insulin resistant. In some variations of the above embodiments, the subject has RKD and has insulin resistance and endothelial dysfunction. In some embodiments, the subject has RKD and diabetes. In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has diabetes. The pharmaceutically effective amount of the compound is also effective to treat one or more complications associated with diabetes. For example, the complication may be selected from the group consisting of: obesity, hypertension, atherosclerosis, coronary heart disease, stroke, peripheral vascular disease, nephropathy, neuropathy, muscle necrosis, diabetic foot ulcers and other diabetic ulcers, retinopathy and metabolic syndrome (syndrome X). As another example, the complication may be metabolic syndrome (syndrome X). In some variations, insulin resistance results in diabetes.

In some embodiments, the subject has RKD and has endothelial dysfunction. In other embodiments, the subject has RKD and cardiovascular disease. In some embodiments, the subject has CVD. In some variations, endothelial dysfunction leads to CVD.

In some embodiments, the subject has endothelial dysfunction and/or insulin resistance. In some embodiments, the subject has fatty liver disease. In some variations, the fatty liver disease is non-alcoholic fatty liver disease. In other variations, the fatty liver disease is alcoholic fatty liver disease. In some variations, the subject suffers from fatty liver disease and one or more of the following conditions: renal or renal disease (RKD), insulin resistance, diabetes, endothelial dysfunction, and cardiovascular disease (CVD).

In some embodiments, the method further comprises identifying a subject in need of treatment for any of the diseases, dysfunctions, drug resistance or disorders listed herein. In some embodiments, the subject has a family or patient history of any disease, dysfunction, drug resistance, or disorder listed herein. In some embodiments, the subject exhibits symptoms of any of the diseases, dysfunctions, drug resistance, or disorders listed herein.

In another aspect of the present invention, there is provided a method of improving glomerular filtration rate or creatinine clearance in a subject comprising: administering to the subject a pharmaceutically effective amount of a compound having the structure of structural formula I or a pharmaceutically acceptable salt, hydrate, or solvate thereof.

In some embodiments, the compound is administered topically. In some embodiments, the compound is administered systemically. In some embodiments, the compound is administered in the following manner: oral, intralipidic, intraarterial, intraarticular, intracranial, intradermal, intralesional, intramuscular, intranasal, intraocular, intrapericardial, intraperitoneal, intrapleural, intraprostatic, intrarectal, intrathecal, intratracheal, intratumoral, intraumbilical, intravaginal, intravenous, intracapsular, intravitreal, liposomal, topical, mucosal, oral, parenteral, rectal, subconjunctival, subcutaneous, sublingual, topical, buccal, transdermal, vaginal, in creme, in a lipid composition, via a catheter, via lavage, via continuous infusion, via inhalation, via injection, via topical delivery, via topical infusion, direct immersion of target cells, or any combination thereof. For example, in some variations, the compound is administered intravenously, intra-arterially, or orally. For example, in some variations, the compound is administered orally.

In some embodiments, the compounds are formulated as hard or soft capsules, tablets, syrups, suspensions, solid dispersions, cachets, or elixirs. In some variations, the soft capsule is a gelatin capsule. In some variations, the compounds are formulated as a solid dispersion. In some variations, the hard capsule, soft capsule, tablet, or cachet also comprises a protective coating. In some variations, the formulated compound comprises a delayed absorber. In some variations, the formulated compound also includes a solubilizable agent or a co-dispersant. In some variations, the compound is dispersed in a liposome, an oil-in-water emulsion, or a water-in-oil emulsion.

In some embodiments, a pharmaceutically effective amount is about 25mg to about 500mg of the compound per daily dose. In some variations, the daily dose is from about 1mg to about 300 mg of the compound. In some variations, the daily dose is from about 10mg to about 200mg of the compound. In some variations, the daily dose is about 25mg of the compound. In other variations, the daily dose is about 75mg of the compound. In still other variations, the daily dose is about 150mg of the compound. In other variations, the daily dose is from about 0.1mg to about 30mg of the compound. In some variations, the daily dose is about 0.5mg to about 20 mg of the compound. In some variations, the daily dose is from about 1mg to about 15mg of the compound. In some variations, the daily dose is from about 1mg to about 10mg of the compound. In some variations, the daily dose is from about 1mg to about 5mg of the compound.

In some embodiments, a pharmaceutically effective amount is 0.01-25 mg of the compound per kg of body weight per day. In some variations, the daily dose is 0.05-20 mg of the compound per kg of body weight. In some variations, the daily dose is 0.1-10 mg of the compound per kg of body weight. In some variations, the daily dose is 0.1-5 mg of the compound per kg of body weight. In some variations, the daily dose is 0.1-2.5 mg of the compound per kg of body weight.

In some embodiments, the pharmaceutically effective amount is administered as a single daily dose. In some embodiments, the pharmaceutically effective amount is administered in two or more daily doses.

In some embodiments, the method of treatment also includes a second therapy. In some variations, the second therapy comprises administering to the subject a pharmaceutically effective amount of a second drug. In some embodiments, the second agent is a cholesterol-lowering agent, an anti-hyperlipidemic agent, a calcium channel blocker, an antihypertensive agent, or an HMG-CoA reductase inhibitor. Non-limiting examples of the second drug are amlodipine (amlodipine), aspirin, ezetimibe (ezetimibe), felodipine (felodipine), lacidipine (lacidipine), lercanidipine (lercanidipine), nicardipine (nicardipine), nifedipine (nifedipine), nimodipine (nimodipine), nisoldipine (nisoldipine), and nitrendipine (nitrendipine). Additional non-limiting examples of the second drug are atenolol (bucindolol), bucindolol (bucindolol), carvedilol (carvedilol), clonidine (clonidine), doxazosin (doxazosin), indoramin (indoramin) labetalol (labetalol), methyldopa (methylopa), metoprolol (metoprolol), nadolol (nadolol), oxprenolol (oxyprenolol), phenoxybenzamine (phenoxybenzamine), phenotolamine (phenolamine), pindolol (pindolol), prazosin (prazosin), propranolol (propranolol), terazosin (terazosin), timolol (timolol), and tolazoline (tolazoline). In some variations, the second drug is a statin. Non-limiting examples of statins are atorvastatin (atorvastatin), cerivastatin (cerivastatin), fluvastatin (fluvastatin), lovastatin (lovastatin), mevastatin (mevastatin), pitavastatin (pitavastatin), pravastatin (pravastatin), rosuvastatin (rosuvastatin), and simvastatin (simvastatin). In some variations, the second agent is a dipeptidyl peptidase-four (DPP-4) inhibitor. Non-limiting examples of DPP-4 inhibitors are sitagliptin (sitagliptin), vildagliptin (vildagliptin), SYR-322, BMS 477118 and GSK 823093. In some variations, the second drug is a biguanide, for example, the biguanide may be metformin (metformin). In some variations, the second drug is a Thiazolidinedione (TZD), non-limiting examples of TZDs are pioglitazone (pioglitazone), rosiglitazone (rosiglitazone), and troglitazone (troglitazone). In some variations, the second agent is a sulfonylurea derivative. Non-limiting examples of sulfonylurea derivatives are tolbutamide (tolbutamide), acetohexamide (acetohexamide), tolazamide (tolazamide), chlorpropamide (chlorpropamide), glipizide (glipizide), glibenclamide (glyburide), glimepiride (glimepiride), gliclazide (gliclazide). In some variations, the second drug is meglitinide (meglitinide), non-limiting examples of which include repaglinide (repaglinide), mitiglinide (mitiglinide), and nateglinide (nateglinide). In some variations, the second drug is insulin. In some variations, the second agent is an alpha-glucosidase inhibitor. Non-limiting examples of alpha-glucosidase inhibitors are acarbose (acarbose), miglitol (miglitol) and voglibose (voglibose). In some variations, the second agent is a glucagon-like peptide 1 analog. Non-limiting examples of glucagon-like peptide 1 analogs are: exenatide (exenatide) and liraglutide (liraglutide). In some variations, the second drug is a gastric inhibitory peptide analog. In some variations, the second agent is a GPR40 agonist. In some variations, the second agent is a GPR119 agonist. In some variations, the second agent is a GPR30 agonist. In some variations, the second agent is a glucokinase activator. In some variations, the second agent is a glucagon receptor antagonist. In some variations, the second drug is a dextrin analogue, a non-limiting example of which is pramlintide (pramlintide). In some variations, the second agent is an IL-1 β receptor antagonist, a non-limiting example of which is anakinra. In some variations, the second agent is an endocannabinoid receptor antagonist or inverse agonist, a non-limiting example of which is rimonabant (rimonabant). In some variations, the second drug is Orlistat (Orlistat). In some variations, the second drug is Sibutramine (Sibutramine). In some variations, the second agent is a growth factor, non-limiting examples of which are TGF- β 1, TGF- β 2, TGF- β 1.2, VEGF, insulin-like growth factor I or II, BMP2, BMP4, BMP7, GLP-1 analogs, GIP analogs, DPP-IV inhibitors, GPR119 agonists, GPR40 agonists, gastrin, EGF, betacellulin, KGF, NGF, insulin, growth hormone, HGF, FGF homologs of FGF, PDGF, leptin, prolactin, placental prolactin, PTHrP, activin, inhibin, and INGAP. Further non-limiting examples of growth factors are parathyroid hormone, calcitonin, interleukin-6 and interleukin-11.

In some embodiments, the subject is a primate. In some variations, the primate is a human. In other variations, the subject is a cow, horse, dog, cat, pig, mouse, rat, or guinea pig.

In some embodiments, the compound is defined as:

wherein Y is: -H, hydroxy, amino, halo or C₁-C₁₄-alkoxy, C₂-C₁₄-alkenyloxy, C₂-C₁₄-alkynyloxy, C₁-C₁₄Aryloxy group, C₂-C₁₄-aralkyloxy group, C₁-C₁₄Alkylamino radical, C₂-C₁₄-alkenylamino, C₂-C₁₄-alkynylamino, C₁-C₁₄Arylamino, C₃-C₁₀-aryl or C₂-C₁₄-aralkylamino, wherein any of these groups may be heteroatom substituted or not; or a pharmaceutically acceptable salt, hydrate or solvate thereof.

In some embodiments, Y is C unsubstituted with heteroatoms₁-C₄-alkylamino, the compounds of the invention being for example:

in some embodiments, Y is C substituted or unsubstituted with a heteroatom₂-C₄-alkylamino, the compounds of the invention being, for example:

in some embodiments, Y is C substituted or unsubstituted with a heteroatom₁-C₄Alkoxy radicals, such as C, not substituted by hetero atoms₁-C₂-alkoxy groups. For example, one of non-limiting examples of such compounds is:

in some embodiments, at least a portion of CDDO-Me is polymorphic, in a crystalline form having an X-ray diffraction pattern (CuK α): the X-ray diffraction pattern of the crystal has obvious diffraction peaks at 8.8, 12.9, 13.4, 14.2 and 17.4 degrees 2 theta. In a non-limiting example, an X-ray diffraction pattern (CuK α) is substantially as shown in fig. 12A and 12B. In other variations, at least a portion of the CDDO-Me is a polymorphic compound, wherein said amorphous X-ray diffraction pattern (CuK α) has a halo peak at about 13.5 ° 2 θ (substantially as shown in FIG. 12C) and a T_g. In some variations, the compound is amorphous. In some variations, the compound is a glassy solid of CDDO-Me having an X-ray powder diffraction pattern (CuK α) with a halo peak at about 13.5 ° 2 θ (as shown in FIG. 12C) and a T_gAs shown. In some variations, T_gValues fall within the range of about 120 ℃ to about 135 ℃. In some variations, T_gValues of about 125 ℃ to about 130 ℃.

In some embodiments, Y is hydroxy, then the compounds of the invention are, for example:

in some embodiments, the compound is:

in some embodiments, the compound is defined as:

wherein Y' is C substituted or unsubstituted with a heteroatom₁-C₁₄-an aryl group; or a pharmaceutically acceptable salt, hydrate or solvate thereof.

In some embodiments, the compound is:

in some variations of the above methods, the compound is substantially free of its optical isomer. In some variations of the above methods, the compound is in the form of a pharmaceutically acceptable salt. In other variations of the above process, the compound is not a salt.

In some embodiments, the compound is prepared as a pharmaceutical composition comprising (i) a therapeutically effective amount of the compound and (ii) an excipient selected from the group consisting of (a) a carbohydrate, carbohydrate derivative, or carbohydrate polymer, (B) a synthetic organic polymer, (C) an organic acid salt, (D) a protein, polypeptide, or peptide, and (E) a high molecular weight polysaccharide. In some variations, the excipient is a synthetic organic polymer. In some variations, the excipient is selected from the group consisting of hydroxypropyl methylcellulose, poly [1- (2-oxo-1-pyrrolidinyl) ethylene ], or copolymers thereof, and methacrylic acid-methyl methacrylate copolymers. In some variations, the excipient is hydroxypropyl methylcellulose phthalate. In some variations, the excipient is PVP/VA. In some variations, the excipient is methacrylic acid-ethyl acrylate copolymer (1: 1). In some variations, the excipient is copovidone (copovidone).

Any embodiment discussed herein with respect to one aspect of the invention is also applicable to the other aspects of the invention, unless explicitly stated otherwise.

Other objects, features and advantages of the present invention are shown in the following detailed description and the accompanying drawings. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.

Drawings

The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein.

FIGS. 1 a-d: RTA402 reduced renal injury after ischemia reperfusion. From the second day, mice were fed orally daily with RTA402 at a dose of 2mg per kg, or vehicle (sesame oil) alone. On day 0, to induce ischemia reperfusion, the left renal artery of the mouse was clamped with a clamp and removed after 17 minutes. (FIG. 1a) on day 1, blood samples were collected from animals whose renal arteries had been clamped and from control animals that had undergone "sham surgery" without clamping of the renal arteries. Blood Urea Nitrogen (BUN) content was determined as a representative of the degree of renal injury. (FIGS. 1b-d) sections of kidneys from rats treated with RTA 402-treatment and vehicle were scored for tissue damage (FIGS. 1b and 1d) and inflammatory status (FIG. 1 c). (FIG. 1d) Black arrows (vehicle group) indicate two of the renal tubules with a number of severe lesions in the outer medulla. The red arrows (RTA 402 group) indicate two of the many intact renal tubules of the outer medulla.

FIGS. 2 a-c: RTA402 reduced cisplatin-induced nephrotoxicity. Rats were fed orally daily from the first day with RTA402 administered at a dose of 10mg per kg, or vehicle (sesame oil) alone. On day 0, rats were administered cisplatin intravenously at a dose of 6mg per kg. On the indicated days, blood samples were taken to determine creatinine (fig. 2a) and Blood Urea Nitrogen (BUN) (fig. 2b) levels as markers of renal injury. The results on day 3 (creatinine) and day 5 (creatinine and BUN) show statistically significant differences between the vehicle-treated group and the RTA 402-treated group. (FIG. 2c) the RTA 402-treated group had fewer proximal tubular injuries compared to the vehicle-treated group.

FIGS. 3 a-d: RTA402 reduced serum creatinine levels in monkeys, dogs, and rats. (FIG. 3a) Cynomolgus monkeys (Cynomolgus monkey) were orally administered RTA402 at the indicated dose once daily for 28 consecutive days, illustrating the reduced percentage of serum creatinine at day 28 in the RTA402 treated group compared to the vehicle treated group. (FIG. 3b) Beagle dogs (Beagle Dog) were orally administered RTA402 at the indicated doses once daily for three months. The animals of the control group were then dosed with vehicle (sesame oil) and the percentage change in serum creatinine relative to baseline at the three month time point is graphically depicted. (FIG. 3c) Sprague-Dawley rats (Sprague-Dawley rats) orally administered RTA402 at the indicated doses once daily for one month, illustrating the percentage of serum creatinine reduced in the RTA 402-treated group compared to the vehicle-treated group at the end of the study. (FIG. 3d) Spela-Dow rats were orally administered amorphous RTA402 once daily for three consecutive months. The percent serum creatinine decreased in the RTA 402-treated group compared to the vehicle-treated group at the end of the study is shown. Note: in figures 3A, 3C, and 3D, "% decrease" on the vertical axis indicates percent change, e.g., -15 readings on the vertical axis indicate a 15% decrease in serum creatinine.

FIGS. 4A-B: RTA402 reduces serum creatinine levels in human cancer patients and increases the estimated glomerular filtration rate (eGFR). Figure 4A is a graph of serum creatinine measurements from patients in the RTA402 treatment group who participated in a phase I cancer treatment clinical trial. The patient orally administers RTA402 once daily for 21 consecutive days in a dose of 5 to 1300 mg daily. The percent reduction in serum creatinine relative to baseline on the indicated study days is shown graphically. Serum creatinine levels were significantly reduced on days 15 and 21. Fig. 4B is a graph of the calculation of the estimated glomerular filtration rate (eGFR) for the patient of fig. 4A. eGFR was significantly increased in both groups, with all patients: n is 24; patient baseline value is greater than or equal to 1.5: n is 5. In FIGS. 4A and 4B, p is ≦ 0.04;represents that p is 0.01,p is less than or equal to 0.01. Note: in fig. 4A, "reduction from baseline%" on the vertical axis indicates a percentage change. For example, a-15 reading on the vertical axis indicates a 15% reduction in serum creatinine.

FIG. 5: RTA402 increases GFR in human cancer patients. The estimated glomerular filtration rate (eGFR) was measured in RTA402 treatment group patients who participated in a clinical trial of several months cancer treatment. All patients who took six months (n-11) were included in the analysis. The medication information for these patients is already presented in example 5 below.

FIG. 6: RTA402 activity correlates with severity. The magnitude of the decrease in heme A1c is expressed as a fraction of the initial baseline value. Higher baseline values (e.g., average baseline values ≧ 7.0% A1c or ≧ 7.6% A1c) indicate greater magnitude of reduction. The intent-to-treat (ITT) group included all patients (n-53), including patients with a normal A1c value at the beginning.

FIG. 7: RTA402 activity is dose-dependent. The magnitude of the decrease in hemoglobin A1c is shown based on the initial baseline value. The bar graph shows the average results for all patients, all patients with a baseline value of A1c of 7.0%, individual dose cohorts in the cohort with a baseline value of 7.0%, patients with stage 4 renal disease (GFR 15-29 ml/min), where n is the number of patients in each group.

FIG. 8: RTA402 reduced Circulating Endothelial Cells (CECs) and iNOS positive CECs. Mean number of CECs (cells/ml) changes from the baseline-elevated group for the intent-to-treat (ITT) group and 28 days post-treatment before RTA treatment are illustrated. About 20% reduction in the intent-to-treat group, about 33% reduction in the baseline-enhanced group (> 5 CEC/ml), and about 29% reduction in CEC in the iNOS-positive group.

FIG. 9: reversible dose-dependent GFR increase over 28 days. RTA402 treatment increased the dose dependence of GFR, and all evaluable patients were included. Patients with stage 4 renal disease increased > 30%.

FIGS. 10A-B: markers and results of reduced severity of diabetic nephropathy. Adiponectin (fig. 10A) and angiotensin II (fig. 10B) were associated with the severity of nephropathy, both elevated in Diabetic Nephropathy (DN) patients, both of which are shown to be improved. Adiponectin predicts that DN patients contain all the causes of mortality and end stage renal disease. All available data is included.

FIGS. 11A-C: RTA402 significantly reduced uremic solutes. The graph shows the average change in BUN (fig. 11A), phosphorus (fig. 11B), and uric acid (fig. 11C) for all patients and patients showing an increase in the baseline value for a particular solute.

FIGS. 12A-C: x-ray powder diffraction (XRPD) spectra of RTA-402 form A and RTA-402 form B. Figure 12A shows form a without micronization. Figure 12B shows micronized form a. Fig. 12C shows form B.

FIG. 13: RTA-402 Modulated Differential Scanning Calorimetry (MDSC) curve type A. Curve segment and glass transition temperature (T) shown in enlarged view_g) And (5) the consistency is achieved.

FIG. 14: RTA-402 Modulated Differential Scanning Calorimetry (MDSC) curve form B. Curve segment and glass transition temperature (T) shown in enlarged view_g) And (5) the consistency is achieved.

FIG. 15: improved bioavailability of form B (amorphous) in cynomolgus monkeys. A graph showing the area under the curve typical for type A and type B after oral administration of a dose of 4.1mg/kg to cynomolgus monkeys. Each data point represents the mean plasma CDDO methyl ester concentration for 8 animals. Error bars represent standard deviation within the sampled population.

Detailed Description

I. The invention

The present invention relates to novel methods of using triterpenoids to treat and prevent renal diseases and related conditions, including diabetes and cardiovascular diseases.

Definition of

The term "amino" as used herein refers to-NH 2; the term "nitro" refers to — NO 2; the term "halo" means-F, -Cl, -Br, or-I; the term "mercapto" refers to-SH; the term "cyano" refers to — CN; the term "silyl" refers to-SiH 3 and the term "hydroxy" refers to-OH.

The term "heteroatom substituted," when used to modify an organic group (e.g., alkyl, aryl, acyl, etc.), means that one or more hydrogen atoms of the group have been replaced with a heteroatom or heteroatom-containing group. Examples of the hetero atom and the hetero atom-containing group include: hydroxy, cyano, alkoxy, ═ O, ═ S, -NO₂、-N(CH₃)₂Amino or-SH. Certain heteroatom-substituted organic groups are more fully defined below.

The term "unsubstituted" when used to modify an organic group (e.g., alkyl, aryl, acyl, etc.) means that none of the hydrogen atoms in the group are replaced with a heteroatom or heteroatom-containing group. The replacement of a hydrogen atom by a carbon atom or by a group containing only carbon and hydrogen atoms is not sufficient to substitute one group b by a heteroatom. For example: -C6H4C ≡ CH is aryl which is not substituted by heteroatoms, and-C6H 4F is aryl which is substituted by heteroatoms. Certain organic groups which are not substituted with heteroatoms are more fully defined below.

The term "alkyl" includes straight-chain alkyl groups, branched-chain alkyl groups, cycloalkyl (alicyclic) groups, cycloalkyl groups wherein the alkyl group is substituted with a heteroatom, and alkyl groups wherein the cycloalkyl group is substituted with a heteroatom. The term "C unsubstituted by hetero atoms_n-alkyl "refers to a group: having a linear or branched, cyclic or acyclic structure, having no carbon-carbon double or triple bonds, a total of n carbon atoms, all non-aromatic, 3 or more hydrogen atoms, and no heteroatoms. For example: c unsubstituted by hetero atoms₁-C₁₀-alkyl has 1 to 10 carbon atoms. Group (b): -CH₃、-CH₂CH₃、-CH₂CH₂CH₃、-CH(CH₃)₂、-CH(CH₂)₂(cyclopropyl), -CH₂CH₂CH₂CH₃、-CH(CH₃)CH₂CH₃、-CH₂CH(CH₃)₂、-C(CH₃)₃、-CH₂C(CH₃)₃Cyclobutyl, cyclopentyl and cyclohexyl are all examples of alkyl groups which are unsubstituted by heteroatoms. The term "heteroatom-substituted C_n-alkyl "refers to a group: a single saturated carbon atom as the point of attachment, no carbon-carbon double or triple bonds, and a linear or branched, cyclic or acyclic structure, a total of n carbon atoms, all non-aromatic, no or 1 or more hydrogen atoms, containing at least one heteroatom, each heteroatom being independently selected from the group: n, O, F, Cl, Br, I, Si, P and S, e.g. C substituted by hetero atoms₁-C₁₀-alkyl has 1 to 10 carbon atoms. The following groups are examples of heteroatom-substituted alkyl groups: trifluoromethyl, -CH₂F、-CH₂Cl、-CH₂Br、-CH₂OH、-CH₂OCH₃、-CH₂OCH₂CH₃、-CH₂OCH₂CH₂CH₃、-CH₂OCH(CH₃)₂、-CH₂OCH(CH₂)₂、-CH₂OCH₂CF₃、-CH₂OCOCH₃、-CH₂NH₂、-CH₂NHCH₃、-CH₂N(CH₃)₂、-CH₂NHCH₂CH₃、-CH₂N(CH₃)CH₂CH₃、-CH₂NHCH₂CH₂CH₃、-CH₂NHCH(CH₃)₂、-CH₂NHCH(CH₂)₂、-CH₂N(CH₂CH₃)₂、-CH₂CH₂F、-CH₂CH₂Cl、-CH₂CH₂Br、-CH₂CH₂I、-CH₂CH₂OH、-CH₂CH₂OCOCH₃、-CH₂CH₂NH₂、-CH₂CH₂N(CH₃)₂、-CH₂CH₂NHCH₂CH₃、-CH₂CH₂N(CH₃)CH₂CH₃、-CH₂CH₂NHCH₂CH₂CH₃、-CH₂CH₂NHCH(CH₃)₂、-CH₂CH₂NHCH(CH₂)₂、-CH₂CH₂N(CH₂CH₃)₂、-CH₂CH₂NHCO₂C(CH₃)₃and-CH₂Si(CH₃)₃。

The term "C unsubstituted by hetero atoms_n-alkenyl "refers to a group: having a linear or branched, cyclic or acyclic structure containing at least one non-aromatic carbon-carbon double bond, but no carbon-carbon triple bonds, a total of n carbon atoms, three or more hydrogen atoms, and no heteroatoms. For example, C not substituted by hetero atoms₂-C₁₀Alkenyl has 2 to 10 carbon atoms. Alkenyl groups which are not substituted with heteroatoms include: -CH ═ CH₂、-CH＝CHCH₃、-CH＝CHCH₂CH₃、-CH＝CHCH₂CH₂CH₃、-CH＝CHCH(CH₃)₂、-CH＝CHCH(CH₂)₂、-CH₂CH＝CH₂、-CH₂CH＝CHCH₃、-CH₂CH＝CHCH₂CH₃、-CH₂CH＝CHCH₂CH₂CH₃、-CH₂CH＝CHCH(CH₃)₂、-CH₂CH＝CHCH(CH₂)₂and-CH ═ CH-C₆H₅. The term "heteroatom-substituted C_n-alkenyl "refers to a group: having a single non-aromatic carbon atom as attachment point, containing at least one non-aromatic carbon-carbon double bond, but no carbon-carbon triple bond, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, containing no or 1 or more hydrogen atoms, having at least one heteroatom, wherein each heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₂-C₁₀Alkenyl has 2 to 10 carbon atoms. Group (b): -CH ═ CHF, -CH ═ CHCl and-CH ═ CHBr are via heteroatomsExamples of unsubstituted alkenyl groups.

The term "C unsubstituted by hetero atoms_n-alkynyl "refers to a group: having a linear or branched, cyclic or acyclic structure, having at least one carbon-carbon triple bond, a total of n carbon atoms, at least one hydrogen atom, no hetero atom, e.g. C, not substituted by hetero atoms₂-C₁₀Alkynyl has 2 to 10 carbon atoms. Group (b): -C ≡ CH, -C ≡ CCH₃and-C ≡ CC₆H₅Examples of alkynyl groups which are unsubstituted with heteroatoms. The term "heteroatom-substituted C_n-alkynyl "refers to a group: having a single nonaromatic carbon atom as the point of attachment, containing at least one carbon-carbon triple bond, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, containing no or 1 or more hydrogen atoms, containing at least one heteroatom, wherein each heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₂-C₁₀Alkynyl has 2 to 10 carbon atoms. The group-C ≡ CSi (CH)₃)₃Examples of alkynyl groups substituted with heteroatoms.

The term "C unsubstituted by hetero atoms_n-aryl "refers to the group: having a single carbon atom as the point of attachment, wherein the carbon atom is part of an aromatic ring structure containing only carbon atoms, a total of n carbon atoms, 5 or more hydrogen atoms, and no heteroatoms, e.g. C not substituted by heteroatoms₆-C₁₀Aryl has 6 to 10 carbon atoms. Examples of aryl groups which are unsubstituted by heteroatoms include phenyl, methylphenyl, (dimethyl) phenyl, -C₆H₄CH₂CH₃、-C₆H₄CH₂CH₂CH₃、-C₆H₄CH(CH₃)₂、-C₆H₄CH(CH₂)₂、-C₆H₃(CH₃)CH₂CH₃、-C₆H₄CH＝CH₂、-C₆H₄CH＝CHCH₃、-C₆H₄C≡CH、-C₆H₄C≡CCH₃Naphthyl and biphenyl derived groups. "aryl unsubstituted with heteroatoms" includes carbocyclic aryl, biaryl, and polycyclic fused hydrocarbon (PAH) derived groups. The term "heteroatom-substituted C_n-aryl "refers to the group: having a single aromatic carbon atom or a single aromatic heteroatom as attachment point, a total of n carbon atoms, containing at least one hydrogen atom and at least one heteroatom, each heteroatom being independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C1-C10-heteroaryl, which is unsubstituted with heteroatoms, has from 1 to 10 carbon atoms. "heteroatom-substituted aryl" includes heteroaryl. It also includes derivatives of the following compounds: pyrrole, furan, thiophene, imidazole,Oxazole, isoOxazole, thiazole, isothiazole, triazole, pyrazole, pyridine, pyrazine, pyridazine, pyrimidine, and the like. Other examples of heteroatom-substituted aryl groups include-C₆H₄F、-C₆H₄Cl、-C₆H₄Br、-C₆H₄I、-C₆H₄OH、-C₆H₄OCH₃、-C₆H₄OCH₂CH₃、-C₆H₄OCOCH₃、-C₆H₄OC₆H₅、-C₆H₄NH₂、-C₆H₄NHCH₃、-C₆H₄NHCH₂CH₃、-C₆H₄CH₂Cl、-C₆H₄CH₂Br、-C₆H₄CH₂OH、-C₆H₄CH₂OCOCH₃、-C₆H₄CH₂NH₂、-C₆H₄N(CH₃)₂、-C₆H₄CH₂CH₂Cl、-C₆H₄CH₂CH₂OH、-C₆H₄CH₂CH₂OCOCH₃、-C₆H₄CH₂CH₂NH₂、 -C₆H₄CH₂CH＝CH₂、-C₆H₄CF₃、-C₆H₄CN、-C₆H₄C≡CSi(CH₃)₃、-C₆H₄COH、-C₆H₄COCH₃、-C₆H₄COCH₂CH₃、-C₆H₄COCH₂CF₃、-C₆H₄COC₆H₅、-C₆H₄CO₂H、-C₆H₄CO₂CH₃、-C₆H₄CONH₂、-C₆H₄CONHCH₃、-C₆H₄CON(CH₃)₂Furyl, thienyl, pyridyl, pyrrolyl, pyrimidinyl, pyrazinyl, imidazolyl, quinolinyl, and indolyl.

The term "C unsubstituted by hetero atoms_n-aralkyl "refers to a group: with a single saturated carbon atom as the point of attachment, there are a total of n carbon atoms, of which at least 6 constitute an aromatic ring structure containing only carbon atoms, 7 or more hydrogen atoms, and no heteroatoms. For example, C not substituted by hetero atoms₇-C₁₀Aralkyl has 7 to 10 carbon atoms. Examples of aralkyl groups which are not substituted with heteroatoms include benzyl (benzyl) and phenethyl. The term "heteroatom-substituted C_n-aralkyl "refers to a group: having a single saturated carbon atom as the point of attachment, a total of n carbon atoms, no or 1 or more hydrogen atoms, at least one heteroatom, wherein the aromatic ring structure contains at least one carbon atom, and wherein each heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₂-C₁₀Heteroaralkyl has 2 to 10 carbon atoms.

The term "C unsubstituted by hetero atoms_nBy acyl is meant such a radicalAnd (3) clustering: having one carbon atom of the carbonyl group as an attachment point, having a linear or branched, cyclic or acyclic structure, a total of n carbon atoms, containing 1 or more hydrogen atoms, a total of one oxygen atom, and no other heteroatoms. For example, C not substituted by hetero atoms₁-C₁₀Acyl has 1 to 10 carbon atoms. Group (b): -COH, -COCH₃、-COCH₂CH₃、-COCH₂CH₂CH₃、-COCH(CH₃)₂、-COCH(CH₂)₂、-COC₆H₅、-COC₆H₄CH₃、-COC₆H₄CH₂CH₃、-COC₆H₄CH₂CH₂CH₃、-COC₆H₄CH(CH₃)₂、-COC₆H₄CH(CH₂)₂and-COC₆H₃(CH₃)₂Examples of acyl groups which are unsubstituted with heteroatoms. The term "heteroatom-substituted C_n-acyl "refers to a group: having a single carbon atom as attachment point (which carbon atom is part of a carbonyl group), having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, containing no or 1 or more hydrogen atoms, and containing at least one further heteroatom in addition to the oxygen atom of the carbonyl group, wherein each further heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₁-C₁₀Acyl has 1 to 10 carbon atoms. The term "heteroatom-substituted acyl" includes carbamoyl, thiocarboxylate and thiocarboxylic acid groups. Group (b): -COCH₂CF₃、-CO₂H、-CO₂CH₃、-CO₂CH₂CH₃、-CO₂CH₂CH₂CH₃、-CO₂CH(CH₃)₂、-CO₂CH(CH₂)₂、-CONH₂、-CONHCH₃、-CONHCH₂CH₃、-CONHCH₂CH₂CH₃、-CONHCH(CH₃)₂、-CONHCH(CH₂)₂、-CON(CH₃)₂、-CON(CH₂CH₃)CH₃、-CON(CH₂CH₃)₂and-CONHCH₂CF₃Are examples of acyl groups substituted with heteroatoms.

The term "C unsubstituted by hetero atoms_n-alkoxy "refers to a group having the structure-OR, wherein R is C unsubstituted with hetero atoms_n-alkyl, as defined above. Alkoxy groups which are not substituted with heteroatoms include: -OCH₃、-OCH₂CH₃、-OCH₂CH₂CH₃、-OCH(CH₃)₂and-OCH (CH)₂)₂. The term "heteroatom-substituted C_n-alkoxy "refers to a group having the structure-OR, wherein R is C substituted with a heteroatom_n-alkyl, as defined above. For example, -OCH₂CF₃Is alkoxy substituted by hetero atom.

The term "C unsubstituted by hetero atoms_n-alkenyloxy "means a group having the structure-OR, wherein R is C unsubstituted by hetero atoms_n-alkenyl, as defined above. The term "heteroatom-substituted C_n-alkenyloxy "refers to a group having the structure-OR, wherein R is C substituted with a heteroatom_n-alkenyl, as defined above.

The term "C unsubstituted by hetero atoms_n-alkynyloxy "refers to a group having the structure-OR, wherein R is C unsubstituted with hetero atoms_nAlkynyl, as defined above. The term "heteroatom-substituted C_n-alkynyloxy "refers to a group having the structure-OR, wherein R is C substituted with a heteroatom_nAlkynyl, as defined above.

The term "C unsubstituted by hetero atoms_n-aryloxy "refers to a group having the structure-OAr, wherein Ar is C unsubstituted with heteroatoms_n-aryl, as defined above. An example of an aryloxy group which is not substituted by hetero atoms is-OC₆H₅. The term "heteroatom-substituted C_n-aryloxy isRefers to a group having the structure-OAr, wherein Ar is C substituted with a heteroatom_n-aryl, as defined above.

The term "C unsubstituted by hetero atoms_nBy aralkoxy is meant having-OR_ArA group of the structure (I), wherein R_ArIs C not substituted by hetero atoms_nAralkyl, as defined above. The term "heteroatom-substituted C_nBy aralkoxy is meant having-OR_ArA group of the structure (I), wherein R_ArIs C substituted by hetero atoms_n-aralkyl radical, C_nAralkyl is as defined above.

The term "C unsubstituted by hetero atoms_n-acyloxy "means a radical having the structure-OAc, where Ac is C unsubstituted by a heteroatom_nAcyl, as defined above. Heteroatom-unsubstituted acyloxy groups include alkylcarbonyloxy and arylcarbonyloxy. For example, -OCOCH₃Examples of acyloxy groups which are not substituted by heteroatoms. The term "heteroatom-substituted C_n-acyloxy "means a radical having the structure-OAc, where Ac is C substituted with a heteroatom_nAcyl, as defined above. Heteroatom-substituted acyloxy groups include alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkoxycarbonyl, aminocarbonyl, and alkylthiocarbonyl groups.

The term "C unsubstituted by hetero atoms_n-alkylamino "refers to a group: having a single nitrogen atom as the point of attachment, containing one or two saturated carbon atoms linked to said nitrogen atom, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, all non-aromatic carbon atoms, containing 4 or more hydrogen atoms, a total of 1 nitrogen atom, and no other heteroatoms. For example, C not substituted by hetero atoms₁-C₁₀Alkylamino has 1 to 10 carbon atoms. The term "C unsubstituted by hetero atoms_n-alkylamino "includes groups having the structure-NHR, wherein R is C which is unsubstituted by a heteroatom_n-alkyl, as defined above. Alkylamino groups which are not substituted with heteroatoms include-NHCH₃、-NHCH₂CH₃、-NHCH₂CH₂CH₃、-NHCH(CH₃)₂、-NHCH(CH₂)₂、-NHCH₂CH₂CH₂CH₃、-NHCH(CH₃)CH₂CH₃、-NHCH₂CH(CH₃)₂、-NHC(CH₃)₃、-N(CH₃)₂、-N(CH₃)CH₂CH₃、-N(CH₂CH₃)₂N-pyrrolidinyl, and N-piperidinyl. The term "heteroatom-substituted C_n-alkylamino "refers to a group: having a single nitrogen atom as attachment point, having one or two saturated carbon atoms attached to the nitrogen atom, having no carbon-carbon double or triple bonds, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, all of which are non-aromatic carbon atoms, having no or 1 or more hydrogen atoms, and having at least one further heteroatom other than the nitrogen atom at the attachment point, each of said further heteroatoms being independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₁-C₁₀Alkylamino has 1 to 10 carbon atoms. The term "heteroatom-substituted C_nThe "alkylamino group" includes a group having the structure-NHR, wherein R is C substituted with a hetero atom_n-alkyl, as defined above.

The term "C unsubstituted by hetero atoms_n-alkenylamino "refers to a group: having a single nitrogen atom as the point of attachment, having one or two carbon atoms attached to the nitrogen atom, having a linear or branched, cyclic or acyclic structure, containing at least one non-aromatic carbon-carbon double bond, a total of n carbon atoms, 4 or more hydrogen atoms, a total of one nitrogen atom, and no other heteroatoms. For example, C not substituted by hetero atoms₂-C₁₀Alkenylamino has 2 to 10 carbon atoms. The term "C unsubstituted by hetero atoms_n-alkenylamino "includes groups having the structure-NHR, where R is C which is unsubstituted by a heteroatom_n-alkenyl, as defined above. C unsubstituted by hetero atoms_nExamples of-alkenylamino also include dienylamino and alkyl (alkenyl) amino. The term "heteroatom-substituted C_n-alkenylamino "refers to a group: having a single nitrogen atom as attachment point and containing at least one non-aromatic carbon-carbon double bond, but no carbon-carbon triple bond, having one or two carbon atoms attached to the nitrogen atom, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, containing no or 1 or more hydrogen atoms, and having at least one further heteroatom other than the nitrogen atom at the attachment point, wherein each further heteroatom is independently selected from the group: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₂-C₁₀Alkenylamino has 2 to 10 carbon atoms. The term "heteroatom-substituted C_n-alkenylamino "includes groups having the structure-NHR, where R is C substituted with a heteroatom_n-alkenyl, as defined above.

The term "C unsubstituted by hetero atoms_n-alkynylamino "refers to the group: having a single nitrogen atom as an attachment point, having one or two carbon atoms attached to the nitrogen atom, having a linear or branched, cyclic or acyclic structure, containing at least one carbon-carbon triple bond, a total of n carbon atoms, containing at least one hydrogen atom, a total of one nitrogen atom, and no other heteroatoms. For example, C not substituted by hetero atoms₂-C₁₀-alkynylamino has 2 to 10 carbon atoms. The term "C unsubstituted by hetero atoms_n-alkynylamino "includes groups having the structure-NHR, wherein R is C which is unsubstituted by a heteroatom_nAlkynyl, as defined above. Alkynylamino includes dialkynylamino and alkyl (alkynyl) amino. The term "heteroatom-substituted C_n-alkynylamino "refers to the group: having a single nitrogen atom as attachment point, having one or two carbon atoms attached to the nitrogen atom, and having at least one non-aromatic carbon-carbon triple bond, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, no or 1 or more hydrogen atoms, and having at least one further heteroatom in addition to the nitrogen atom at the attachment point, wherein each further heteroatom isThe heteroatoms of (a) are each selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₂-C₁₀-alkynylamino has 2 to 10 carbon atoms. The term "heteroatom-substituted C_n-alkynylamino "includes groups having the structure-NHR, wherein R is C substituted with a heteroatom_nAlkynyl, as defined above.

The term "C unsubstituted by hetero atoms_nBy arylamino is meant a group: having a single nitrogen atom as the point of attachment, having at least one aromatic ring structure attached to the nitrogen atom-wherein the aromatic ring structure contains only carbon atoms-n total carbon atoms, 6 or more hydrogen atoms, one nitrogen atom in total, and no other heteroatoms. For example, C not substituted by hetero atoms₆-C₁₀Arylamino groups have 6 to 10 carbon atoms. The term "C unsubstituted by hetero atoms_nArylamino includes groups having the structure-NHR, where R is C which is unsubstituted by a heteroatom_n-aryl, as defined above. Arylamino groups which are unsubstituted with heteroatoms include diarylamino groups and alkyl (aryl) amino groups. The term "heteroatom-substituted C_nBy arylamino is meant a group: having a single nitrogen atom as the point of attachment, a total of n carbon atoms, containing at least one hydrogen atom, and having at least one additional heteroatom in addition to the nitrogen atom at the point of attachment, wherein at least one of said carbon atoms is contained in one or more aromatic ring structures, wherein each additional heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₆-C₁₀Arylamino groups have 6 to 10 carbon atoms. The term "heteroatom-substituted C_n-arylamino "includes groups having the structure-NHR, where R is C substituted with a heteroatom_n-aryl, as defined above. Heteroatom-substituted arylamino groups include heteroarylamino groups.

The term "C unsubstituted by hetero atoms_n-aralkylamino "means a group: having a single nitrogen atom as attachment point, having one or two saturated carbon atoms bound to the nitrogen atom, in totalHaving n carbon atoms, wherein at least 6 carbon atoms form an aromatic ring structure containing only carbon atoms, 8 or more hydrogen atoms, for a total of one nitrogen atom, and no other heteroatoms. For example, C not substituted by hetero atoms₇-C₁₀Aralkylamino has 7 to 10 carbon atoms. The term "C unsubstituted by hetero atoms_n-aralkylamino "includes groups having the structure-NHR, wherein R is C which is unsubstituted by a heteroatom_nAralkyl, as defined above. Aralkylamino includes diaralkylamino. The term "heteroatom-substituted C_n-aralkylamino "means a group: having a single nitrogen atom as attachment point, having at least one or two saturated carbon atoms attached to the nitrogen atom, a total of n carbon atoms, containing no or 1 or more hydrogen atoms, at least one of said carbon atoms being incorporated into an aromatic ring, having at least one additional heteroatom in addition to the nitrogen atom at the attachment point, wherein each heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₇-C₁₀Aralkylamino has 7 to 10 carbon atoms. The term "heteroatom-substituted C_n-aralkylamino "includes groups having the structure-NHR, wherein R is C substituted with a heteroatom_nAralkyl, as defined above. The term "heteroatomic substituted aralkylamino" embraces the term "heteroaralkylamino".

The term "amido" includes N-alkylamido, N-arylamido, N-aralkamido, acylamino, alkylcarbonylamino, arylcarbonylamino, ureido. group-NHCOCH₃Examples of amide groups which are not substituted with heteroatoms. The term "C unsubstituted by hetero atoms_nBy amido "is meant a group: having a single nitrogen atom as an attachment point, having a carbonyl group attached to the nitrogen atom via a carbon atom thereof, having a linear or branched, cyclic or acyclic structure, having a total of n carbon atoms, containing 1 or more hydrogen atoms, a total of one oxygen atom, a total of one nitrogen atom, and no other hetero atoms. For example, C not substituted by hetero atoms₁-C₁₀Amide groups have 1 to 10 carbon atoms. The term "not substituted by hetero atomsC of (A)_n-amido "includes groups having the structure-NHR, where R is C which is unsubstituted by a heteroatom_nAcyl, as defined above. The term "heteroatom-substituted C_nBy amido "is meant a group: having a single nitrogen atom as attachment point and having a carbonyl group attached to the nitrogen atom via a carbon atom thereof, having a linear or branched, cyclic or acyclic structure, having a total of n aromatic or nonaromatic carbon atoms, containing no or 1 or more hydrogen atoms, and having at least one further heteroatom other than the oxygen atom of the carbonyl group and the nitrogen atom of the attachment point, wherein each further heteroatom is independently selected from the group consisting of: n, O, F, Cl, Br, I, Si, P and S. For example, C substituted by hetero atoms₁-C₁₀Amide groups have 1 to 10 carbon atoms. The term "heteroatom-substituted C_n-amido "includes groups having the structure-NHR, where R is C which is unsubstituted by a heteroatom_nAcyl, as defined above. group-NHCO₂CH₃Are examples of heteroatom-substituted amido groups.

Further, the atoms comprising the compounds of the present invention are intended to include all isotopic forms of these atoms. Isotopes used herein include atoms of the same atomic number but different mass numbers. As a general example and without limitation, isotopes of hydrogen include tritium and deuterium, while isotopes of carbon include¹³C and¹⁴C. similarly, it is contemplated that one or more carbon atoms of the compounds of the present invention may be replaced by a silicon atom. Similarly, it is contemplated that one or more of the oxygen atoms of the compounds of the present invention may also be replaced by a sulfur or selenium atom.

Any undefined valence bond in an atom of the structure shown in this application implicitly represents a hydrogen atom bound to the atom.

The words "a" and "an", when used in the claims and/or in this patent specification with the term "comprising", may mean "one", but also correspond to the meaning of "one or more", "at least one", and "one" or more than one ".

In this application, the term "about" is used to indicate that a value includes the inherent error variation of the equipment and method used to determine the value, or the variation present in the subject.

The terms "comprising", "having" and "including" are open-ended linking verbs. Any form or tense of one or more of these verbs, such as "comprising", "having", "including", and "including", is also open-ended. For example, any method that "comprises," "has," or "includes" one or more steps is not limited to having the one or more steps, but also includes other steps not listed.

The term "effective" as used in this patent specification and/or claims means sufficient to obtain a desired, expected, or intended result.

The term "hydrate," when used to modify a compound, means that the compound has less than one (e.g., hemihydrate), one (e.g., monohydrate), or more than one (e.g., dihydrate) water molecule associated with each compound molecule, e.g., the compound in solid form.

The term "IC" herein₅₀"refers to the amount of inhibitor at which 50% of the maximal response is obtained.

An "isomer" of a first compound is an independent compound, in which the constituent atoms of each molecule are the same as in the first compound, but the three-dimensional configuration of these atoms is different.

The term "patient" or "subject" herein refers to a living mammal, such as a human, monkey, cow, sheep, goat, dog, cat, mouse, rat, guinea pig, or transgenic species thereof. In certain embodiments, the patient or subject is a primate. Non-limiting examples of human subjects are adults, adolescents, infants, and fetuses.

By "pharmaceutically acceptable" is meant that it can be used to prepare generally safe, non-toxic pharmaceutical compositions, is biologically and otherwise undesirable, and is useful for veterinary as well as human medical uses.

By "pharmaceutically acceptable salt" is meant a salt of a compound of the invention which is pharmaceutically acceptable as defined above and which has the desired pharmacological activity. Such salts include acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, or with organic acids such as 1, 2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, 2-naphthalenesulfonic acid, 3-phenylpropionic acid, 4' -methylenebis (3-hydroxy-2-ene-1-carboxylic acid), 4-methylbicyclo [2.2.2] oct-2-ene-1-carboxylic acid, acetic acid, aliphatic mono-and dicarboxylic acids, aliphatic sulfuric acids, aromatic sulfuric acids, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, carbonic acid, cinnamic acid, citric acid, cyclopentanepropionic acid, ethanesulfonic acid, fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, glycolic acid, heptanoic acid, hexanoic acid, hydroxynaphthoic acid, lactic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, glycolic acid, heptanoic acid, caproic acid, and the like, Maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, muconic acid, o- (4-hydroxybenzoyl) benzoic acid, oxalic acid, p-chlorobenzenesulfonic acid, phenyl-substituted alkanoic acids, propionic acid, p-toluenesulfonic acid, pyruvic acid, salicylic acid, stearic acid, succinic acid, tartaric acid, tert-butylacetic acid, trimethylacetic acid, and the like. Pharmaceutically acceptable salts also include base addition salts formed by reaction of an acidic proton present with an inorganic or organic base. Acceptable inorganic bases include sodium hydroxide, sodium carbonate, potassium hydroxide, aluminum hydroxide, and calcium hydroxide. Acceptable organic bases include ethanolamines, diethanolammines, triethanolammines, tromethamine, N-methylglucamine, and the like. It will be appreciated that the particular anion or cation used to form any salt of the invention is not critical, as long as the salt as a whole is pharmacologically acceptable. Handbook of Pharmaceutical Salts: there are other examples of pharmaceutically acceptable salts and methods of making and using them within Properties, and Use (handbook of pharmaceutical salts: Properties and uses) (edited by P.H. Stahl and C.G. Wermuth, Verlag Helvetica ChimicaActa, 2002).

By "predominantly one enantiomer" herein is meant that the compound contains at least about 85% of the same enantiomer, or more preferably at least about 90% of the same enantiomer, or even more preferably at least about 95% of the same enantiomer, or most preferably at least about 99% of the same enantiomer. Similarly, the phrase "substantially free of other optical isomers" means that the composition contains at most about 15% of the other enantiomer or diastereomer, more preferably at most about 10% of the other enantiomer or diastereomer, even more preferably at most about 5% of the other enantiomer or diastereomer, and most preferably at most about 1% of the other enantiomer or diastereomer.

"preventing" or "prevention" includes: (1) inhibiting onset of a disease in a subject or patient who may be at risk of and/or predisposed to the disease, but does not yet experience or display any or all of the conditions or symptoms of the disease, and/or (2) slowing onset of the conditions or symptoms of the disease in a subject or patient who may be at risk of and/or predisposed to the disease, but does not yet experience or display any or all of the conditions or symptoms of the disease.

The term "saturated" if referring to an atom means that the atom is only singly bonded to other atoms.

"stereoisomers" or "optical isomers" are isomers of particular compounds in which the same atom is bonded to the same other atom but the atoms differ in their three-dimensional spatial configuration. "enantiomers" are stereoisomers of particular compounds that are mirror images of each other, including the left and right hands. "diastereoisomers" are stereoisomers of a particular compound, but not enantiomers thereof.

By "therapeutically effective amount" or "pharmaceutically effective amount" is meant an amount administered to a subject or patient for the treatment of a disease that is sufficient to treat the disease.

"treating" or "treating" includes (1) inhibiting the disease in a subject or patient who is experiencing or exhibiting the condition or symptom of the disease (e.g., arresting further development of the condition and/or symptom), (2) ameliorating the disease in a subject or patient who is experiencing or exhibiting the condition or symptom of the disease (e.g., reversing the condition and/or symptom), and/or (3) obtaining any measurable amount of remission of the disease in a subject or patient who is experiencing or exhibiting the condition or symptom of the disease.

The term "water-soluble" as used herein means that the compound is soluble in water, has a solubility of at least 0.010 mole/liter, or is classified as soluble according to the literature.

Other abbreviations herein are as follows: DMSO ═ dimethyl sulfoxide; NO ═ nitric oxide; iNOS ═ inducible nitric oxide synthase; COX-2 ═ cyclooxygenase-2; NGF ═ nerve growth factor; IBMX ═ isobutylmethylxanthine; FBS is fetal bovine serum; GPDH ═ glycerol 3-phosphate dehydrogenase; RXR ═ retinoic acid X receptor; TGF- β ═ β type transforming growth factor; IFN γ or IFN- γ ═ interferon- γ; LPS ═ bacterial endotoxin lipopolysaccharide; TNF α or TNF- α ═ tumor necrosis factor- α; IL-1 β ═ interleukin-1 β; GAPDH ═ 3-glyceraldehyde phosphate dehydrogenase; MTBE ═ methyl tert-butyl ether; MTT ═ 3- (4, 5-dimethylthiazole-2) -2, 5-diphenyltetrazolium bromide salt; TCA ═ trichloroacetic acid; HO-1 ═ inducible heme oxygenase.

The above definitions supersede any conflicting definition in any reference incorporated herein by reference.

Synthesis of triterpene Compounds

Triterpenoids are synthesized in plants via squalene cyclization, which is used in medicine in many asian countries; some of them (e.g., oleanolic acid and ursolic acid) are known to have anti-inflammatory and anti-cancer effects (Huang et al, 1994; Nishino et al, 1988). However, the biological activity of such natural molecules is relatively weak, and therefore, synthesis of new analogs has been performed to enhance their efficacy (Honda et al, 1997; Honda et al, 1998). Subsequent studies have identified a number of synthetic compounds with higher activity than the natural triterpenoids.

2-cyano-3, 12-dioxooleanane-1, 9(11) -dien-28-oic acid (CDDO, RTA-402) and related compounds (e.g., CDDO-Me, TP-225, CDDO-Im) were discovered in an ongoing effort to improve the anti-inflammatory and anti-proliferative activities of oleanolic acid and ursolic acid analogs (Honda et al, 1997, 1998, 1999, 2000a, 2000b, 2002; Suh et al, 1998; 1999; 2003; Place et al, 2003; Liby et al, 2005). In the case of induction of cytoprotective genes through signal transduction of Keap1-Nrf 2-Antioxidant Response Element (ARE), structural activity evaluation was recently made on 15 triterpenoids, and it was found that the importance of michael receptor groups on ring a and ring C, and the nitrile group on ring a C-2 and the substituent on ring C-17 influence the pharmacodynamic action in vivo (Yates et al, 2007).

In general, CDDO is the prototype of a large number of compounds belonging to a group of agents that have shown multiple uses. For example, CDDO-Me and CDDO-Im have been reported to modulate transforming growth factor-beta (TGF-. beta.) or Smad signaling in various classes of cells (Suh et al, 2003; Minns et al, 2004; Mix et al, 2004). Both ARE known to be potent inducers of heme oxygenase-1 and Nrf2/ARE signaling (Liby et al, 2005), and a series of synthetic Triterpenoid (TP) analogs of oleanolic acid have been shown to be potent inducers of phase 2 responses, elevating nad (p) H-quinone oxidoreductase and heme oxygenase-1 (HO-1), heme oxygenase-1 being the primary protectant that protects cells from oxidative and electrophilic stresses (Dinkova-kostowa et al, 2005). Like the previously identified phase 2 inducers, TP analogs have been shown to utilize the signal pathway of the antioxidant response element, Nrf2-Keap 1.

RTA402, bardoxolone (bardoxolone) methyl, is a compound used in conjunction with the methods within the present invention, a clinically developed Antioxidant Inflammation Modulator (AIM) for inflammation and cancer related indications that inhibits immune-mediated inflammation by restoring redox balance in inflamed tissues. It also induces the cell protective transcription factor Nrf2, and inhibits the activity of pro-oxidant and pro-inflammatory transcription factors NF-kB and STAT 3. In vivo, RTA402 has shown significant single-agent anti-inflammatory activity in several animal models of inflammation, such as renal damage in the cisplatin model, and acute renal injury in the ischemia-reperfusion model. In addition, it has been found that serum creatinine levels are significantly reduced in patients treated with RTA 402.

In one aspect of the invention, the compounds of the invention are useful for treating a subject suffering from renal disease resulting from an increased level of oxidative stress in one or more tissues. Acute or chronic inflammation may occur with oxidative stress. Oxidative stress can be caused by the following conditions: acute exposure to external agents, such as ionizing radiation or cytotoxic chemotherapeutic drugs (e.g., doxorubicin); trauma or other acute tissue injury; ischemia/reperfusion injury; poor blood circulation or anemia; local or systemic hypoxia or hyperoxia; or other abnormal physiological state such as hyperglycemia or hypoglycemia.

Thus, treatment of a disease involving oxidative stress alone or oxidative stress aggravated by inflammation may comprise administering to the subject a therapeutically effective amount of a compound of the invention, such as described above or throughout this patent specification. Prophylactic treatment can be performed before a foreseeable oxidative stress (e.g. organ transplantation or treatment of cancer patients) occurs, or treatment can be performed on existing oxidative stress and inflammation.

It has now been found that newer amide derivatives of CDDO are likely to be effective agents because, for example, these derivatives have the ability to cross the blood brain barrier. In addition to CDDO formamide (CDDO-MA) (as reported by Honda et al in 2002), the present invention provides for the use of other CDDO amide derivatives, such as CDDO acetamide (CDDO-EA) and fluorinated amide derivatives of CDDO (e.g., 2, 2, 2-trifluoroethylamide derivatives of CDDO, i.e., CDDO-TFEA).

The compounds of the present invention can be prepared according to the methods described by Honda et al (1998), Honda et al (2000b), Honda et al (2002), Yates et al (2007), and U.S. Pat. Nos. 6,326,507 and 6,974,801, which are incorporated herein by reference.

Non-limiting examples of triterpenoids that may be used according to the methods within the present invention are as follows.

The compounds used in the present invention (e.g., as shown in the above table) are structurally similar to RTA-402 and, as noted above, in many cases exhibit similar biological properties. Table 1 summarizes in vitro results for several more examples of these compounds, where RAW264.7 macrophages were pretreated with different concentrations (nM) of DMSO or drug for 2 hours, followed by 20ng/ml of IFN γ for 24 hours. The concentration of NO in the medium was measured using a Griess reagent system; cell viability was determined with WST-1 reagent. NQO1 CD represents the concentration required to induce doubling of NQO1 expression, NQO1 being an Nrf2 regulated antioxidant enzyme in Hepa1c1c7 murine hepatoma cells (Dinkova-Kostova et al, 2005). These results indicate several orders of magnitude more activity than the parent oleanolic acid molecule. Some of the reasons include: activation of Nrf2 to induce antioxidant pathways provides important protection against oxidative stress and inflammation. Compounds related to RTA402 may also provide significant benefits similar to those provided by RTA402 in this application. Accordingly, these related compounds may be used to treat and/or prevent diseases such as: renal or renal disease (RKD), insulin resistance, diabetes, endothelial dysfunction, fatty liver disease, cardiovascular disease (CVD) and related conditions.

TABLE 1 inhibition of IFN γ -induced NO production

Honda et al (2002), incorporated herein by reference, discuss the synthesis of CDDO-MA. The synthesis of CDDO-EA and CDDO-TFEA was proposed by Yates et al (2007), which is incorporated herein by reference, and is shown in scheme 1 below.

Polymorphic forms of CDDO-Me

Polymorphic forms of the compounds of the invention, for example forms A and B of CDDO-Me, may be used according to the methods of the invention. Form B showed unexpectedly better bioavailability than form a (figure 15). Specifically, if monkeys orally ingest both forms of gelatin capsules at equal doses, the bioavailability of CDDO-Me form B is higher than form a (U.S. patent application No. 12/191,176, filed 8/13 of 2008).

The "form A" (RTA-402) of CDDO-Me is unsolvated (non-hydrated) and is characterized by a unique crystal structure having the space group P4 as well as a packing structure₃2₁2 (number 96) unit cell size ofThree molecules are stacked in a helical fashion along the crystalline b-axis by virtue of the stacking structure. In some embodiments, form a can also be characterized by an X-ray powder diffraction (XRPD) pattern (cuka) comprising significant diffraction peaks at approximately 8.8, 12.9, 13.4, 14.2, and 17.4 ° 2 Θ. In some variations, the X-ray powder diffraction pattern (CuK α) of form a is substantially as shown in fig. 12A and 12B.

The CDDO-Me 'form B' is different from the form A, the form B is a single phase and lacks definite crystalsAnd (5) structure. The type B sample did not exhibit a long distance (more than about)) And (4) molecular relevance. Furthermore, the thermal analysis showed the glass transition temperature (T) of the B-type sample_g) Between about 120 c and about 130 c (fig. 14). Unlike it, disordered nanocrystalline phases do not show T_gShowing only the melting temperature (T)_m) Above this temperature, the crystal structure becomes liquid. Form B is characterized by an X-ray powder diffraction (XRPD) spectrum (FIG. 12c) distinct from that of form A (FIG. 12A or 12B). Form B has no well-defined crystalline structure and therefore no distinct XRPD peaks (e.g. the representative XRPD peak of form a), and is characterized by a general "halo" XRPD pattern which shows three or less dominant diffraction halos, and amorphous form B belongs to the "X-ray amorphous" solid and to the "glassy" material of such solids.

Forms A and B of CDDO-Me are readily prepared from various solutions of this compound. Form B can be prepared, for example, by rapid evaporation or slow evaporation in MTBE, THF, toluene or ethyl acetate. Form a can be made by several methods including fast evaporation, slow evaporation or slow cooling of CDDO-Me in ethanol or methanol. CDDO-Me was prepared from acetone solution, if it was fast evaporation to make its form a, if it was slow evaporation to make its form B.

Various characterization methods can be used together to distinguish between CDDO-Me forms A and B, as well as other CDDO-Me forms. Illustrative of techniques suitable for identifying them are solid-state Nuclear Magnetic Resonance (NMR), X-ray powder diffraction (compare fig. 12A and 12B with fig. 12C), X-ray diffraction crystal analysis, Differential Scanning Calorimetry (DSC) (compare fig. 13 with fig. 14), dynamic vapor adsorption/Desorption (DVS), Karl Fischer analysis (KF), hot stage microscopy, modulated differential screening calorimetry, fourier transform infrared spectroscopy (FT-IR), and raman spectroscopy. For example, analysis of XRPD and DSC data can identify form a, form B, and the hemibenzoate (hemibenzanate) form of CDDO-Me (U.S. patent application No. 12/191,176 filed on 8/13 of 2008).

Additional details of the CDDO-Me polymorphic form are found in united states provisional application No. 60/955,939 filed on 8/15, 2007 and united states non-provisional application No. 12/191,176 filed on 8/13, 2008, both of which are incorporated herein by reference in their entirety.

Use of triterpenes for treating chronic kidney disease, insulin resistance/diabetes, endothelial dysfunction/cardiovascular disease

The compounds and methods of the present invention are useful for treating a variety of renal/renal diseases, including acute and chronic indications. Generally, the method comprises administering to the subject a pharmaceutically effective amount of a compound of the invention.

Inflammation contributes significantly to Chronic Kidney Disease (CKD). There is also a close mechanistic link between oxidative stress and renal dysfunction. The NF-. kappa.B signaling pathway plays an important role in the progression of CKD, because NF-. kappa.B regulates the transcription of MCP-1, and MCP-1 is a chemokine responsible for recruiting single cells or macrophages, triggering inflammation, and ultimately damaging the kidney (Wardle, 2001). The Keap1/Nrf2/ARE pathway controls transcription of several genes encoding antioxidant enzymes, including heme oxygenase-1 (HO-1). Deletion of the female mouse Nrf2 gene can lead to lupus-like glomerulonephritis (Yoh et al, 2001; Ma et al, 2006). In addition, several studies have shown that the response caused by renal injury and inflammation induces the expression of HO-1, and that this enzyme and its products (bilirubin and carbon monoxide) have a renal protective effect (Nath et al, 2006).

The glomeruli and the surrounding glomerular capsule constitute the basic functional units of the kidney. Glomerular Filtration Rate (GFR) is a standard measure of renal function. Creatinine clearance is often used as a measure of GFR, while serum creatinine content is often used as a surrogate measure of creatinine clearance. For example, an excessively high serum creatinine content is generally considered to be renal insufficiency, and a decrease in serum creatinine with time is considered to be an improvement in renal function. The normal level of creatinine in the blood is about 0.6 to 1.2 milligrams (mg) per deciliter (dl) for adult males and about 0.5 to 1.1 milligrams per deciliter for adult females.

Acute Kidney Injury (AKI) can occur following ischemic reperfusion, treatment with certain agents such as cisplatin and rapamycin, and intravenous injection of radiocontrast agents for medical imaging. Like CKD, inflammation and oxidative stress contribute to acute kidney injury. The molecular mechanisms by which radiocontrast agents induce renal disease (RCN) are not well understood, however, it is likely that a combination of factors, including long-term vasoconstriction, autoregulation of damaged kidneys, and direct toxicity of the contrast agents, contribute to renal failure (Tumlin et al, 2006). Vasoconstriction results in reduced renal blood flow and causes ischemia reperfusion and the production of reactive oxygen species. These conditions strongly induce HO-1, which has been shown to prevent ischemia reperfusion injury in several different organs including the kidney (Nath et al, 2006). In particular, it has been demonstrated that induction of HO-1 can exert a protective effect in a rat model of RCN (Goodman et al, 2007). Reperfusion is in part induced by activation of NF- κ B signaling to induce an inflammatory response (Nichols, 2004). Targeting NF- κ B has been proposed as a therapeutic strategy to prevent organ damage (Zingarelli et al, 2003).

Without being bound by theory, the efficacy of the compounds of the invention (e.g., RTA 402) results primarily from the addition of α, β -unsaturated carbonyl groups. In vitro assays, most of the activity of the compounds can be inhibited by the addition of: dithiothreitol (DTT), N-acetylcysteine (NAC), Glutathione (GSH), a thiol-containing moiety that interacts with an α, β -unsaturated carbonyl group (Wang et al, 2000; Ikeda et al, 2003; 2004; Shishodia et al, 2006). Biochemical assays have demonstrated that RTA402 interacts directly with and inhibits the activity of a key cysteine residue (C179) on IKK β (see below) (Shishodia et al, 2006; Ahmad et al, 2006). IKK β controls activation of NF-. kappa.B via the "classical" pathway, which involves the induction of I.kappa.B degradation via phosphorylation, releasing NF-. kappa.B dimers to the nucleus. In macrophages, this pathway is responsible for the production of a number of proinflammatory molecules in response to TNF α and other proinflammatory stimuli.

RTA402 also suppresses JAK/STAT signal pathways at multiple levels. Following activation by ligands (e.g., interferons and interleukins), JAK proteins are recruited to transmembrane receptors (e.g., IL-6R). JAKs then phosphorylate the intracellular portion of the receptor, recruiting STAT transcription factors. Subsequently, JAKs phosphorylate STATs into dimers and translocate to the nucleus where transcription of several genes associated with inflammation is activated. RTA402 inhibits constitutive STAT3 phosphorylation and STAT3 phosphorylation induced by IL-6 and dimer formation and binds directly to STAT3(C259) and cysteine residues in the kinase domain of JAK1 (C1077). Biochemical assays have also established that triterpenoids interact directly with critical cysteine residues on Keap1 (Dinkova-Kostova et al, 2005). Keap1 is an actin-linked protein that sequesters the transcription factor Nrf2 in the cytoplasm under normal conditions (Kobayashi and Yamamoto, 2005). Oxidative stress oxidizes regulatory cysteine residues on Keap1 and releases Nrf 2. Nrf2 then translocates to the nucleus, binding to the Antioxidant Response Element (ARE), activating transcription of many antioxidant and anti-inflammatory genes. Another target of the Keap1/Nrf2/ARE pathway is heme oxygenase-1 (HO-1). HO-1 breaks down heme into bilirubin and carbon monoxide, and has a number of antioxidant and anti-inflammatory effects (Maines and Gibbs, 2005). Recently, triterpenoids such as RTA402 have been shown to be effective in inducing HO-1(Liby et al, 2005). RTA402 and many structural analogs have been shown to be effective inducers of expression of other secondary proteins (Yates et al, 2007).

RTA402 is a potent inhibitor of NF-. kappa.B activation. Furthermore, RTA402 activated the Keap1/Nrf2/ARE pathway and induced HO-1 expression. RTA402 has been shown to be active in two animal models of AKI, as described below. In addition, it has been found that most patients who have received RTA402 treatment have a reduced serum creatinine level and also an improved glomerular filtration rate (see examples below). Significant improvements have been found in phase II studies in diabetic nephropathy patients. This finding suggests that RTA402 can be used to improve renal function in diabetic nephropathy patients by inhibiting renal inflammation and increasing glomerular filtration rate.

As mentioned above, both diabetes and essential hypertension are major risk factors for the development of chronic kidney disease, and ultimately, renal failure. These two symptoms, along with indications for systemic cardiovascular diseases such as hyperlipidemia, are common in the same patient, especially in patients with clinical obesity. Although the underlying etiology is not completely clear, it has been suggested that vascular endothelial dysfunction is an important pathological factor in systemic cardiovascular disease, chronic renal disease, and diabetes (see, e.g., zocicali, 2006). It has also been suggested that acute or chronic oxidative stress in vascular endothelial cells can lead to endothelial dysfunction and is closely linked to the chronic inflammatory process, and thus, if a drug can relieve the oxidative stress of vascular endothelium and its associated inflammation, the drug may alleviate the dysfunction and restore the endothelial system balance. Without being bound by theory, the compounds of the invention have shown extraordinary ability to improve the clinical parameters in patients with abnormal values of these parameters by stimulating endogenous antioxidant mechanisms regulated by Nrf2, parameters associated with renal function (e.g., serum creatinine and estimated glomerular filtration rate), parameters associated with glycemic control and insulin resistance (e.g., heme A1c), and parameters associated with systemic cardiovascular disease (e.g., circulating endothelial cells). Currently, it is often necessary to use combination therapies in these patients to improve glycemic control and cardiovascular disease, including angiotensin converting enzyme inhibitors or angiotensin II receptor blockers to reduce hypertension and slow the progression of chronic renal disease. The compounds of the invention enable meaningful clinical improvement of the measure of all these parameters, in particular renal function parameters, simultaneously. Thus, these compounds are a significant improvement over existing therapies. In certain aspects, the compounds of the invention may treat these conditions as one therapy, or in combination with other therapies, but with fewer therapies than current combination therapies.

These findings also suggest that administration of RTA402 can be used to protect a patient from renal injury, such as that caused by exposure to radiocontrast agents (e.g., radiocontrast-induced nephropathy) and other conditions. In one aspect, the compounds of the invention are useful for treating acute renal injury induced by ischemia reperfusion and/or chemotherapy. For example, the results shown in examples 2 and 3 below demonstrate that RTA402 is protective in animal models of ischemia reperfusion and chemotherapy-induced acute kidney injury.

Serum creatinine levels were measured in several animal models receiving RTA402 treatment. It was observed that the serum creatinine levels in the treated animals of cynomolgus monkeys, beagle dogs, and spra-dawley rats were significantly reduced compared to the baseline or control animals (fig. 3A-D). This effect was observed in rats using RTA402 in two different forms, crystalline and amorphous.

RTA402 can reduce serum creatinine levels in a patient. For example, it is observed that a cancer patient treated with RTA402 has improved disease. Nephrotoxicity in humans is a dose-limiting side effect of cisplatin treatment. It is believed by scientists that cisplatin-induced damage to the proximal tubule is mediated by increased inflammation, oxidative stress, and apoptosis (Yao et al, 2007). Serum creatinine levels have been measured in chronic renal disease (CKD) patients participating in the RTA402 public label phase II clinical trial (example 6). The study was designed to have multiple endpoints, categories insulin resistance, endothelial dysfunction/CVD, CKD, and include measurement of heme A1c (A1c), which is widely used at the phase 3 endpoint of glycemic control.

Glycated hemoglobin (A1c) is a small fraction of hemoglobin bound by glucose, also known as glycosylated hemoglobin or glucosylated hemoglobin. High Performance Liquid Chromatography (HPLC) is used to separate A1c from other heme a components in blood by charge and size differences. Since A1c is not affected by short-term fluctuations in blood glucose concentration (e.g., a three meal diet), blood samples can be drawn for the A1c test regardless of whether or not a meal has been consumed. Healthy and non-diabetic patients have an A1c content of less than 7% of total heme, with a normal range of 4-5.9%. Patients with poorly controlled diabetes can be 8.0% or higher. It has been shown that if the A1c content can be kept around 7%, diabetic complications can be delayed or prevented.

Six months of treatment with the most recently approved drug typically reduced the A1c level by only 0.4 to 0.80, with a generally lesser degree of improvement on day 28. Six months after treatment with the two approved drugs sitagliptin and pramlintide acetate, the reduction in glycated hemoglobin is as follows (Aschner et al, 2006; Goldstein et al, 2007; Pullman et al, 2006).

In contrast, treatment of refractory diabetes with RTA402, except for standard of care, decreased A1c within 28 days. The treatment showed: the reduction in treatment-intended patients was 0.34 (n-21) and in high baseline (baseline value ≧ 7.0) patients was 0.50 (n-16). These results are described in more detail in the examples section below, with reference to fig. 6 and 7.

In another aspect, the compounds of the present invention may also be used to improve insulin sensitivity and/or improve glycemic control. For example, in the study detailed in example 6, the results of hyperinsulinemic euglycemic clamp tests showed that RTA402 treatment improved glycemic control. The euglycemic clamp test for hyperinsulinemia is the standard method for studying and quantifying insulin sensitivity. The method measures the amount of glucose required to compensate for the increase in insulin without causing hypoglycemia (DeFronzo et al, 1979).

A typical procedure is as follows: at 10-120mU/m²Rate per minute, insulin infusion via peripheral vein. To compensate for insulin infusion, 20% glucose was infused to maintain blood glucose levels between 5 and 5.5 mmol/l. Every 5 to 10 minutes, blood glucose levels were measured to determine the glucose infusion rate.

It is generally advantageous to assess hepatic responses with low-dose insulin infusions, and peripheral (i.e., muscle and fat) insulin action with high-dose insulin infusions.

The results obtained are generally evaluated as follows: the rate of glucose infusion determines the insulin sensitivity at the last 30 minutes of the experiment. If a high rate is required (7.5 mg/min or higher), the patient is insulin sensitive. A very low rate (4.0 mg/min or less) indicates that the body is resistant to the action of insulin. Between 4.0 and 7.5 mg/min, the results may be ambiguous and may suggest "impaired glucose tolerance" as an early sign of insulin resistance.

The methods within the present invention can be used to improve kidney function. Example 6 shows that treatment with RTA402 improves six indicators of renal function and renal status, including serum creatinine-based eGFR, creatinine clearance, BUN, cystatin C, adiponectin, and angiotensin II. RTA402 was also shown to increase GFR in a dose-dependent manner with high response rates (86%; n-22). Fig. 9 also shows that the improvement in GFR was reversible at 28 days after drug withdrawal.

In some embodiments, the methods of treatment of the present invention may improve the levels of adiponectin and/or angiotensin II. Adiponectin is generally elevated with angiotensin II in DN patients and is associated with the severity of renal disease. Adiponectin (also known as Acrp30, apM1) is a hormone that regulates many metabolic processes including glucose regulation and fatty acid catabolism. Adipose tissue secretes adiponectin into the bloodstream, which is rich in adiponectin relative to many other hormones. The content of this hormone in adults is inversely related to the percentage of body fat, but the association is not clear in infants and young children. This hormone may play a role in inhibiting metabolic disorders that may cause type 2 diabetes, obesity, atherosclerosis and non-alcoholic fatty liver disease (NAFLD). Adiponectin can be used to predict mortality from a variety of causes and end stage renal disease in patients with DN.

The compounds and methods of the present invention are useful in a variety of contexts for the treatment of cardiovascular disease (CVD). The treatment methods of the invention have been found to reduce Circulating Endothelial Cells (CEC) in patients. CEC is a marker of endothelial dysfunction and vascular injury. Endothelial dysfunction is a systemic inflammatory process associated with cardiovascular and peripheral organ injury. CEC increases are usually associated with the onset, progression and death of CVD, as well as chronic kidney disease and a decrease in GFR, and their normal level has consistently been ≦ 5 cells/mL.

Endothelial dysfunction is typically characterized by an inability of arteries and arterioles to fully dilate in response to an appropriate stimulus, and thus, there is a detectable difference between subjects with endothelial dysfunction and those with healthy normal endothelium. This difference can be detected in a number of ways, including acetylcholine iontophoresis, intra-arterial administration of various vasoactive drugs, localized heating of the skin, or inflating a blood pressure cuff to high pressure to induce temporary arterial occlusion. Detection may also be performed in the coronary arteries. These techniques are believed to stimulate the endothelium to release Nitric Oxide (NO) and possibly other substances that may diffuse into the surrounding vascular smooth muscle, causing vasodilatation.

For example, according to the phase II study results (example 6), patients treated 28 days later with RTA402 had reduced cardiovascular inflammation markers as indicated by a reduced number of circulating endothelial cells. CEC reduction in the intended treatment group (n ═ 20) was 27%; the high baseline group (n ═ 14) was 40% less (p ═ 0.02), and CEC levels were normal in nine patients after treatment. These results are consistent with a reversal of endothelial dysfunction.

It has been found that the treatment methods of the invention can reduce matrix metallopeptidase-9 (MMP-9), soluble adhesion molecules, and tumor necrosis factor (TNF α) in a substantial proportion of patients. High levels of these substances are associated with cardiovascular adverse consequences.

Pharmaceutical formulations and routes of administration

The compounds of the present invention are administered to a patient according to a general drug administration regimen, taking into account drug toxicity, if any. And contemplates repeating the treatment cycle as needed.

The compounds of the invention can be administered via a variety of methods, such as orally or by injection (subcutaneously, intravenously, intraperitoneally, etc.). Depending on the application route, the active compounds can be protected from acids or other natural conditions of the deactivating compounds by a coating. It may also be administered in a continuous infusion or infusion mode at the site of disease or injury. Specific formulation examples, found in U.S. patent application No. 12/191,176 filed on 13/8/2008 (which is incorporated herein by reference in its entirety), include polymer-based CDDO-Me dispersions that have shown improved oral bioavailability. It will be appreciated by those skilled in the art that other manufacturing methods may produce compounds having properties and uses comparable to the dispersions of the present invention (see Repka et al, 2002, and references cited therein). Such alternative methods include, but are not limited to, solvent evaporation, extrusion (e.g., hot melt extrusion), and other techniques.

If the therapeutic compound is not administered parenterally, it may be desirable to protect the coating with, or co-administer with, a substance to avoid loss of activity. For example, the therapeutic compound may be administered to the patient in a suitable carrier, such as a liposome or diluent. Pharmaceutically acceptable diluents include saline and aqueous buffer solutions. Liposomes include water-in-oil-in-water CGF emulsions as well as conventional liposomes (Strejan et al, 1984).

The therapeutic compounds may also be administered parenterally, intraperitoneally, intraspinally, or intracerebrally. Dispersions can be made in glycerol, liquid polyethylene glycols and mixtures thereof and in oils; . Under ordinary conditions of storage and use, the preparation may contain a preservative to prevent the growth of microorganisms.

Pharmaceutical compositions suitable for injectable administration include sterile aqueous solutions (where the drug is water-soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the composition must be sterile and must be fluid for easy injection with a syringe. Has stable property and is preserved under the conditions of production and storage to prevent the pollution of microorganisms such as bacteria, fungi and the like. The carrier may be a solvent or dispersion medium containing: such as water, ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. For example, proper fluidity can be maintained by coating with lecithin or the like, by maintaining the desired particle size of the dispersion in the case of dispersion, or by using a surfactant or the like. Microbial contamination can be prevented by using various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, sodium chloride or polyalcohols such as mannitol and sorbitol in the composition. Prolonged absorption can be achieved by including in the injectable compositions an absorption delaying agent, for example, aluminum monostearate or gelatin.

Sterile injectable solutions can be prepared in the following manner: the desired amount of the therapeutic compound is dissolved in a suitable solvent, one or several of the ingredients listed above are added as required, after which it is filter sterilized. The therapeutic compound is typically dispersed in a sterile vehicle which contains a basic dispersion medium and the required other ingredients from those listed above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods are vacuum drying and freeze-drying to produce a powder of the active ingredient (i.e., the therapeutic compound) and any desired additional ingredient from a solution thereof which has been sterile-filtered.

The therapeutic compound may be administered orally, for example, using an inert diluent or an ingestible food carrier. The therapeutic compound and other ingredients may be contained in hard or soft gelatin capsules, compressed into tablets or added directly to the diet of the patient. The therapeutic compounds for oral administration may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, cachets and the like. The percentage of therapeutic compound in the composition and formulation can, of course, vary. The therapeutic compound is present in the pharmaceutical compositions for such therapeutic uses in an amount to obtain a suitable dosage.

Parenteral compositions formulated in dosage unit form are particularly advantageous for ease of administration and uniformity of dosage. Dosage unit form, as used herein, refers to physically discrete units suitable as unitary dosages for the subjects to be treated; each unit containing a predetermined quantity of therapeutic compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. The specification for the dosage unit forms of the invention is dictated by and directly determined by the following conditions: (a) the unique characteristics of the therapeutic compound and the particular therapeutic effect to be achieved, and (b) the inherent limitations of the formulation techniques used to treat the selected condition in the patient.

The therapeutic compounds may also be applied topically to the skin, eye or mucosa. If local delivery to the lung is desired, the therapeutic compound may be formulated as a dry powder or aerosol for administration by inhalation.

The actual dosage used for administering a compound of the present invention or a composition comprising a compound of the present invention to a subject may be determined by physical and physiological factors such as age, sex, body weight, severity of symptoms, type of disease being treated, previous or current therapeutic intervention, self-morbidity of the subject, and the route of administration. These factors can be determined by the skilled artisan. The concentration of the active ingredient in the composition is generally determined by the practitioner responsible for administration, as well as the appropriate dosage to be administered to the individual subject. The dosage may be adjusted by the individual physician, if any complications arise.

In some embodiments, a pharmaceutically effective amount is about 0.1mg to about 500mg of the compound per daily dose. In some variations, the daily dose is from about 1mg to about 300 mg of the compound. In some variations, the daily dose is from about 10mg to about 200mg of the compound. In some variations, the daily dose is about 25mg of the compound. In other variations, the daily dose is about 75mg of the compound. In still other variations, the daily dose is about 150mg of the compound. In a further variation, the daily dose is from about 0.1mg to about 30mg of the compound. In some variations, the daily dose is from about 0.5mg to about 20 mg of the compound. In some variations, the daily dose is from about 1mg to about 15mg of the compound. In some variations, the daily dose is from about 1mg to about 10mg of the compound. In some variations, the daily dose is from about 1mg to about 5mg of the compound.

In some embodiments, the pharmaceutically effective amount is 0.01 to 25mg of compound per kg of body weight per daily dose. In some variations, the daily dose is 0.05 to 20 mg compound per kg body weight. In some variations, the daily dose is 0.1 to 10mg compound per kg body weight. In some variations, the daily dose is 0.1 to 5mg compound/kg body weight. In some variations, the daily dose is 0.1 to 2.5 mg compound/kg body weight.

In some embodiments, the pharmaceutically effective amount is 0.1 to 1000mg of compound per kg of body weight per daily dose. In some variations, the daily dose is 0.15 to 20 mg compound per kg body weight. In some variations, the daily dose is 0.20 to 10mg compound per kg body weight. In some variations, the daily dose is 0.40 to 3mg compound per kg body weight. In some variations, the daily dose is 0.50 to 9mg compound per kg body weight. In some variations, the daily dose is 0.60 to 8mg compound per kg body weight. In some variations, the daily dose is 0.70 to 7mg compound/kg body weight. In some variations, the daily dose is 0.80 to 6mg compound per kg body weight. In some variations, the daily dose is 0.90 to 5mg compound per kg body weight. In some variations, the daily dose is from about 1mg to about 5mg of compound per kilogram of body weight.

An effective dose is typically from about 0.001mg/kg to about 1,000mg/kg, from about 0.01mg/kg to about 750mg/kg, from about 0.1mg/kg to about 500mg/kg, from about 0.2mg/kg to about 250mg/kg, from about 0.3mg/kg to about 150mg/kg, from about 0.3mg/kg to about 100mg/kg, from about 0.4mg/kg to about 75mg/kg, from about 0.5mg/kg to about 50mg/kg, from about 0.6mg/kg to about 30mg/kg, from about 0.7mg/kg to about 25mg/kg, from about 0.8mg/kg to about 15mg/kg, from about 0.9mg/kg to about 10mg/kg, from about 1mg/kg to about 5mg/kg, from about 100mg/kg to about 500mg/kg, From about 1.0mg/kg to about 250mg/kg or from about 10.0mg/kg to about 150mg/kg, in a single or more dose per day for one or more days (depending of course on the mode of administration and on the factors discussed above). Other suitable dosage ranges include 1mg to 10,000 mg daily, 100mg to 10,000 mg daily, 500mg to 10,000 mg daily, and 500mg to 1,000mg daily. In some particular embodiments, the daily amount administered is less than 10,000 mg, for example in the range of 750mg to 9,000 mg per day.

The effective dose may be less than 1 mg/kg/day, less than 500 mg/kg/day, less than 250 mg/kg/day, less than 100 mg/kg/day, less than 50 mg/kg/day, less than 25 mg/kg/day, less than 10 mg/kg/day or less than 5 mg/kg/day. It may also be in the range of 1 mg/kg/day to 200 mg/kg/day. For example, a unit dose for treating a diabetic patient is an amount that reduces blood glucose by at least 40% compared to an untreated subject. In another embodiment, the unit dose is that amount required to lower blood glucose to within ± 10% of the blood glucose level of a subject without diabetes.

In other non-limiting examples, the dose per administration is about 1 microgram/kg/body weight, about 5 microgram/kg/body weight, about 10 microgram/kg/body weight, about 50 microgram/kg/body weight, about 100 microgram/kg/body weight, about 200 microgram/kg/body weight, about 350 microgram/kg/body weight, about 500 microgram/kg/body weight, about 1 mg/kg/body weight, about 5 mg/kg/body weight, about 10 mg/kg/body weight, about 50 mg/kg/body weight, about 100 mg/kg/body weight, about 200 mg/kg/body weight, about 350 mg/kg/body weight, about 500 mg/kg/body weight, about, To about 1000 mg/kg/body weight or more, any range derivable therein. In a non-limiting example, the ranges derived from the numbers listed herein are: about 1 mg/kg/body weight to about 5 mg/kg/body weight, about 5 mg/kg/body weight to about 100 mg/kg/body weight, about 5 micrograms/kg/body weight to about 500 mg/kg/body weight, and the like, can be administered based on the above numbers.

In certain embodiments, a pharmaceutical composition of the invention may comprise, for example, at least about 0.1% of a compound of the invention. In other embodiments, the compounds of the present invention may comprise from about 2% to about 75%, or between about 25% and about 60%, by weight of the unit, and any range derivable therein.

Administration of a single dose or multiple doses is contemplated. The desired time interval for administration of multiple doses can be determined by one of ordinary skill in the art and requires only routine experimentation. For example, the agent may be administered to the subject once every 12 hours, twice daily. In some embodiments, the administration is once daily.

The agents may be administered on a routine schedule. A routine schedule herein refers to a predetermined specified period of time. The time periods covered by the routine schedule may be the same or different in length, as long as the schedule is predetermined. For example, a routine schedule may set a set number of days or weeks of administration twice daily, bidaily, every three days, every four days, every five days, every six days, weekly, monthly, or intervening. Alternatively, the predetermined routine schedule may set the first week to be administered twice daily, followed on a daily basis for months, etc. In other embodiments, the agents provided herein can be administered orally, and the time of oral administration may or may not be dependent on eating. Thus, the medicament may be taken sooner and/or later, regardless of whether or not food has been consumed or is about to be consumed.

Non-limiting specific formulations include CDDO-Me polymer dispersions (see U.S. patent application No. 12/191,176 filed on 8/13/2008, which is incorporated herein by reference), some of which reports exhibit bioavailability higher than micronized form a or nanocrystalline form a formulations. Furthermore, it has been demonstrated that polymer dispersion based formulations provide a more surprising improvement in oral bioavailability over micronized type B formulations. Such as methacrylic acid copolymer, type C and HPMC-P formulations, showed the greatest bioavailability in experimental monkeys.

Combination therapy

In addition to being used as monotherapy, the compounds of the invention may also be used in combination therapy. An effective combination therapy may comprise a single composition or pharmaceutical formulation of two agents, or two different compositions or formulations administered simultaneously, wherein one composition comprises a compound of the invention and the other composition comprises a second agent. Alternatively, the therapy may be administered before or after the treatment with the other agent, at intervals ranging from minutes to months.

Various combinations may be employed, with "a" representing a compound of the invention and "B" representing a second agent, non-limiting examples of which are as follows:

A/B/A B/A/B B/B/A A/A/B A/B/B B/A/A A/B/B/B B/A/B/B

B/B/B/A B/B/A/B A/A/B/B A/B/A/B A/B/B/A B/B/A/A

B/A/B/A B/A/A/B A/A/A/B B/A/A/A A/B/A/A A/A/B/A

other anti-inflammatory agents are contemplated for administration in conjunction with the treatment of the present invention, for example, other COX inhibitors including arylcarboxylic acids (salicylic acid, acetylsalicylic acid, diflunisal, choline magnesium trisalicylate, salicylates, paracetamol (benomylate), flufenamic acid, mefenamic acid, meclofenamic acid (triflumic acid)), arylalkanoic acids (diclofenac), fenclofenac (fenclofenac), alclofenac (alclofenac), fentiaprofenic acid (fentiazac), ibuprofen (ibuprofen), flurbiprofen (flurbiprofen), ketoprofen (ketoprofen), naproxen (naproxen), fenoprofen (fenoprofen), fenbufen (fenbufen), suprofen (fenprofen), suprofen (suprofen), indoprofen (indoprofen), tiaprofenic acid (tiaprofenic acid), fenpropicin (fenpropicin), piroprofen (piroprofen), ibuprofen (oxyphenicol (bromfenac), ibuprofen (bromfenac), and ibuprofen (bromfenac), ibuprofen (ibuprofen), ibuprofen (bromfenac), and ibuprofen (bromfenac), ibuprofen (bromphen (brom, Oxyphenbutazone, azapropazone (azapropazone), fipronone (feprazone), piroxicam (piroxicam) and isoxicam (isoxicam). See U.S. patent 6,025,395, which is incorporated herein by reference.

Food and nutritional supplements that are reported to be beneficial for the treatment or prevention of the following diseases may be used in conjunction with the compounds of the present invention: parkinson's disease, alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, rheumatoid arthritis, inflammatory bowel disease, and all other diseases where an excess of Nitric Oxide (NO) or prostaglandin production is implicated as causative agent, such as acetyl levocarnitine, octacosanol, evening primrose oil, vitamin B6, tyrosine, phenylalanine, vitamin C, levodopa or a combination of several antioxidants.

Other specific secondary therapies include immunosuppressants (for transplantation of autoimmune-related RKD), antihypertensive drugs (for hypertension-related RKD, e.g., angiotensin converting enzyme inhibitors and angiotensin receptor blockers), insulin (for diabetic RKD), lipid or cholesterol lowering agents (e.g., HMG-CoA reductase inhibitors such as atorvastatin (atorvastatin) or simvastatin (simvastatin)), therapies for hyperphosphatemia or CKD-related hyperparathyroidism (e.g., sevelamer acetate, cinacalcet (cinacalcet)), dialysis, and food restrictions (e.g., protein, salt, fluid, potassium, phosphorus).

VIII example

The following examples are included to illustrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventor to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. Those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.

Example 1: materials and methods

Triterpenoids are synthesized as previously described by Honda et al (1998), Honda et al (2000b), Honda et al (2002), and Yates et al (2007), all of which are incorporated herein by reference.

Example 2: experimental results of ischemia reperfusion of mice

In the mouse model of acute ischemic renal failure, the renal arteries were clamped for approximately twenty minutes. Thereafter, the clamps are removed and the kidneys are reperfused with blood. Ischemia reperfusion results in renal injury, which can be assessed by Blood Urea Nitrogen (BUN) levels, and reduced renal function, which increases BUN levels. As shown in fig. 1a-d, surgery-induced ischemia-reperfusion increased BUN content to around 2-fold. However, the BUN content of animals treated by oral administration of RTA402 at a dose of 2mg/kg once daily was significantly reduced (p < 0.01) compared to animals treated with vehicle, starting two days prior to surgery, similar to animals receiving sham surgery (FIGS. 1 a-c). The renal injury and inflammatory histology measures also improved significantly after RTA402 treatment (fig. 1 d). These data show that RTA402 can protect against ischemia reperfusion-induced tissue damage.

Example 3: experimental results of rat chemotherapy-induced acute kidney injury

In another model of acute kidney injury, cisplatin, an anti-cancer drug, was injected intravenously into rats. Nephrotoxicity in humans is a dose-limiting side effect of cisplatin treatment. Cisplatin-induced proximal tubular injury is thought to be mediated by increased inflammation, oxidative stress, and apoptosis (Yao et al, 2007). Rats treated with 6mg/kg of cisplatin as a single dose developed renal insufficiency, and were measured to have increased creatinine and BUN contents in blood. If rats were orally fed RTA402 at a dose of 10mg/kg on the day before cisplatin administration and continued daily, significant reductions in creatinine and BUN levels in the blood could be achieved (FIGS. 2 a-b). The renal histological evaluation demonstrated that the extent of proximal tubular injury was improved in RTA 402-treated animals compared to vehicle-only treated animals (fig. 2 c).

Example 4: serum creatinine content reduction in several species

During toxicity testing, serum creatinine was determined for several animal species treated with RTA 402. It was observed that serum creatinine levels were significantly reduced in cynomolgus monkeys, beagle dogs, and sper-dow rats compared to baseline values or control animals (fig. 3 a-d). This effect was also seen when the crystalline and amorphous RTA402 was administered to rats.

Example 5: serum creatinine reduction and eGFR enhancement in cancer patients

Serum creatinine was measured in patients participating in the RTA402 phase I clinical trial. These patients received a 5 to 1,300 mg/day dose of RTA402 once daily for 21 of 28 days. Eight days after treatment began, serum creatinine was found to decrease by more than 15% and continued until the end of the cycle (fig. 4A). Treatment of patients for six or more cycles with RTA402 can maintain this reduction in serum creatinine. In a group of patients with preexisting renal impairment (a baseline serum creatinine of at least 1.5 mg/dl), the serum creatinine levels were also significantly reduced following treatment with RTA 402. The serum creatinine levels in these patients decreased gradually throughout the cycle such that the level on day 21 was approximately 25% below baseline (fig. 4A). The following table summarizes the results.

Patients treated with RTA402 exhibited significant improvement in their estimated glomerular filtration rate (eGFR) (fig. 4B).

Figure 5 shows the results of eleven cancer patients after at least six months of treatment with RTA402, demonstrating a sustained improvement in eGFR. Some of the patients participated in the phase I study, others participated in the RTA402 (in combination with gemcitabine) study in pancreatic cancer patients. The results are summarized in Table 2 below.

Example 6 phase 2 study in diabetic nephropathy patients

Serum creatinine was determined in patients with chronic renal disease (CKD) who participated in the RTA402 public label phase II clinical trial. These patients received RTA402 once daily for 28 days in three doses: 25mg, 75mg and 150 mg.

This study was designed with multiple endpoints, categories of insulin resistance, endothelial dysfunction/CVD and CKD, summarized as follows:

the primary measure of the results of this study was to determine the effect of oral administration of three different dose strengths of RTA402 on glomerular filtration rate (estimated by the MDRD formula) in diabetic nephropathy patients.

The secondary outcome metrics include: (1) evaluating the safety and tolerability of the patient population for oral administration of three different doses of RTA 402; (2) after oral administration of three different doses of RTA402 to diabetic nephropathy patients, the effect of RTA402 on serum creatinine content, creatinine clearance and urinary albumin/creatinine ratio of these patients was evaluated; (3) evaluating the effect of oral administration of three different doses of RTA402 on heme A1c in all patients enrolled in the study and on the insulin response resulting from hyperinsulinemic euglycemic clamp test in patients enrolled in only one of the study centers; (4) the effect of three different doses of RTA402 on a marker panel of inflammation, renal injury, oxidative stress and endothelial cell dysfunction was evaluated.

The patient populations enrolled in the study all suffered from type 2 diabetes and CKD. Most patients have been diagnosed with a twenty year poor glycemic control. CKD is determined by an increase in serum creatinine (SCr) content. Most patients have been diagnosed with cardiovascular disease (CVD), and most are receiving standard of care (SOC) treatment for diabetes, CKD, and CVD (e.g., insulin, ACEI/ARB, β -blockers, diuretics, and statins). The baseline demographic information is summarized as follows:

age (age)

59

Course of diabetes mellitus (year)	15.4
		Diabetic nephropathy	100％
Non-renal diabetic complications1	100％
		Hypertension (hypertension)	100％
Hgb A1c(％)	7.9％
		Ineffective oral antihyperglycemic agents	90％
Using ACEI/ARB	80％
		Use of statins	50％

¹Including neuropathy and retinopathy

All values are mean values; n is 10; batch 1 Total 10 patients completed the study

Patient inclusion criteria were as follows: (1) diagnosing type 2 diabetes; (2) serum creatinine in the female is 1.3-3.0 mg/dl (115-265. mu. mol/l), including the threshold, and in the male is 1.5-3.0 mg/dl (133-265. mu. mol/l), including the threshold; (3) the patient must agree to take effective contraceptive means; (4) the urine pregnancy test results 72 hours before the patient receives the first dose of the drug must be negative; (5) patients are willing and able to coordinate with the whole of the scheme, and can communicate effectively; (6) patients were willing and able to provide written informed consent to participate in this clinical study.

Patient elimination criteria were as follows: (1) (ii) has type 1 (insulin-dependent; juvenile onset) diabetes; (2) patients with known non-diabetic nephropathy (patients who can receive both nephrosclerosis and diabetic nephropathy) or receive kidney transplants; (3) suffering from the following cardiovascular diseases: participating unstable angina pectoris and myocardial infarction in three months after research, and receiving coronary artery bypass grafting operation or percutaneous coronary angioplasty or stent in three months after research; transient ischemic attack occurs within three months after study participation; cerebrovascular accidents occurred within three months after participation in the study; suffering from obstructive valvular heart disease or hypertrophic cardiomyopathy; treatment of second or third degree atrioventricular block with a pacemaker was unsuccessful; (4) chronic (> 2 weeks) immunosuppressive therapy, including corticosteroids (excluding inhaled or nasal steroids), was required within three months after study participation; (5) evidence of liver dysfunction including total bilirubin > 1.5 mg/dl (> 26 μ M/l) or liver transaminase (aspartate transaminase [ AST ] or alanine transferase [ ALT ]) > 1.5 fold of the upper normal limit; (6) a female who is pregnant, nursing, or scheduled to become pregnant; (7) together with any other clinical conditions and which the investigator believes may be potentially dangerous to the patient's health during the study or may affect the study outcome; (8) known to be allergic to the components of the study drug; (9) known to be allergic to iodine; (10) within 30 days prior to study participation, receiving a diagnosis or intervention requiring contrast agent; (11) when any of the following drugs is used, there is a change or dose adjustment: during the first 3 months of study participation, ACE inhibitors, angiotensin II blockers, non-steroidal anti-inflammatory drugs (NSAIDs) or COX-2 inhibitors were used; other antihypertensive or antidiabetic drugs were administered within 6 weeks prior to study participation; (12) (ii) a drug or alcohol abuse experience or a positive result for any drug abuse test (positive urine drug test and/or breathalyzer test); (13) another clinical study involving study medication or marketed products has been, or will be, involved within 30 days prior to study participation; (14) due to language disability, mental retardation or impaired brain function, communication or cooperation with researchers is not possible.

By the end of 9 months in 2008, 32 out of 60 patients participated in the study, all but one patient received insulin and oral antihyperglycemic drugs meeting the regulatory standards.

It was observed that RTA402 plus care compliance with the guardian standard for 28 days decreased the percentage of glycated hemoglobin in refractory diabetic patients. This treatment showed a decrease of approximately 0.25 (n-56) in treatment-intended patients and a decrease of 0.50 (n-35) in high baseline (baseline value ≧ 7.0) patients. The reduction in the percentage of glycated hemoglobin as a function of baseline severity is shown in figure 6, while the reduction as a function of dose is shown in figure 7. Patients with advanced (stage 4) renal disease (GFR 15-29 ml/min) had an average percentage reduction of glycated hemoglobin of about 0.77. All reductions were statistically significant.

The hyperinsulinemic euglycemic clamp test showed that 28 days of treatment also improved glycemic control and insulin sensitivity in the patient as shown by the glucose clearance (GDR) assay. After 28 days of treatment, the GDR improved, with statistically significant improvement (P.ltoreq.0.02) in severely impaired patients (GDR < 4). Hyperinsulinemic euglycemic clamp tests were performed at baseline (day 1) and at the end of the study on day 28. The test measures the rate of Glucose (GINF) infusion required to compensate for the increased insulin content without causing hypoglycemia; the GDR can be derived using this value.

In short, the hyperinsulinemic euglycemic clamp test takes about 2 hours. At 10-120mU/m²The rate per minute is via peripheral intravenous infusion of insulin. To compensate for insulin infusion, 20% glucose needs to be infused to keep the blood glucose level between 5 and 5.5 mmol/l. Blood glucose levels were measured every 5 to 10 minutes to determine the glucose infusion rate. The rate of glucose infusion over the last 30 minutes of the experiment was used to determine insulin sensitivity, which was determined by the rate of glucose metabolism (M) (expressed in mg/kg/min).

Protocol guidelines for the hyperinsulinemic euglycemic clamp test are as follows:

1) the diet was discontinued for 8-10 hours before clamping.

2) Jaw test vital signs and body weight were measured the morning.

3) An retrograde line was made in one hand with a catheter 11/4 ", 18-20 for sampling.

4) A number IV tube with 2 three-way taps and a j-ring extension tube was prepared. The tube was inserted into a one liter bag containing 0.9% NaCl to facilitate operation under KVO (vein patency, approximately 10 ml/hour) until the procedure was initiated.

5) The heating pad is placed within the pillowcase with the pad separating the heating pad from the subject's hand (for collection of shunted arteriolized blood via intravenous catheterization).

6) The temperature generated by the heating pad (approximately 150F./65℃.) was monitored before and during the clamp test to maintain arterialization.

7) An 11/4' 18-20 catheter is used, and another pipeline is opened at the far end of the forearm opposite to the extraction side to be used as an infusion line; a number IV tube with 2 three-way taps was prepared.

8) A 500 ml bag containing 20% dextrose was hung and connected to the infusion side port.

9) Preparation of insulin infusion solutions

a) From a 500 ml bag containing 0.9% NaCl, 53 ml (50 ml over-run) of saline was removed and discarded

b) An 8 ml blood sample was drawn from the subject by aseptic technique and injected into the tiger top tube

c) The top tube of the tiger was centrifuged, 2 ml of serum was extracted and injected into a 500 ml bag containing 0.9% NaC

d) 100 units of insulin were added to the serum bag and mixed well (0.2U insulin/ml)

e) Connecting the IV pipe fitting and the two-way air hole angle nail, and putting the IV pipe fitting and the two-way air hole angle nail into a 0.9% NaCl bag

f) Placed on a Baxter pump

10) Time and draw all basal blood samples (fasting blood glucose baseline values were taken before preparation for insulin infusion).

11) The sensitizing dose and 60mU/m are calculated²Insulin infusion rate required for insulin infusion amount. Background insulin inhibits endogenous hepatic glucose production. The lean subject can be treated with 40mU/m²Is administered in an amount of 80mU/m to an obese subject suffering from insulin resistance²Amount of (2)And (5) preparing. 60mU/m²Should be sufficient to inhibit the BMI to be 27-40 kg/m²The recommended population of study subjects of (1). If the BMI is modified, the suggested 60mU/m²The amount of insulin infusion may need to be adjusted.

12) Samples of 0.5 ml were taken every five minutes and the glucose infusion rate (mg/kg/min) was determined or adjusted using the YSI glucometer reading. This protocol requires any additional laboratory tests, all of which increase blood volume requirements. The clamp test lasted 120 minutes, which is considered sufficient time to determine insulin sensitivity.

13) And labeling and storing the printed files of all YSI glucometers as source files.

14) The glucose infusion rate for the last 30 minutes of the euglycemic clamp test was adjusted with spatial correction. This will be used to determine the rate of glucose metabolism (Mmg/kg/min), which represents the subject's sensitivity to insulin.

As shown in fig. 8, RTA402 can reduce Circulating Endothelial Cells (CECs). Mean numbers of CECs expressed in cells/ml for the intended treatment group (ITT) and the high basal group before and 28 days after RTA treatment are shown. The reduction in the intended treatment group was about 20% and the reduction in the high basal group (> 5 CEC/ml) was about 33%. The reduction in the iNOS-positive CEC portion was about 29%. Normalization of the CEC values was observed in 11 patients (< 5 cells/ml) in 19 patients on the high baseline group.

CECs were isolated from whole blood using CD146 Ab (an antibody to the CD146 antigen, expressed in endothelial cells and leukocytes). After separation, CellSearch was used^TMCEC was identified systemically with CD105 Ab (antibody specific for endothelial cells) conjugated to FITC (fluorescein isothiocyanate). The leukocytes were stained with fluorescent conjugates of CD45 Ab and then gated out. For an overview of this process, see Blann et al, (2005), which is incorporated herein by reference in its entirety. CEC samples can be assessed for the presence of iNOS via immunostaining. RTA402 treatment reduced iNOS-positive CECs by approximately 29%, further showing that RTA402 reduced inflammation in endothelial cells.

RTA402 has been shown to significantly improve eight indicators of renal function and renal status, including serum creatinine-based eGFR (fig. 9), creatinine clearance, BUN (fig. 11A), serum phosphorus (fig. 11B), serum uric acid (fig. 11C), cystatin C, adiponectin (fig. 10A), and angiotensin II (fig. 10B). Adiponectin predicts mortality and end-stage renal disease in patients with DN for various reasons. Adiponectin and angiotensin II were elevated in DN patients and correlated with nephropathy severity (fig. 10A-B). FIGS. 11A-C show the effect on BUN, phosphorus and uric acid.

Patients treated with a higher dose (75 or 150 mg) of RTA402 had a modest increase in proteinuria (approximately 20% to 25%). This is consistent with multiple studies showing that better GFR performance is associated with increased proteinuria. For example, treatment with ramipril (ramipril) (ACE inhibitor) is more effective in slowing the rate of eGFR decline in long-term clinical studies in over 25,000 patients than with telmisartan (angiotensin receptor blocker) or a combination of ramipril (ramipril) and telmisartan (telmisartan) (Mann et al, 2008). In contrast, the ramipril (ramipril) test group increased proteinuria more than the other two test groups. Although the increase in proteinuria was minimal in the combination treatment trial group, the primary renal results obtained with either drug alone were superior to the combination treatment. Other studies have demonstrated that drugs that reduce GFR, such as ACE inhibitors, also reduce proteinuria (Lozano et al, 2001; Sengul et al, 2006). Other studies have demonstrated that drugs that acutely increase GFR, such as certain calcium channel blockers, can increase proteinuria to 60% during short-term dosing (Agodoa et al, 2001; Viberti et al, 2002).

All of the compositions and/or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and/or methods and in the steps or in the sequence of steps of the methods described herein without departing from the concept, spirit and scope of the invention. More specifically, it will be apparent that certain agents which are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.

IX. reference

The following references, which are specifically incorporated by reference herein, are provided to complement the illustrative procedural or other details herein.

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Claims (27)

1. Use of a compound having the structure:

2. the use of claim 1, wherein the subject has acute ischemic renal failure, chemotherapy-induced acute kidney injury, cancer, diabetic nephropathy, and/or chronic kidney disease.

3. The use according to claim 1, wherein at least a portion of the compound is in a crystalline form having a CuK α X-ray diffraction pattern as shown in figure 12A or figure 12B.

4. The use according to claim 1, wherein at least a portion of the compound is amorphous having a CuK α X-ray diffraction pattern having a halo peak at 13.5 ° 2 θ, as shown in figure 12C, and a T of 120 ℃ to 135 ℃_g。

5. The use as claimed in claim 4, wherein T is_gIs from 125 ℃ to 130 ℃.

6. The use according to claim 1, wherein the daily dose of the medicament comprises from 10mg to 200mg of the compound.

7. The use according to claim 4, wherein the daily dose of the medicament comprises from 0.1mg to 30mg of the compound.

8. The use of any one of claims 1-7, wherein the subject has or displays one or more symptoms of chronic kidney disease.

9. The use of any one of claims 1-7, wherein the compound or pharmaceutically acceptable salt thereof is used in the manufacture of a medicament for improving kidney function in a subject having diabetic nephropathy.

10. The use of any one of claims 1-7, wherein the subject has a reduced estimated glomerular filtration rate.

11. The use of any one of claims 1-7, wherein the subject has an increased level of serum creatinine.

12. The use of any one of claims 1-7, wherein the subject has an increased level of blood urea nitrogen.

13. The use of any one of claims 1-7, wherein the subject has increased levels of adiponectin in blood.

14. The use of any one of claims 1-7, wherein the subject has an increased level of angiotensin II.

15. The use of any one of claims 1-7, wherein the subject has insulin resistance or exhibits one or more symptoms of insulin resistance.

16. The use of claim 15, wherein the subject has an increased level of heme Alc.

17. The use of claim 15, wherein the subject has an increased blood glucose level.

18. The use of claim 15, wherein the subject has reduced insulin sensitivity as determined by the hyperinsulinemic euglycemic clamp test.

19. The use of claim 15, wherein the subject has reduced glucose clearance.

20. The use of any one of claims 1-7, wherein the compound or pharmaceutically acceptable salt thereof is used in the manufacture of a medicament for improving renal function in a subject having cardiovascular disease or exhibiting one or more symptoms of cardiovascular disease.

21. The use of claim 20, wherein the subject has an increased number of circulating endothelial cells in the blood.

22. The use of claim 21, wherein the circulating endothelial cells are iNOS positive circulating endothelial cells.

23. The use of any one of claims 1-7, wherein the subject is a human.

24. The use of any one of claims 1-7, wherein the medicament is formulated for oral, intra-arterial, or intravenous administration.

25. The use of claim 24, wherein the medicament is formulated as a hard or soft capsule or tablet.

26. The use of claim 24, wherein the medicament is formulated as a solid dispersion comprising (i) the compound and (ii) an excipient.

27. The use of claim 24, wherein the medicament is formulated to comprise (i) a therapeutically effective amount of the compound and (ii) an excipient, wherein the excipient is methacrylic acid-ethyl acrylate copolymer 1: 1.