ZA200301044B

ZA200301044B - Anticonvulsant derivatives useful for preventing the development of Type II diabetes mellitus and Syndrome X.

Info

Publication number: ZA200301044B
Application number: ZA200301044A
Authority: ZA
Inventors: Carlos Plata-Salaman; Jeffrey Crooke
Original assignee: Ortho Mcneil Pharm Inc
Priority date: 2000-07-07
Filing date: 2003-02-06
Publication date: 2004-05-06

Description

ANTICONVULSANT DERIVATIVES USEFUL FOR PREVENTING THE

DEVELOPMENT OF TYPE Il DIABETES MELLITUS AND SYNDROME X . CROSS REFERENCE TO RELATED APPLICATIONS . This application claims priority from United States provisional application Serial

No. 60/217,141 filed July 07, 2000 and United States provisional application Serial No. 60/270,022 filed, February 20, 2001, the contents of which are hereby incorporated by reference.

BACKGROUND OF THE INVENTION

Compounds of Formula (1):

X 1

RS CH,0OSO,NHR : R2 4 3

R R (I) are structurally novel antiepileptic compounds that are highly effective anticonvulsants in animal tests (MARYANOFF, B.E, NORTEY, S.0., GARDOCKI, J.F., SHANK, R.P.

AND DODGSON, S.P. J. Med. Chem. 1987, 30, 880-887; MARYANOFF, B.E,,

COSTANZO, M.J., SHANK, R.P., SCHUPSKY, J.J., ORTEGON, M.E., AND VAUGHT

J.L. Bioorg. Med. Chem. Lett. 1993, 3, 2653-2656; SHANK, R.P., GARDOCKI, J.F.,

VAUGHT, J.L., DAVIS, C.B., SCHUPSKY, J.J., RAFFA, R.B., DODGSON, S.J,

NORTEY, S.0., MARYANOFF, B.E. Epilepsia 1994, 35, 450-460; MARYANOFF BE,

COSTANZO MJ, NORTEY SO, GRECO MN, SHANK RP, SCHUPSKY JJ, ORTEGON

MP, VAUGHT JL. J. Med. Chem. 1998, 41, 1315-1343). These compounds are covered by three US Patents: No.4,513,006, No.5,242,942, and No0.5,384,327. One of these compounds 2,3:4,5-bis-O-(1-methylethylidene)-R-D-fructopyranose sulfamate known as topiramate has been demonstrated in clinical trials of human epilepsy to be ] effective as adjunctive therapy or as monotherapy in treating simple and complex partial seizures and secondarily generalized seizures (E. FAUGHT, B.J. WILDER, R.E. . RAMSEY, R.A. REIFE, L D. KRAMER, G.W. PLEDGER, R.M. KARIM et. al., Epilepsia 1995, 36 (S4), 33; S.K. SACHDEO, R.C. SACHDEOQ, R.A. REIFE, P. LIM and G.

PLEDGER, Epilepsia 1995, 36 (S4), 33; T.A. GLAUSER, Epilepsia 1999, 40 (S5),

S$71-80; R.C. SACHDEO, Clin. Pharmacokinet. 1998, 34, 335-346), and is currently marketed for the treatment of seizures in patients with simple and complex partial epilepsy and seizures in patients with primary or secondary generalized seizures in the

United States, Europe and most other markets throughout the world.

Compounds of Formula (1) were initially found to possess anticonvulsant ) activity in the traditional maximal electroshock seizure (MES) test in mice (SHANK,

R.P., GARDOCKI, J.F., VAUGHT, J.L., DAVIS, C.B., SCHUPSKY, J.J., RAFFA, R.B.,

DODGSON, S.J., NORTEY, S.0., and MARYANOFF, B.E., Epilepsia 1994, 35, 450- 460). Subsequent studies revealed that Compounds of Formula (1) were also highly effective in the MES test in rats. Topiramate was also found to effectively block seizures in several rodent models of epilepsy (J. NAKAMURA, S. TAMURA, T.

KANDA, A. ISHII, K. ISHIHARA, T. SERIKAWA, J. YAMADA, and M. SASA, Eur. J.

Pharmacol. 1994, 254. 83-89), and in an animal model of kindled epilepsy (A.

WAUQUIER and S. ZHOU, Epilepsy Res. 1996, 24, 73-77).

More recently compounds of formula (I) have been found to be effective for maintaining weight loss, as disclosed in WIPO publication WO00/61140, for the treatment of obesity, as disclosed in U.S. Patent No. 6,071,537 (WO 9800130), for lowering blood glucose levels, as disclosed in WIPO publication W0O00/61139 and for lowering lipids as disclosed in WIPO publication WO00/61137. Thakur et al in WIPO publication W099/44581 disclose the use of topiramate for the treatment of diabetes.

Type Il diabetes mellitus (non-insulin-dependent diabetes mellitus or NIDDM) is a metabolic disorder involving dysregulation of glucose metabolism and insulin resistance, and long-term complications involving the eyes, kidneys, nerves, and blood vessels. Type ll diabetes mellitus usually develops in adulthood (middle life or later) and is described as the body's inability to make either sufficient insulin (abnormal insulin secretion) or its inability to effectively use insulin (resistance to insulin action in target organs and tissues). More particularly, patients suffering from Type li diabetes mellitus have a relative insulin deficiency. That is, in these patients, plasma insulin . levels are normal to high in absolute terms, although they are lower than predicted for the level of plasma glucose that is present.

Type il diabetes mellitus is characterized by the following clinical signs or symptoms: persistently elevated plasma glucose concentration or hyperglycemia; polyuria; polydipsia and / or polyphagia; chronic microvascular complications such as retinopathy, nephropathy and neuropathy; and macrovascular complications such as hyperlipidemia and hypertension which can lead to blindness, end-stage renal disease, limb amputation and myocardial infarction.

Syndrome X, also termed Insulin Resistance Syndrome (IRS), Metabolic

Syndrome, or Metabolic Syndrome X, is a disorder that presents risk factors for the development of Type Il diabetes mellitus and cardiovascular disease including glucose intolerance, hyperinsulinemia and insulin resistance, hypertriglyceridemia, hypertension and obesity.

The diagnosis of Type II diabetes mellitus includes assessment of symptoms and measurement of glucose in the urine and blood. Blood glucose level determination is necessary for an accurate diagnosis. More specifically, fasting blood glucose level determination is a standard approach used. However, the oral glucose tolerance test (OGTT) is considered to be more sensitive than fasted blood glucose level. Type Il diabetes mellitus is associated with impaired oral glucose tolerance (OGT). The OGTT thus can aid in the diagnosis of Type Il diabetes mellitus, although generally not necessary for the diagnosis of diabetes (Emancipator K, Am J Clin

Pathol 1999 Nov;112(5):665-74; Type 2 Diabetes Mellitus, Decision Resources Inc.,

March 2000). The OGTT allows for an estimation of pancreatic beta-cell secretory function and insulin sensitivity, which helps in the diagnosis of Type Il diabetes mellitus and evaluation of the severity or progression of the disease (e.g., Caumo A, Bergman

RN, Cobelli C,. J Clin Endocrinol Metab 2000, 85(11):4396-402). More particularly, the

OGTT is extremely helpful in establishing the degree of hyperglycemia in patients with multiple borderline fasting blood glucose levels that have not been diagnosed as diabetics. In addition, the OGTT is useful in testing patients with symptoms of Type II diabetes mellitus where the possible diagnosis of abnormal carbohydrate metabolism has to be clearly established or refuted.

Thus, impaired glucose tolerance is diagnosed in individuals that have fasting . blood glucose levels less than those required for a diagnosis of Type Il diabetes mellitus, but have a plasma glucose response during the OGTT between normal and ‘ diabetics. Impaired glucose tolerance is considered a prediabetic condition, and impaired glucose tolerance (as defined by the OGTT) is a strong predictor for the development of Type Il diabetes mellitus (Haffner SM, Diabet Med 1997 Aug;14 Suppl 3:512-8).

: Type Il diabetes mellitus is a progressive disease associated with the reduction of pancreatic function and/or other insulin-related processes, aggravated by increased ’ plasma glucose levels. Thus, Type ll diabetes mellitus usually has a prolonged prediabetic phase and various pathophysiological mechanisms can lead to pathological hyperglycemia and impaired glucose tolerance, for instance, abnormalities in glucose utilization and effectiveness, insulin action and/or insulin production in the prediabetic state (Goldberg RB, Med Clin North Am 1998

Jul;82(4):805-21).

The prediabetic state associated with glucose intolerance can also be associated with a predisposition to abdominal obesity, insulin resistance, hyperlipidemia, and high blood pressure, that is, Syndrome X (Groop L, Forsblom C,

Lehtovirta M, Am J Hypertens 1997 Sep;10(9 Pt 2):172S-180S; Haffner SM, J

Diabetes Complications 1997 Mar-Apr;11(2):69-76; Beck-Nielsen H, Henriksen JE,

Alford F, Hother-Nielson O, Diabet Med 1996 Sep;13(9 Supp! 6):578-84).

Thus, defective carbohydrate metabolism is pivotal to the pathogenesis of Type

II diabetes mellitus and impaired glucose tolerance (Dinneen SF, Diabet Med 1997 Aug;14 Suppl 3:519-24). In fact, a continuum from impaired glucose tolerance and impaired fasting glucose to definitive Type Il diabetes mellitus exists (Ramlo-Halsted

BA, Edelman SV, Prim Care 1999 Dec;26(4):771-89).

Early intervention in individuals at risk to develop Type Ii diabetes mellitus, focusing on reducing the pathological hyperglycemia or impaired glucose tolerance may prevent or delay the progression towards Type Il diabetes mellitus and associated complications and/or Syndrome X. Therefore, by effectively treating impaired oral glucose tolerance and / or elevated blood glucose levels, one can prevent or inhibit the progression of the disorder to Type Il diabetes mellitus or Syndrome X. . Many anti-diabetic agents typically prescribed for the treatment of Type Hl diabetes mellitus and/or Syndrome X, for example, sulfonylureas and ‘ thiazolidinediones, have an undesired side effect of increasing body weight. Increased body weight in patients with prediabetic conditions or with diagnosed Type II diabetes mellitus or Syndrome X results in deleterious effects due to accentuation of the metabolic and endocrine dysregulation, and obesity per se is a pivotal risk factor for

L] the development and progressive worsening of Type Il diabetes mellitus. Thus itis desirable to have an anti-diabetic agent which maintains or lowers body weight.

DISCLOSURE OF THE INVENTION

. 5

It has now been found that compounds of the following formula (I):

X 1

CH,0SO,NHR

R

Rr?

R* R® (1) wherein X is O or CH2, and R', R?, R%, R* and R® are as defined hereinafter are useful in preventing the development of Type Il diabetes mellitus and Syndrome X.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The sulfamates of the invention are of the following formula (i):

X 1

RS CH,0OSO,NHR

R* R® 0) wherein

X is CH2 or oxygen;

R1 is hydrogen or alkyl; and

Rz, R3, R4 and Rs are independently hydrogen or lower alkyl and, when Xis

CHa, R4 and Rs may be alkene groups joined to form a benzene ring and, when X is oxygen, Rz and R3 and/or R¢ and Rs together may be a methylenedioxy group of the following formula (11):

O— . "( 3

RY o—3 (I) wherein i WO 02/03984 PCT/US01/21404

Rs and R” are the same or different and are hydrogen, lower alkyl or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring.

R1 in particular is hydrogen or alkyl of about 1 to 4 carbons, such as methyl, ethyl and iso-propyl. Alkyl throughout this specification includes straight and branched } 5 chain alkyl. Alkyl groups for R?, R3, R¢, Rs, R¢ and R7 are of about 1 to 3 carbons and include methyl, ethyl, iso-propyl and n-propyl. When X is CH2, R* and RS may combine to form a benzene ring fused to the 6-membered X-containing ring, i.e., R* and Rs are defined by the alkatrienyl group =C-CH=CH-CH=.

A particular group of compounds of formula (1) is that wherein X is oxygen and both R2 and R3 and R¢ and Rs together are methylenedioxy groups of the formula (II), wherein Re and R7 are both hydrogen both alkyl or combine to form a spiro cyclopentyl or cyclohexyl ring, in particular where Re and R7 are both alkyl such as methyl. A second group of compounds is that wherein X is CH2 and R* and RS are joined to form a benzene ring. A third group of compounds of formula (1) is that wherein both R2 and

Re& are hydrogen.

The compounds of formula (i) may be synthesized by the following methods: (a) Reaction of an alcohol of the formula RCH20H with a chlorosulfamate of the formula CISO2NH2 or CISO2NHR! in the presence of a base such as potassium - butoxide or sodium hydride at a temperature of about -20° to 25° C and in a solvent such as toluene, THF, or dimethylformamide wherein R is a moiety of the following formula (111):

X

RS

0

RY Ra (b) Reaction of an alcohol of the formula RCH20H with sulfurylchloride of ] the formula SO2CI2 in the presence of a base such as triethylamine or pyridine at a . temperature of about -40° to 25° C in a solvent such as diethyl ether or methylene chloride to produce a chlorosulfate of the formula RCH20S02CH.

The chlorosulfate of the formula RCH20S02CI may then be reacted with an amine of the formula R'NH? at a temperature of abut 40° to 25° C in a solvent such as methylene chloride or acetonitrile to produce a compound of formula (1). The reaction conditions for (b) are also described by T. Tsuchiya et al. in Tetrahedron Lelt., 1978, 3365. (c) Reaction of the chlorosuifate RCH20S02CI with a metal azide such as sodium azide in a solvent such as methylene chloride or acetonitrile yields an azidosulfate of the formula RCH20S02N3 as described by M. Hedayatullah in

Tetrahedron Lett. 1975, 2455. The azidosulfate is then reduced to a compound of formula (I) wherein R' is hydrogen by catalytic hydrogenation, e.g. with a noble metal and Ho or by heating with copper metal in a solvent such as methanol.

The starting materials of the formula RCH20H may be obtained commercially or as known in the art. For example, starting materials of the formula RCH20H wherein both R2 and R3 and R¢ and RS are identical and are of the formula (II) may be obtained by the method of R. F. Brady in Carbohydr. Res. 1970, 14, 35 or by reaction of the trimethylsilyl enol ether of a RECOR? ketone or aldehyde with fructose at a temperature of about 25° C, in a solvent such a halocarbon, e.g. methylene chloride in the presence of a protic acid such as hydrochloric acid or a Lewis Acid such as zinc chloride. The trimethylsilyl enol ether reaction is described by G. L. Larson et al. in J.

Org. Chem. 1973, 38, 3935.

Further, carboxylic acids and aldehydes of the formulae RCOOH and RCHO may be reduced to compounds of the formula RCH,OH by standard reduction techniques, e.g. reaction with lithium aluminum hydride, sodium borohydride or borane-THF complex in an inert solvent such a diglyme, THF or toluene at a temperature of about 0° to 100° C, e.g. as described by H.O. House in "Modern

Synthetic Reactions”, 2nd Ed., pages 45 to 144 (1972).

The compounds of formula I: may also be made by the process disclosed US

Patents: No.4,513,006, No.5,242,942, and No.5,384,327, which are incorporated by reference herein.

The compounds of formula (I) include the various individual isomers as well as the racemates thereof, e.g., the various alpha and beta attachments, i.e., below and . 30 above the plane of the drawing, of R2, R3, R* and RS on the 6-membered ring.

Preferably, the oxygen of the methylenedioxy group (ll) are attached on the same side . of the 6-membered ring.

The following Examples are set forth to aid in the understanding of the invention, and are not intended and should not be construed to limit in any way the invention set forth in the claims which follow thereafter.

EXAMPLE 1

It is known that ob/ob mice when allowed to eat ad libitum, develop abnormally high blood levels of insulin (hyperinsulinemia), glucose (hyperglycemia), lesions of the skin, and a high level of glycosylated hemoglobin, all of which are hallmark signs of

Type ll diabetes mellitus (R.R. HENRY, Ann. Intern. Med. 1996, 124, 97-103; G.W.

EDELSON, Clin. Podiatr. Med. Surg. 1998, 15, 41-48; P.R. JOHNSON, M.R.

GREENWOOD, B.A. HORWITZ and J.S. STERN, Annu. Rev. Nutr. 1991, 11, 325- 353). Based on this knowledge of the ob/ob mouse model, two studies were designed to determine the effect of compounds of formula (1) in these mice models.

In the first study, a uniform population of mice was divided into three groups; in } one group of control mice no topiramate was added to the food for the entire120-day study period. In a second group topiramate was added to the food in amounts sufficient to give a daily dose of 20 mg/kg for 84 days, then a daily dose of 180 mg/kg for 36 days. A third group received a daily dose of 60 mg/kg for the entire 120 days.

After completion of the120-day period, the mice were sacrificed and blood was obtained and prepared for subsequent analyses of glucose, insulin and triglycerides in plasma.

A statistical analysis of the results revealed that glucose was significantly lower in both groups of mice treated with topiramate than in the control group (Table 1).

Insulin levels were also lower in both groups of mice treated with topiramate but the difference was statistically significant only in the first group (Table 1).

Table 1. Effect of topiramate on blood glucose, insulin and triglycerides in ob/ob mice : (N) mg/dL + SEM ng/mL + SEM mg/dL + SEM (10) 180: 36 days P=0.02 P=0.0004 P=0.075

(10) P=0.007 P=0.058 P=0.099

P values were calculated using Student's two-tailed t-test. All P values were obtained from a comparison of the topiramate (TPM)-treated groups to the control group. *,

Body weights at the beginning of topiramate administration and at the end of the study (that is following 120 days of treatment with topiramate in the food) were similar in the control and topiramate-treated groups: 48.2 + 1.1 g on day -1 for the control group and 48.4 + 0.9 on day —1 for the TPM 2 group; and 62.7 + 1.4 g on day 119 for the control group and 62.2 + 1.2 on day 119 for the TPM 2 group. Body weight gain for the control and TPM 2 groups from day —1 to day 119 was also similar. Percentage differences in parentheses are versus control values. 0

In the second study, a uniform population of mice was divided into two groups.

In one group of control mice, no topiramate was added to the food for the entire 118- day period. In a second group, topiramate was added to the food in amounts sufficient to give a daily dose of 60 mg/kg for 6 days, then a daily dose of 180 mg/kg for 112 days. During the course of the dosing period the mice were examined two-times a week for skin lesions. When skin lesions were evident, the severity was estimated based on the number and size of the lesions, and given a score ranging from mild to severe (Table 2). For four of the control mice their health deteriorated to the point that they either died or had to be euthanized. Three of these four control mice had lesions (two mice were classified as severe and one mice as mild). In addition to the four control mice that did not survive the 118-day period, other mice developed skin lesions ranging from mild to severe by the end of the study (Table 2). By comparison none of the nine mice treated with topiramate ever developed skin lesions (Table 2). After the 118-day period was completed, all surviving mice were sacrificed and blood was obtained and prepared for subsequent analysis of plasma glucose and insulin, and glycosylated hemoglobin. In this second study, the level of blood glucose was significantly lower in the topiramate-treated mice relative to the control mice (P<0.05, 276 x 49 mg/dL mean + SEM, n = 7 for the control group and 131 + 13 mg/dL mean + ) SEM, n = 9 for the topiramate-treated group at the end of the study; the difference between groups is 52%), whereas insulin levels did not differ between the two groups. . Glycosylated hemoglobin was significantly higher in the surviving control mice than in the topiramate-treated mice (6.09 = 0.8 (n = 7) versus 3.16 + 0.1 (n = 9), mean + SEM,

P<0.01, 48% reduction with topiramate treatment). Also, throughout the 118 days of study 2, the average body weight of the two groups of mice did not differ, that is, on day —1 body weights were 43.6 + 0.8 g (n = 7) for the control group and 42.8 + 1.19 (n = 9) for the topiramate-treated group; and 55.2 + 2.4 g on day 118 for the control group and 55.1 + 1.3 g on day 118 for the topiramate-treated group. The body weight gain was similar in both groups throughout the study. Therefore, the beneficial effects of topiramate on the biological markers of the disease cannot be secondary to the potential beneficial effect that might arise from a loss of body weight. The data suggest that topiramate redirects metabolic and endocrine activities to improve the diabetic syndrome in the ob/ob mice, activities that are independent of topiramate’s effect on body weight.

Table 2. Skin lesions in control and topiramate-treated ob/ob mice

CONTROL TOPIRAMATE (180 mg/kg/day)

Severe Moderate Mild Severe Moderate Mild Day n=7 N=9 1 1 1 3 0 0 0 0 118 1 1 2 4 0 0 0 0 105 n=8 N=9 2 0 3 5 0 97 n=9 N=9 2 1 3 6 0 0 0 0 96 0 1 3 4 0 0 0 0 83 n=10 N=9 0 1 1 2 0 70 n=11 N=9 0 0 2 2 0 0 0 0 61 0 0 2 2 0 0 0 0 53 0 0 0 0 0 0 0 0 41

For each mouse, lesions were scored as: none observed. Mild: one or two small lesions (less than 5 mm in the longest dimension). Moderate: one or multiple lesions (more than 5 mm but less than 8 mm in the longest dimension). Severe: multiple lesions (more than 8 mm in the longest dimension). Day indicates the day during the 118-day period.

Thus when ob/ob mice were given topiramate admixed into food over a period ’ 20 of four months, the blood levels of glucose and insulin, and the level of glycosylated hemoglobin were significantly lower than in control ob/ob mice not given topiramate. ‘ Furthermore, none of the mice that received topiramate at a dose of 180 mg/kg developed lesions of the skin, whereas control ob/ob mice developed lesions. These results demonstrate that topiramate either reduced or prevented the development of all the hallmark signs of Type Il diabetes mellitus in an obese-diabetic animal model,

even when body weight was not affected. The results also suggest that topiramate can reduce cellular resistance to insulin. This is known to be a primary factor in Type

Il diabetes mellitus (R.R. HENRY, Ann. Intern. Med. 1996, 124, 97-103; J.D.

McGARRY, Am. J. Clin. Nutr. 1998, 67, 5003-5048; J.M. OLEFSKY and J.J. NOLAN,

Am. J. Clin. Nutr. 1995, 61, 980S-986S).

Syndrome X, also termed Insulin Resistance Syndrome (IRS), Metabolic

Syndrome, or Metabolic Syndrome X, is a disorder that presents risk factors for the development of Type 2 diabetes mellitus and cardiovascular disease including glucose intolerance, hyperinsulinemia and insulin resistance, and dyslipidemia (eg, high triglycerides). When ob/ob mice were given topiramate admixed into food over a period of four months, the blood levels of glucose, insulin, and triglycerides were significantly lower than in control ob/ob mice not given topiramate. These results demonstrate that topiramate can reduce or prevent pathophysiological signs associated with Syndrome X and thus prevent its development.

EXAMPLE 2

Six to seven week old female C57 BLK S/J-m"/* Lepr® mice (db/db) and heterozygous littermates were purchased from The Jackson Laboratory (Bar Harbor,

ME). Upon arrival the mice were quarantined for 5 days and housed in pairs in shoe- box cages containing ALPHA-dri® bedding (Shepherd Speciality Papers, Inc.,

Kalamazoo, MI). The mice were maintained at an ambient temperature of 21 to 23°C on a 12hour-12hour light-dark schedule and given access to water and food ad libitum.

The diet was comprised of NIH (National Institutes of Health) Rat and Mouse/Auto 6F

Reduced Fat Diet No. 5K52 (PMI Nutrition International Inc., Brentwood, MO).

The vehicle, used as a reference and for test compounds, was 0.5% methylcellulose dissolved in water. The compounds were either fully dissolved or uniformly suspended in the vehicle when administered to the mice.

The db/db mice were randomly separated into five groups of eight, as were the heterozygous littermates. The groups were as follows: one vehicle control group and : four groups that each received one of four doses of topiramate (TPM) (10, 30, 100, or 300 mg/kg, respectively). Topiramate or vehicle was administered orally by gavage ‘ once a day during the 8" hour of the light portion of the light-dark cycle.

Between 18 and 24 hours after the last dose was administered, the mice were anesthetized with CO,/O, (70:30) and blood from the retro-orbital sinus puncture was collected into 2 mL heparinized snap-top polypropylene tubes, then placed inice.

Plasma was separated from blood cells by centrifugation (15 minutes at 1600 g). For samples not assayed immediately, the plasma was transferred into 96-well plates and frozen at -70°C.

Glucose and triglycerides were assayed using standard procedures for blood clinical laboratories. Specifically, the samples were analyzed using an automated

Hitachi 717 autoanalyzer (Boehringer Mannheim/Hitachi 717 Autoanalyzer, Boehringer

Mannheim Laboratory Systems Division, Indianapolis, IN). A statistical analysis of data for the drug-treated groups compared to data for the vehicle group was performed using the one-way analysis of variance (ANOVA) with Dunnett's Multiple

Comparisons test.

Following the procedure described above, the effect of topiramate on plasma glucose and triglyceride levels was determined for female diabetic db/db mice and littermates, following 11 days oral dosage, with results as listed in Table 3 and 4.

Body weights are in Table 5. The abbreviation N represents the number of animals per study group.

Table 3: Plasma Glucose

Homozygous Diabetic Mice Heterozygous Non-Diabetic Mice

Treatment, | N Plasma Conc. % Change | N Plasma Conc. % Change (mg/dL + SEM) (mg/dL + SEM)

Vehicle 8 147 + 4 -

Topiramate | 8 382 +46 27.4 7 145+ 6 1.2 10 mg/kg P<0.05

Topiramate | 7 344 + 31 -34.6 7 152+ 6 +3.8 mg/kg P<0.01

Topiramate | 8 333 +42 -36.6 8 148 +7 +0.8 100 mg/kg P<0.01

Topiramate | 7 207 + 41 -60.5 8 12117 -17.2 300 mg/kg P<0.01 P<0.01

Table 4: Plasma Triglycerides

Homozygous Diabetic Mice Heterozygous Non-Diabetic Mice

Treatment N PlasmaConc. % Change | N Plasma Conc. % Change (mg/dL + SEM) (mg/dL + SEM)

Vehicle 8 141 £10 -

i WO 02/03984 PCT/US01/21404

Topiramate 8 277 +30 -18.9 7 141+ 13 -0.3 10 mg/kg

Topiramate | 7 303 +29 -11.2 7 116 +6 -17.5 : 30 mg/kg

Topiramate | 8 227 +15 -33.7 8 92+6 -34.8 100 mg/kg P<0.01 P<0.01

Topiramate 7 199 +16 -41.7 8 138 £15 -1.8 300 mg/kg P<0.01

Table 5: Body Weights

Heterozygous Non-Diabetic Mice

Treatment N Change in Body N Change in Body

Vehicle 8 0.7+04

Topiramate 8 1.7+£0.9 7 0.5+0.2 10 mg/kg

Topiramate 7 26+1.0 7 00x03 30 mg/kg

Topiramate 8 1.5+£07 8 02+0.2 100 mg/kg

Topiramate 7 07x24 8 -22+0.5 300 mg/kg P<0.05

The results show that topiramate decreased blood glucose in a dose- dependent manner by 27% (P<0.05 vs. control), 35% (P<0.01 vs. control), 37% (P<0.01 vs control), and 61% (P<0.01 vs control) for dosage levels of 10, 30, 100 and 300 mg/kg/day respectively. Topiramate dosing at 100 and 300 mg/kd/day also significantly (P<0.01) decreased plasma triglyceride levels by up to 42% versus diabetic controls. The results demonstrate that topiramate significantly ameliorates the diabetic condition of the homozygous diabetic mice and that this activity of topiramate is not dependent on a reduction in body weight.

EXAMPLE 3

Six week old male Zucker diabetic fatty (ZDF/Gmi-Fa) rats were purchased from Genetic Models, Inc. Indianapolis, Indiana. The rats were housed in groups of four in hanging metal cages, at an ambient temperature of 68-72°F, on a 12 hour-12 hour light-dark schedule and given access to water and food ad libitum. Lean rats (ZDF/GMI -+/+ or +/fa) were used as normal, non-diabetic controls. The diet was comprised of LabDiet 5008 breeding formula (PM! Nutrition Int'l, Brentwood, MO).

The rats were randomly separated into four groups of eight. The groups were as follows: one vehicle control group and three groups that each received one of three doses of TPM (30, 100, or 300 mg/kg, respectively). Topiramate or vehicle was administered orally by gavage once a day, for 14 consecutive days, during the 8" hour of the light portion of the light-dark cycle.

At the start of the study, the rats were bled through a tail vein and plasma glucose and triglyceride levels were determined. Between 18 and 24 hours after the last dose was administered, blood samples were again taken via tail clip on day 1 (fed animals) and day 14 (fed animals). The plasma was collected into 2 mL heparinized snap-top polypropylene tubes, then placed in ice. Plasma was separated from blood cells by centrifugation (20 minutes at 1800 g). For samples not assayed immediately, the plasma was transferred into 96-well plates and frozen at -70°C.

Body weights for the animals were determined at the start of the study and again after 14 days oral dosing. Glucose and triglycerides were assayed using standard procedures for blood clinical laboratories. Specifically, the samples were analyzed using an automated Hitachi 717 autoanalyzer (Boehringer Mannheim/Hitachi 717 Autoanalyzer, Boehringer Mannheim Laboratory Systems Division, Indianapolis,

IN). A statistical analysis of data for the drug-treated groups compared to data for the vehicle group was performed using the one-way analysis of variance (ANOVA) with

Dunnett's Multiple Comparisons test.

Following the procedure described above, the effect of topiramate on plasma glucose, triglyceride levels and body weight changes was determined for male Zucker rats, orally dosed for 14 days, with results as listed in Table 6-8. The abbreviation N : represents the number of animals per study group.

i WO 02/03984 PCT/USO1/21404

Table 6: Plasma Glucose Levels “Treatment N Plasma Conc. % Change from % of Diabetic (mg/dL) + sem Diabetic Control Control “Diabetic Vehicle 8 8296x177

Control 30 mg/kg P<0.01

Topiramate 7 170.3 + 14.7 -47.5 52.5 100 mg/kg P<0.01

Topiramate 8 166.4 + 11.1 -49.5 50.5 300 mg/kg P<0.01

Table 7: Plasma Triglyceride Levels “Treatment _N Plasma Conc. % Change from % of Diabetic (mg/dL) + sem Diabetic Control Control “Diabetic Control 7 6059 +401 “Topiramate 7 4824%105 204 796 30 mg/kg P<0.05

Topiramate 7 498.6 + 58.8 -17.7 82.3 100 mg/kg

Topiramate 8 423.0 + 33.6 -30.2 69.8 300 mg/kg P<0.01

Table 8: Body Weight Change (grams) “Treatment _________N Body WeightChange

Gms + sem “Diabefic Vehicle Control 8 +108.0+21 “Topiramate, 30mglkg 8 +955x72

Topiramate, 100 mg/kg 7 +75.7 + 2.7 (P<0.01)

Topiramate, 300 mg/kg 8 +64.4 + 6.0 (P<0.01)

As indicated by the data in the Tables above, topiramate decreased blood glucose levels by 25-50% (P<0.01). The reduction in blood glucose levels at 30 mg/kg/day occurred without significant body weight change. Topiramate at 30 and

Claims

: PCT/US01/21404

1. Use of a compound of the formula I: : a SE aia

R . . R2 ‘RY R SC wherein ; X is CH, or oxygen; Ris hydrogen or alkyl; and RZ, R3, R* and RS are independently hydrogen or lower alkyl and, when X is CH,, R* and R® may be alkene groups joined to form a benzene ring and, when X is oxygen, R? and R® and/or R* and R® together may be a methylenedioxy group of the following formula (ll): Co 0 | . “ EE RY | a 3 an : wherein RS and R7 are the same or different and are hydrogen, lower alkyl or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring, : in the manufacture of a medicament for treating or preventing the development of Type Il diabetes mellitus in mammals afflicted with such condition, by administering a therapeutically amount of said medicament.

2. Use of claim 1 wherein the compound of formula (I) is topiramate. :

3. Use of claim 1, wherein the therapeutically effective amount is from about 10 to 650 mg. AMENDED SHEET

PCT/US01/21404

4. Use of claim 1, wherein the amount is of from about 16 to 325 mg once or twice daily. .

5. Use of a compound of the formula I: 3 5 TX _CH,0SONHR' R R? : A 0) = wherein . 5 X is CH, or oxygen; R' is hydrogen or alkyl; and R2, R3, R* and R°® are independently hydrogen or lower alkyl and, when X is CH?, R* and R® may be alkene groups joined to form a benzene ring and, when X is oxygen, R? and R® and/or R* and R® together may be a methylenedioxy group of the following formula (ll}:

O . - YY BE | : o—3 (in) wherein ’ R® and R” are the same or different and are hydrogen, lower alkyl or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring, : in the manufacture of a medicament for treating or preventing the development of Syndrome X (Insulin Resistance Syndrome, Metabolic Syndrome, or Metabolic Syndrome X) in mammals afflicted with such condition, by administering a therapeutically effective amount of said AMENDED SHEET

PCT/US01/21404 medicament.

6. Use of claim 5 wherein the compound of formula (I) is topiramate.

7. Use of claim 5, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

8. Use of claim 5, wherein the therapeutically effective amount is from about 10 to 650 mg daily.

9S. Use of claim 5, wherein the amount is of from about 16 to 325 mg once or twice daily.

10. Use of a compound of the formula I: x - RS CH,;OS0O,NHR .R* R3 (") wherein } : X is CH, or oxygen; ) R! is hydrogen or alkyl; and ’ : RZ, R3, R* and R® are independently hydrogen or lower alkyl and, when X is CH2, R* and R® may be alkene groups joined to form a : benzene ring and, when X is oxygen, RZ and R3 and/or R* and R® together may be a methylenedioxy group of the following formula (li): AMENDED SHEET

PCT/US01/21404 RNs O 3 (in ~ wherein - RS and R7 are the same or different and are hydrogen, lower alkyl : or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring, in the manufacture of a medicament for treating impaired oral glucose tolerance in mammals afflicted with such condition, by administering a therapeutically effective amount of said medicament. N © 11. Use of claim 10 wherein the compound of formula (I) is topiramate. : 12. Use of claim 10, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

13. Use of claim 10, wherein the therapeutically effective amount is from about 10 to 650 mg daily. :

14. Use of claim 10, wherein the amount is of from about 16 to 325 mg once or twice daily.

15. Use of a compound of the formula I: AMENDED SHEET 26 J

PCT/US01/21404 I A _CH,0SC,NHR'

R . RZ .R* R® IE) wherein : X is CH, or oxygen; R! is hydrogen or alkyl; and : RZ, R3, R* and R® are independently hydrogen or lower alkyl and, when X is CH?, R* and R® may be alkene groups joined to form a benzene ring and, when X is oxygen, RZ and R® and/or R% and R® together may be a methylenedioxy group of the following formula (li):

o . : } +X | n wherein R® and R’ are the same or different and are hydrogen, lower alkyl or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring, in the manufacture of a medicament for treating or preventing the development of skin lesions associated with Type Il diabetes mellitus or : Syndrome X in mammals afflicted with such condition, by administering a therapeutically effective amount of said medicament.

18. Use of claim 15 wherein the compound of formula (I) is AMENDED SHEET

PCT/US01/21404 topiramate.

17. Use of claim 15, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

18. Use of claim 15, wherein the therapeutically effective amountis from about 10 to 650 mg daily.

19. Use of claim 15, wherein the amount is of from about 16 to 325 mg once or twice daily.

20. Use of a compound of the formula I: : X 1 RS CH,OSC,NHR 2 4 3 R R | (1) wherein X is CH, or oxygen; R! is hydrogen or alkyl; and ~~ RZ, R3 R*and R® are independently hydrogen or lower alkyl and, : when X is CHZ, R* and R® may be alkene groups joined to form a benzene ring and, when X is oxygen, RZ and R® and/or R* and R® together may be a methylenedioxy group of the following formula (ll): AMENDED SHEET

PCT/US01/21404 . i ®) . : } . . tS ~3 | : R" _ : O 3 ti) : wherein R6 and R’ are the same or different and are hydrogen, lower alkyl or are alkyl and are joined to form a cyclopentyl or cyclohexyl ring, in the manufacture of a medicament for improving defective insulin sensitivity in mammals afflicted with such condition, by administering a therapeutically effective amount of said medicament.

21. Use of claim 20 wherein the compound of formula (I) is ‘topiramate.

22. Use of claim 20, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

23. Use of claim 20, wherein the therapeutically effective amount is from about 10 to 650 mg daily.

24. Use of claim 20, wherein the amount is of from about 16 to 325 mg once or twice daily.

25. A substance or composition for use in a method for treating or preventing the development of Type Il diabetes mellitus in mammals afflicted with such condition, said substance or composition comprising a compound of formula |, as defined in claim 1, and said method AMENDED SHEET

PCT/US01/21404 comprising administering a therapeutically effective amount of said substance or composition.

26. A substance or composition for use in a method of treatment of claim 25, wherein the compound of formula (I) is topiramate.

27. A substance or composition for use in a method of treatment of claim 25, wherein the therapeutically effective amount is from about 10 to 650 mg.

28. A substance or composition for use in a method of treatment of claim 25, wherein the amount is of from about 16 to 325 mg once or twice daily.

29. A substance or composition for use in a method for treating or _preventing the development of Syndrome X (Insulin Resistance Syndrome, Metabolic Syndrome, or Metabolic Syndrome X) in mammals afflicted with such condition, said substance or composition comprising a compound of the formula | as defined in claim 5, and said method comprising administering a therapeutically effective amount of said substance or composition.

30. A substance or composition for use in a method of treatment of claim 29, wherein the compound of formula (I) is topiramate. :

31. A substance or composition for use in a method of treatment of claim 29, wherein the therapeutically effective amount is from about 10 to 1000 mg daily. AMENDED SHEET

PCT/US01/21404 : 32. A substance or composition for use in a method of treatment of claim 29, wherein the therapeutically effective amount is from about 10 to 650 mg.

33. A substance or composition for use in a method of treatment of claim 29, wherein the amount is of from about 16 to 325 mg once or twice daily.

34. A substance or composition for use in a method for treating impaired oral glucose tolerance in mammais afflicted with such condition, said substance or composition comprising a compound of the formula | as defined in claim 10, and said method comprising administering a therapeutically effective amount of said substance or composition.

35. A substance or composition for use in a method of treatment of claim 34, wherein the compound of formula (I) is topiramate.

36. A substance or composition for use in a method of treatment of : claim 34, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

37. A substance or composition for use in 2a method of treatment of claim 34, wherein the therapeutically effective amount is from about 10 : to 650 mg.

38. A substance or composition for use in a method of treatment of claim 34, wherein the amount is of from about 16 to 325 mg once or AMENDED SHEET

PCT/US01/21404 twice daily.

39. A substance or composition for use in a method for treating or preventing the development of skin lesions associated with Type ll diabetes mellitus or Syndrome X in mammals afflicted with such condition, said substance or composition comprising a compound of the formula | as defined in claim 15, and said method comprising administering a therapeutically effective amount of said substance or composition.

40. A substance or composition for use in a method of treatment of claim 39, wherein the compound of formula (I) is topiramate.

41. A substance or composition for use in a method of treatment of claim 39, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

42. A substance or composition for use in a method of treatment of claim 38, wherein the therapeutically effective amount is from about 10 to 650 mg.

43. A substance or composition for use in a method of treatment of claim 39, wherein the amount is of from about 16 to 325 mg once or twice daily. :

44. A substance or composition for use in a method for improving defective insulin sensitivity in mammals afflicted with such condition, said substance or composition comprising a compound of the formula AMENDED SHEET

PCT/US01/21404 therapeutically effective amount of said substance or composition.

45. A substance or composition for use in a method of treatment of claim 44, wherein the compound of formula {!) is topiramate.

46. A substance or composition for use in a method of treatment of claim 44, wherein the therapeutically effective amount is from about 10 to 1000 mg daily.

47. A substance or composition for use in a method of treatment of claim 44, wherein the therapeutically effective amount is from about 10 to 650 mg.

48. A substance or composition for use in a method of treatment of claim 44, wherein the amount is of from about 16 to 325 mg once or twice daily.

49. Use according to any one of claims 1 to 24, substantially as herein described and illustrated.

50. A substance or composition for use in a method of treatment or prevention according to any one of claims 25 to 48, substantially as

. herein described and illustrated.

51. A new use of a compound as defined in any one of claims 1 to 24, or a substance or compaosition for a new use in a method of treatment or prevention, substantially as herein described. AMENDED SHEET