TREATMENT OF METABOLIC DISORDERS IN FELINE ANIMALS

Abstract

One or more SGLT2 inhibitors or pharmaceutically acceptable forms thereof are provided for use in the treatment and/or prevention of a metabolic disorder in a feline animal, preferably where the metabolic disorder is one or more selected from the group consisting of ketoacidosis, pre-diabetes, diabetes mellitus type 1 or type 2, insulin resistance, obesity, hyperglycemia, impaired glucose tolerance, hyperinsulinemia, dyslipidemia, dysadipokinemia, subclinical inflammation, systemic inflammation, low grade systemic inflammation, hepatic lipidosis, atherosclerosis, inflammation of the pancreas, neuropathy and/or Syndrome X (metabolic syndrome) and/or loss of pancreatic beta cell function and/or where the remission of the metabolic disorder, preferably diabetic remission, is achieved and/or maintained.

Claims

1. A method of treatment or prevention of a metabolic disorder in a feline animal comprising administering to the feline animal one or more SGLT2 inhibitors or pharmaceutically acceptable crystalline forms thereof, wherein: the metabolic disorder is one or more selected from the group consisting of ketoacidosis, pre-diabetes, diabetes mellitus type 1 or type 2, insulin resistance, obesity, hyperglycemia, impaired glucose tolerance, hyperinsulinemia, dyslipidemia, dysadipokinemia, subclinical inflammation, systemic inflammation, low grade systemic inflammation, hepatic lipidosis, atherosclerosis, inflammation of the pancreas, neuropathy and/or Syndrome X (metabolic syndrome) and/or loss of pancreatic beta cell function and/or the remission of the metabolic disorder is achieved and/or maintained; and the one or more SGLT-2 inhibitors is administered at a dose of 0.01 to 5.0 mg/kg body mass per day.

2. The method of claim 1, wherein the one or more SGLT-2 inhibitors is selected from the group consisting of the following compounds or pharmaceutically acceptable forms thereof: a glucopyranosyl-substituted benzene derivative of the formula (1) ##STR00029## wherein R.sup.1 denotes cyano, Cl or methyl; R.sup.2 denotes H, methyl, methoxy or hydroxyl; and R.sup.3 denotes cyclopropyl, hydrogen, fluorine, chlorine, bromine, iodine, methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, iso-butyl, tert-butyl, 3-methyl-but-1-yl, cyclobutyl, cyclopentyl, cyclohexyl, 1-hydroxy-cyclopropyl, 1-hydroxy-cyclobutyl, 1-hydroxy-cyclopentyl, 1-hydroxy-cyclohexyl, ethinyl, ethoxy, difluoromethyl, trifluoromethyl, pentafluoroethyl, 2-hydroxyl-ethyl, hydroxymethyl, 3-hydroxy-propyl, 2-hydroxy-2-methyl-prop-1-yl, 3-hydroxy methyl-but-1-yl, 1-hydroxy-1-methyl-ethyl, 2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl, 2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl, 2-methoxy-ethyl, 2-ethoxy-ethyl, hydroxy, difluoromethyloxy, trifluoromethyloxy, 2-methyloxy-ethyloxy, methylsulfanyl, methyl sulfinyl, methlysulfonyl, ethyl sulfinyl, ethyl sulfonyl, trim ethyl silyl, (R)-tetrahydrofuran-3-yloxy or (S)-tetrahydrofuran-3-yloxy or cyano, or a derivative thereof wherein one or more hydroxyl groups of the β-D-glucopyranosyl group are acylated with groups selected from (C.sub.1-18-alkyl)carbonyl, (C.sub.1-18-alkyl)oxycarbonyl, phenylcarbonyl and phenyl-(C.sub.1-3-alkyl)-carbonyl; Dapagliflozin, represented by formula (3): ##STR00030## Canagliflozin, represented by formula (4): ##STR00031## Empagliflozin, represented by formula (5): ##STR00032## Luseogliflozin, represented by formula (6): ##STR00033## Tofogliflozin, represented by formula (7): ##STR00034## Ipragliflozin, represented by formula (8): ##STR00035## Ertugliflozin, represented by formula (9): ##STR00036## Atigliflozin, represented by formula (10): ##STR00037## Remogliflozin, represented by formula (11): ##STR00038## a thiophene derivative of the formula (12) ##STR00039## wherein R denotes methoxy or trifluoromethoxy; 1-(β-D-glucopyranosyl)-4-methyl-3-[5-(4-fluorophenyl)-2-thienylmethyl]benzene; represented by formula (13); ##STR00040## a spiroketal derivative of the formula (14): ##STR00041## wherein R denotes methoxy, trifluoromethoxy, ethoxy, ethyl, isopropyl or tert. butyl; a pyrazole-O-glucoside derivative of the formula (15) ##STR00042## wherein R.sup.1 denotes C.sub.1-3-alkoxy, L.sup.1, L.sup.2 independently of each other denote H or F, R.sup.6 denotes H, (C.sub.1-3-alkyl)carbonyl, (C.sub.1-6-alkyl)oxycarbonyl, phenyloxycarbonyl, benzyloxycarbonyl or benzylcarbonyl; a compound of the formula (16): ##STR00043## Sergliflozin, represented by formula (17): ##STR00044## a compound represented by formula (18): ##STR00045## wherein R.sup.3 is selected from cyclopropyl, ethyl, ethinyl, ethoxy, (R)-tetrahydrofuran-3-yloxy or (S)-tetrahydrofuran-3-yloxy.

3. The method of claim 2, wherein the one or more SGLT-2 inhibitors is the glucopyranosyl-substituted benzene derivative of the formula (1), and R.sup.3 is selected from cyclopropyl, ethyl, ethinyl, ethoxy, (R)-tetrahydrofuran-3-yloxy or (S)-tetrahydrofuran-3-yloxy.

4. The method of claim 1, wherein the one or more SGLT-2 inhibitors is selected from the group consisting of the following compounds or pharmaceutically acceptable forms thereof: a glucopyranosyl-substituted benzene derivative of the formula (1) ##STR00046## wherein R.sup.1 denotes cyano, Cl or methyl; R.sup.2 denotes H, methyl, methoxy or hydroxy; and R.sup.3 denotes cyclopropyl, hydrogen, fluorine, chlorine, bromine, iodine, methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, iso-butyl, tert-butyl, 3-methyl-but-1-yl, cyclobutyl, cyclopentyl, cyclohexyl, 1-hydroxy-cyclopropyl, 1-hydroxy-cyclobutyl, 1-hydroxy-cyclopentyl, 1-hydroxy-cyclohexyl, ethinyl, ethoxy, difluoromethyl, trifluoromethyl, pentafluoroethyl, 2-hydroxyl-ethyl, hydroxymethyl, 3-hydroxy-propyl, 2-hydroxy-2-methyl-prop-1-yl, 3-hydroxy-3-methyl-but-1-yl, 1-hydroxy-1-methyl-ethyl, 2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl, 2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl, 2-methoxy-ethyl, 2-ethoxy-ethyl, hydroxy, difluoromethyloxy, trifluoromethyloxy, 2-methyloxy-ethyloxy, methyl sulfanyl, methyl sulfinyl, methlysulfonyl, ethylsulfinyl, ethyl sulfonyl, trimethylsilyl, (R)-tetrahydrofuran-3-yloxy or (S)-tetrahydrofuran-3-yloxy or cyano, or a derivative thereof wherein one or more hydroxyl groups of the β-D-glucopyranosyl group are acylated with groups selected from (C.sub.1-18-alkyl)carbonyl, (C.sub.1-18-alkyl)oxycarbonyl, phenylcarbonyl and phenyl-(C.sub.1-3-alkyl)-carbonyl; Dapagliflozin, represented by formula (3): ##STR00047## Canagliflozin, represented by formula (4): ##STR00048## Empagliflozin, represented by formula (5): ##STR00049## Ertugliflozin, represented by formula (9): ##STR00050## a compound represented by formula (18): ##STR00051## wherein: R.sup.3 is selected from cyclopropyl, ethyl, ethinyl, ethoxy, (R)-tetrahydrofuran-3-yloxy or (S)-tetrahydrofuran-3-yloxy.

5. The method of claim 1, wherein: the metabolic disorder is diabetes, pre-diabetes, diabetes mellitus type 1, diabetes mellitus type 2 and/or one or more clinical conditions associated with diabetes; and the one or more clinical conditions is one or more conditions selected from the group consisting of ketoacidosis, insulin resistance, obesity, hyperglycemia, impaired glucose tolerance, hyperinsulinemia, dyslipidemia, dysadipokinemia, subclinical inflammation, systemic inflammation, low grade systemic inflammation, hepatic lipidosis, atherosclerosis, inflammation of the pancreas, neuropathy, Syndrome X (metabolic syndrome), loss of pancreatic beta cell function and/or diabetic remission.

6. The method of claim 1, wherein the metabolic disorder is pre-diabetes, diabetes mellitus type 1 or diabetes mellitus type 2.

7. The method of claim 1, wherein the metabolic disorder is pre-diabetes or diabetes, and the method effects an elimination of one or more owner-observed signs.

8. The method of claim 7, wherein the owner-observed signs are selected from the group consisting of lethargy, polyuria, polydipsia, weight loss, and polyphagia, that occur secondary to hyperglycemia of untreated animals.

9. The method of claim 1, wherein the feline animal is obese.

10. The method of claim 1, wherein the feline animal is suffering from diabetes.

11. The method of claim 1, wherein the feline animal is suffering from pre-diabetes or diabetes mellitus type 2.

12. The method of claim 1, wherein the feline animal is a cat.

13. The method of claim 1, wherein the pharmaceutically acceptable crystalline form thereof is a crystalline complex between the one or more SGLT2 inhibitors or pharmaceutically acceptable forms thereof and one or more amino acids.

14. The method of claim 13, wherein the pharmaceutically acceptable crystalline form thereof is proline or L-proline.

15. The method of claim 1, wherein the one or more SGLT-2 inhibitors is administered orally or parenterally.

16. The method of claim 1, wherein the one or more SGLT-2 inhibitors is administered as a liquid.

17. The method of claim 1, wherein the one or more SGLT-2 inhibitors is administered as a tablet.

18. The method of claim 1, wherein the one or more SGLT-2 inhibitors is administered only once per day.

19. The method of claim 1, wherein the one or more SGLT-2 inhibitors is administered in combination with insulin.

20. The method of claim 1, wherein the metabolic disorder is diabetes, and the method effects an improvement of glycemic control in otherwise healthy cats with diabetes mellitus not previously treated with insulin.

Description

BRIEF DESCRIPTION OF THE FIGURES

[0230] FIG. 1 shows the correlation between compound A plasma level and urinary glucose excretion normalized to creatinine (gluc/crea). There is a clear logarithmic-linear relationship.

[0231] FIGS. 2A and 2B show the blood glucose and insulin secretion profiles in an intravenous glucose tolerance test (ivGTT) of normal lean cats according to Hoenig (Mol Cell Endocrinal 2002, 197(1-2): 221-229) (iv GTT [1 g/kg]) and of insulin resistant obese cats before (dotted line—pretests, “pre”) and after 4 weeks of treatment with compound A (solid line) (FIG. 2B). The increased and prolonged second phase of the insulin resistant obese cats used in the present study was significantly improved by treatment with compound A.

[0232] FIGS. 3A and 3B show area-under-curve (AUC) values of blood insulin and a surrogate insulin sensitivity index (blood insulin-to-glucose relationship as expressed by the modified Belfiore index) in insulin resistant cats during an intravenous glucose tolerance test (ivGTT) before (“pre”) and after (“post”) 4 weeks of treatment with compound A or its vehicle (“control”). Treatment with compound A leads to a significant reduction of Insulin AUC (panel A), and significantly improved insulin sensitivity FIG. 3B).

[0233] FIGS. 4A and 4B show time courses of blood glucose concentrations [mmol/L] after insulin challenge in insulin resistant cats during an intravenous insulin tolerance test (ivITT) before (dotted line—pretests, “pre”) and after 4 weeks of treatment (solid line) with compound A or its vehicle (“control”). In untreated animals (controls) insulin sensitivity (IS) decreased throughout the study (FIG. 4A). In comparison, treatment with compound A was associated with a significant improvement in IS (FIG. 4B).

[0234] FIGS. 5A and 5B show time courses of non-esterified fatty acid (NEFA) levels in blood [mEq/L] after insulin challenge in insulin resistant cats during an ivITT before (dotted line—pretests, “pre”) and after 4 weeks of treatment (solid line) with compound A or its vehicle (“control”). In untreated animals (controls) NEFA elimination significantly worsened throughout the study period (FIG. 5A), whereas it was significantly improved by treatment with compound A (FIG. 5B).

[0235] FIG. 6. shows that blood leptin concentrations significantly decreased over the study period in the treated cats.

[0236] FIG. 7 shows a reduction of the respiratory exchange ratio (RER) (indicating increased lipid utilization) in treated animals, as measured by indirect calorimetry.

[0237] FIG. 8 shows that β-hydroxybutyrate levels in blood (β-HB/BHB) increased following 4 weeks of treatment with compound A.

[0238] FIG. 9 shows the positive correlation between the change of blood leptin concentration and the change of RER before and after 4 weeks of treatment with compound A or vehicle (control).

[0239] FIG. 10 shows the negative correlation between β-hydroxybutyrate levels in blood (β-HB/BHB) and the RER after 4 weeks of treatment with compound A.

[0240] FIG. 11 shows an X-ray powder diffraction pattern of a representative batch of a crystalline complex of compound A with L-proline (1:1).

[0241] FIG. 12 shows a DSC/TG diagram of a representative batch of a crystalline complex of compound A with L-proline (1:1).

[0242] FIG. 13 shows mean blood glucose from the 9 hour glucose curve by visit day.

[0243] FIG. 14 shows the serum fructosamine by visit day.

[0244] FIG. 15 shows preliminary data from four cats demonstrate that fasting insulin concentrations increased compared to a simultaneous decrease of the mean glucose values (from a 9 hour blood glucose curve) at Day 7 compared to Day −1. Afterwards insulin concentrations reached a plateau which can be explained by already nearly normalized glucose concentration. This reflects the normal physiological situation in fasted animals: when glucose is within the normal range (fasted state) no increase of insulin concentrations are expected to be present any more. This preliminary data from the fasting insulin values from four cats support the claimed indications “loss of pancreatic beta cell function” and “remission of the metabolic disorder, preferably diabetic remission” since it demonstrates the increase in insulin concentrations and decrease in glucose concentrations back to a normalized values and therefore reflects the return to a normal physiological response.

EXAMPLES

[0245] The following examples show the beneficial therapeutic effects on glycemic control and/or insulin resistance, etc., of using one or more SGLT2 inhibitors in feline animals, according to the present invention. These examples are intended to illustrate the invention in more detail without any limitation of the scope of the claims.

Example 1 Pharmacokinetics (PK)/Pharmacodynamics (PD) of Compound a Single Oral Dosing in Cats

[0246] Compound A was administered to overnight fasted cats. The groups (n=3 per group) received a single oral administration of either vehicle alone (water) or vehicle containing the SGLT2 inhibitor Compound A at a dose of 0.01 mg/kg, 0.1 mg/kg and 1 mg/kg. PK/PD measurements were taken until day 4 after a single administration of compound A or its vehicle.

TABLE-US-00002 TABLE 2 Pharmacokinetic data, single dose (0.01/0.1/1.0 mg/kg) 0.01 0.1 1.0 Parameter mg/kg mg/kg mg/kg .sub.tmax [hour] mean 1 1.3 1 C.sub.max [nmol/L] mean 9 77 1173 AUC.sub.0.fwdarw.∞ [nmol .Math. h/l] mean 30 358 5379 T.sub.1/2 [hour] mean 1.2 2.9 5.4

[0247] Pharmacodynamic Data: [0248] A prominent increase of urinary glucose concentration was evident at doses>0.01 mg/kg already 8 h after administration (mean group values: controls 1.4 mmol/L; 0.01 mg/kg-1.4 mmol/L; 0.1 mg/kg-46.1 mmol/L; 1 mg/kg-239.3 mmol/L) and was persistent for more than 24 h. [0249] None of the three doses of compound A altered the blood glucose level in cats as compared to normal reference values. [0250] None of the three doses of compound A altered the renal function of cats.

[0251] Urinary glucose excretion increase is clearly dose and plasma compound exposure dependent (logarithmic-linear correlation), as shown in Error! Reference source not found.

Example 2 the Effect of Compound a on Urinary and Blood Glucose after Repeated Dosing in Cats

[0252] Compound A was administered to overnight fasted cats. The groups (n=3 per group) received a once daily oral administration of either vehicle alone (PillPocket®) or vehicle containing the SGLT2 inhibitor (dry compound) at a dose of 1 mg/kg and 3 mg/kg for 3 consecutive days. Urinary glucose and blood glucose were measured. [0253] A prominent increase of urinary glucose concentration was evident at both doses already 8 h after administration. The maximal urinary concentration was not further elevated after repeated dosing and was similar at doses of 1 mg/kg and 3 mg/kg (mean values—281 mmol/L and 209 mmol/L, respectively). [0254] Neither dose of compound A altered the blood glucose level in cats as compared to normal reference values.

[0255] In respect to urinary glucose excretion it is thus estimated that the ED.sub.50 is <1 mg/kg.

Example 3 the Effect of Compound a on Urinary and Blood Glucose after Repeated Dosing in Cats

[0256] Compound A was administered to freely fed normoglycemic, obese cats. The groups (n=6 per group) received a once daily oral administration of either vehicle alone (gelatin capsules) or vehicle containing the SGLT2 inhibitor (dry compound) at a dose of 1 mg/kg for 4 weeks. Urinary glucose and blood glucose were measured. [0257] Urinary glucose concentrations were significantly elevated at the end of the study—controls 0.6 mmol/L; 1 mg/kg-489 mmol/L. [0258] No alterations of blood glucose levels were observed.

Example 4 Treatment of Pre-Diabetes: Prevention of Manifest Type 2 Diabetes in Cats

[0259] The efficacy of SGLT2 inhibition in accordance with the invention in the treatment of pre-diabetes characterized by pathological fasting glucose and/or impaired glucose tolerance and/or insulin resistance can be tested using clinical studies. In studies over a shorter or longer period (e.g. 2-4 weeks or 1-2 years) the success of the treatment is examined by determining the fasting glucose values and/or the glucose values after a meal or after a loading test (oral glucose tolerance test or food tolerance test after a defined meal) after the end of the period of therapy for the study and comparing them with the values before the start of the study and/or with those of a placebo group. In addition, the fructosamine value can be determined before and after therapy and compared with the initial value and/or the placebo value. A significant drop in the fasting or non-fasting glucose and/or fructosamine levels demonstrates the efficacy of the treatment of pre-diabetes. Additionally a significant reduction in the number of patients who develop manifest type 2 diabetes when treated with a pharmaceutical composition according to this invention as compared to another form of treatment, demonstrates the efficacy in preventing a transition from pre-diabetes to manifest diabetes.

Example 5 Treatment of Pre-Diabetes: Improvement of Insulin Resistance in Cats

[0260] The following example shows the beneficial effect of compound A in insulin resistant obese cats. Compound A was administered to freely fed normoglycemic, insulin resistant, obese cats. The groups (n=6 per group) received a once daily oral administration of either vehicle alone (gelatin capsules) or vehicle containing the SGLT2 inhibitor (dry compound) at a dose of 1 mg/kg for 4 weeks. The following experiments were performed prior to treatment and at the end of the 4 week treatment period approximately 24 h after the last administration of compound/vehicle.

[0261] An intravenous glucose tolerance test (ivGTT, 0.8 g/kg dextrose) was performed in overnight fasted cats. Blood was collected via jugular vein catheters. Blood samples were taken at −5, 0, 5, 10, 15, 30, 45, 60, 90, 120, 180 min relative to glucose application.

[0262] Glucose and insulin excursion were quantified by calculating the baseline corrected glucose AUC. An intravenous insulin tolerance test (ivITT, 0.05 U/kg regular insulin) was performed in overnight fasted cats. Blood was collected via jugular vein catheters. Blood samples were taken at −5, 0, 15, 30, 60, 90, 120, 180 min relative to insulin application.

[0263] The excursion of glucose and non-esterified fatty acids (NEFA) was quantified by calculating baseline corrected glucose and NEFA AUC.

[0264] The significance of differences of means between groups is evaluated by repeated-measures two-factor (time & treatment) ANOVA and post hoc multiple comparisons versus control or the respective baseline readings.

[0265] The glucose excursion during the ivGTT did not change during the study period or due to the treatment. The insulin excursion was not altered throughout the study period in control cats, but was significantly reduced in treated cats as compared to baseline values (p<0.05).

[0266] As shown in Error! Reference source not found, as compared to lean cats, in the obese cats used in the present study, the insulin secretion profile exhibited a reduced first phase, and an increased and prolonged second phase. As shown in panel B of Error! Reference source not found, treatment with compound A led to a significant improvement of second phase insulin secretion profile.

[0267] Insulin sensitivity was significantly increased in treated cats as compared to baseline values (p<0.05). This was demonstrated by calculating the relationship between glucose and insulin in terms of the modified Belfiore Index (1/log(ΔAUCgluc*ΔAUCins).

[0268] Area-under-curve values of blood insulin and the blood insulin-to-glucose relationship as represented by the modified Belfiore index for insulin sensitivity in insulin resistant cats during an i.v. glucose tolerance test (ivGTT) before (“pre”) and after (“post”) 4 weeks of treatment with compound A or its vehicle (“control”) are shown in Error! Reference source not found.

[0269] The glucose excursion during the ivITT significantly worsened throughout the study period in the control animals (p<0.05) (see Error! Reference source not found, panel A). This was similar for the elimination of NEFAs (see Error! Reference source not found, panel A). In contrast, in cats treated with compound A the glucose curve did not change throughout the study period (see Error! Reference source not found, panel B), and NEFA elimination was significantly improved by the compound A treatment (p<0.01; see Error! Reference source not found, panel B).

[0270] These data indicate that in obese cats insulin resistance is significantly improved after a 4 week treatment with compound A. As insulin resistance is a characteristic feature of pre-diabetes the data strongly indicate that compound A is capable of treating pre-diabetes in feline animals.

[0271] In clinical studies in diabetic cats running for different lengths of time (e.g. 2 weeks to 12 months) the success of the improvement in insulin resistance can be checked by the measuring baseline blood glucose, blood fructosamine and blood insulin levels and then monitoring the development of those levels in individual cats throughout the study period. Also the glucose and insulin values after a meal or after a loading test (glucose tolerance test or insulin tolerance test) after the end of the period of therapy for the study can be compared with the values before the start of the study and/or with those of diabetic cats who have been treated with other medications.

Example 6 Treatment of Type 2 Diabetes in Cats

[0272] Treating cats with type 2 diabetes with the pharmaceutical composition according to the invention, in addition to producing an acute improvement in the glucose metabolic situation, prevents deterioration in the metabolic situation in the long term. This can be observed if cats are treated for a shorter or longer period, e.g. 2-4 weeks or 3 months to 1 year, with the pharmaceutical composition according to the invention and are compared to the metabolic situation prior to treatment or with cats that have been treated with insulin or other antidiabetic medication. There is evidence of therapeutic success if daily mean blood glucose and fructosamine level are reduced as compared to pretreatment level. Further evidence of therapeutic success is obtained if a significantly smaller percentage of the cats treated with a pharmaceutical composition according to the invention, compared with cats who have been treated with other medications, undergo transient deterioration in the glucose metabolic position (e.g. hyper- or hypoglycemia).

Example 7 Improvement of Pancreatic Beta Cell Function

[0273] In clinical studies in diabetic cats running for different lengths of time (e.g. 4 weeks to 12 months) the success of the treatment is checked using the measurement of baseline blood glucose, blood fructosamine and blood insulin level and the corresponding relation between the parameter in the individual cat. Additionally, e.g. arginine stimulation may be employed to test the pancreatic beta cell ability to secrete insulin.

[0274] A significant rise in the blood insulin level (either baseline or after arginine stimulation) during or at the end of the study, compared with the initial value or compared with a placebo group, or a group given a different therapy, proves the efficacy of a pharmaceutical composition according to the invention in the improvement of pancreatic beta cell function in diabetic cats (FIG. 15).

Example 8 Diabetic Remission

[0275] In clinical studies in diabetic cats running for a longer period (e.g. 3 months to 1 year) the success of the treatment is checked using the measurement of baseline blood glucose, blood fructosamine and blood insulin level and the corresponding relation between the parameter in the individual cat. There is evidence of therapeutic success if laboratory values are reduced as compared to pre-treatment level without the need of insulin injections (FIG. 15).

[0276] In case compound A was employed in a combination with e.g. insulin or other drugs effectively reducing hyperglycemia the feline animal may be weaned off insulin or the other drug and still have a glycemic control in normal ranges.

[0277] Most advantageously, the feline animal may be weaned of compound A.

Example 9 Reduction of Hyperglycemia

[0278] In clinical studies in diabetic cats running for different lengths of time (e.g. 1 day to 12 months) the success of the treatment in cats with hyperglycemia is checked by determining the blood glucose or blood fructosamine level. A significant fall in these values during or at the end of the study, compared with the initial value or compared with a placebo group, or a group given a different therapy, proves the efficacy of a pharmaceutical composition according to the invention in the reduction of hyperglycemia in cats.

Example 10 Body Composition and Body Fat Reduction

[0279] The following example shows the beneficial effect of compound A in obese cats. Compound A was administered to freely fed obese cats. The groups (n=6 per group) received a once daily oral administration of either vehicle alone (gelatin capsules) or vehicle containing the SGLT2 inhibitor (dry compound) at a dose of 1 mg/kg for 4 weeks. The following experiments were performed prior to treatment and at the end of the 4 week treatment period approximately 24 h after the last administration of compound/vehicle. As shown in Error! Reference source not found, blood leptin concentrations significantly decreased over the study period in the treated cats (mean values: pre: 2482 pmol/L, post: 2213 pmol/L, p<0.05).

[0280] Indirect calorimetry shows the influence of the treatment on energy metabolism. Respiratory exchange ratios (RER; ratio between the amount of CO.sub.2 exhaled and 02 inhaled; see Error! Reference source not found.) indicated significantly increased fatty acid metabolism (lipid utilization) in treated animals (mean RER values: 0.749 pre-treatment, 0.728 post-treatment; p<0.01).

[0281] Increased lipid utilization was also mirrored in increased blood β-hydroxybutyrate concentrations (β-HB/BHB), as shown in Error! Reference source not found. The increase of blood β-hydroxybutyrate concentrations did not exceed normal reference values.

[0282] These changes in the relevant data throughout the study show a significant correlation and indicate that treatment shows a beneficial effect on body composition.

[0283] Thus, the data show a positive correlation between the change of blood leptin concentration and the change of RER before and after 4 weeks of treatment with compound A (Error! Reference source not found), and a negative correlation between the blood β-hydroxybutyrate levels (β-HB/BHB) and the RER (Error! Reference source not found).

[0284] Liver parameters were unchanged, and no ketones were detected in the urine. Thus, shifting of the metabolism of lipids and carbohydrates was within normal physiological ranges.

[0285] In consequence, a 4 week treatment in obese cats clearly shows that dysadipokinemia was improved and additionally shifting metabolic substrate utilization from glucose to lipid represents a clear benefit in the treatment of obese cats. The data strongly indicate that Compound A is capable of treating pre-diabetes in feline animals

Example 11 Pilot Trial of Compound a in Client-Owned Diabetic Cats

[0286] The following data are from 4 diabetic cats which had been prospectively treated orally with 1 mg/kg once daily Compound A for 28 days. Diagnosis of diabetes mellitus had been made on the basis of blood glucose>250 mg/dl (13.9 mmol/L) at screening, either glucosuria or serum fructosamine>400 μmon, and the persistence of at least one clinical condition/sign consistent with diabetes mellitus [lethargy, polyuria, polydipsia, polyphagia, weight loss, and/or plantigrade posture of hind legs (DM polyneuropathy)].

[0287] Results revealed that the mean (FIG. 13) blood glucose values of the 9 hour blood glucose curve were substantially decreased in all 4 cats compared to baseline by the end of the study. The decrease was already present at day 7 and unexpectedly to such an extent comparable to long-term insulin therapy. For comparison, comparable reduction in mean blood glucose was not observed in 14 cats treated with VETSULIN® (insulin, Intervet, Inc.) until day 14 (NADA 141-236, Freedom of Information Summary, VETSULIN®). Serum fructosamine confirmed this good glycemic control and was also decreased to below 350 μmon (excellent control according to laboratory interpretive guidelines) in all cats by day 28 (FIG. 14). In contrast, the mean serum fructosamine for cats treated with VETSULIN® was 546 by day 30, and remained elevated at 462 on day 60 (NADA 141-236, Freedom of Information Summary, VETSULIN®).

[0288] All cats showed improvement in at least one clinical condition/sign, and 3 of 4 cats showed improvement in at least 3 clinical conditions/signs as assessed by the owner. All cats improved in overall diabetes control as assessed by the Investigator. Urinary glucose excretion was decreased in all cats by the end of the study. No hypoglycemia (defined as blood glucose less than 70 mg/dL) was reported.

[0289] In conclusion, these data demonstrate that Compound A represents can be used to treat diabetic cats with a once daily oral therapy comparable to long-term twice daily insulin therapy.

Example 12 Preparation of 1-cyano-2-(4-cyclopropyl-benzyl)-4-(β-D-glucopyranos-1-yl)-benzene (Compound A)

[0290] The following example of synthesis serves to illustrate a method of preparing 1-cyano-2-(4-cyclopropyl-benzyl)-4-(β-D-glucopyranos-1-yl)-benzene (compound A). A method of preparing its crystalline complex with L-proline is also described. It is to be regarded only as a possible method described by way of example, without restriction of the scope of the invention. The terms “room temperature” and “ambient temperature” are used interchangeably and denote temperatures of about 20° C. The following abbreviations are used: [0291] DMF dimethylformamide [0292] NMP N-methyl-2-pyrrolidone [0293] THF tetrahydrofuran

Preparation of 4-bromo-3-hydroxymethyl-1-iodo-benzene

[0294] ##STR00020##

[0295] Oxalyl chloride (13.0 mL) is added to an ice-cold solution of 2-bromo-5-iodo-benzoic acid (49.5 g) in CH.sub.2Cl.sub.2 (200 mL). DMF (0.2 mL) is added and the solution is stirred at room temperature for 6 h. Then, the solution is concentrated under reduced pressure and the residue is dissolved in THF (100 mL). The resulting solution is cooled in an ice-bath and LiBH.sub.4 (3.4 g) is added in portions. The cooling bath is removed and the mixture is stirred at room temperature for 1 h. The reaction mixture is diluted with THF and treated with 0.1 M hydrochloric acid. Then, the organic layer is separated and the aqueous layer is extracted with ethyl acetate. The combined organic layers are dried (Na.sub.2SO.sub.4) and the solvent is evaporated under reduced pressure to give the crude product.

[0296] Yield: 47.0 g (99% of theory)

Preparation of 4-bromo-3-chloromethyl-1-iodo-benzene

[0297] ##STR00021##

[0298] Thionyl chloride (13 mL) is added to a suspension of 4-bromo-3-hydroxymethyl-1-iodo-benzene (47.0 g) in dichloromethane (100 mL) containing DMF (0.1 mL). The mixture is stirred at ambient temperature for 3 h. Then, the solvent and the excess reagent is removed under reduced pressure. The residue is triturated with methanol and dried.

[0299] Yield: 41.0 g (82% of theory)

Preparation of 4-bromo-1-iodo-3-phenoxymethyl-benzene

[0300] ##STR00022##

[0301] Phenol (13 g) dissolved in 4 M KOH solution (60 mL) is added to 4-bromo chloromethyl-1-iodo-benzene (41.0 g) dissolved in acetone (50 mL). NaI (0.5 g) is added and the resulting mixture is stirred at 50° C. overnight. Then, water is added and the resulting mixture is extracted with ethyl acetate. The combined extracts are dried (Na.sub.2SO.sub.4) and the solvent is evaporated under reduced pressure. The residue is purified by chromatography on silica gel (cyclohexane/ethyl acetate 19:1).

[0302] Yield: 38.0 g (79% of theory)

Preparation of 1-bromo-4-(1-methoxy-D-glucopyranos-1-yl)-2-(phenoxymethyl)-benzene

[0303] ##STR00023##

[0304] A 2 M solution of iPrMgCl in THF (11 mL) is added to dry LiCl (0.47 g) suspended in THF (11 mL). The mixture is stirred at room temperature until all the LiCl is dissolved. This solution is added dropwise to a solution of 4-bromo-1-iodo-3-phenoxymethyl-benzene (8.0 g) in tetrahydrofuran (40 mL) cooled to −60° C. under argon atmosphere. The solution is warmed to −40° C. and then 2,3,4,6-tetrakis-O-(trimethylsilyl)-D-glucopyranone (10.7 g, 90% pure) in tetrahydrofuran (5 mL) is added. The resulting solution is warmed to −5° C. in the cooling bath and stirred for another 30 min at this temperature. Aqueous NH.sub.4Cl solution is added and the resultant mixture is extracted with ethyl acetate. The combined organic extracts are dried over sodium sulfate and the solvent is removed under reduced pressure. The residue is dissolved in methanol (80 mL) and treated with methane sulfonic acid (0.6 mL) to produce the more stable anomer solely. After stirring the reaction solution at 35-40° C. overnight, the solution is neutralized with solid NaHCO.sub.3 and the methanol is removed under reduced pressure. The remainder is diluted with aqueous NaHCO.sub.3 solution and the resulting mixture is extracted with ethyl acetate. The combined extracts are dried over sodium sulfate and the solvent is evaporated to yield the crude product that is submitted to reduction without further purification.

[0305] Yield: 7.8 g (93% of theory)

Preparation of 1-bromo-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl) (phenoxymethyl)-benzene

[0306] ##STR00024##

[0307] Boron trifluoride diethyletherate (4.9 mL) is added to a solution of 1-bromo-4-(1-methoxy-D-glucopyranos-1-yl)-2-(phenoxymethyl)-benzene (8.7 g) and triethylsilane (9.1 mL) in dichloromethane (35 mL) and acetonitrile (50 mL) cooled to −20° C. at such a rate that the temperature maintains below −10° C. The resultant solution is warmed to 0° C. over a period of 1.5 h and then treated with aqueous sodium hydrogen carbonate solution. The resulting mixture is stirred for 0.5 h, the organic solvent is removed and the residue is extracted with ethyl acetate. The combined organic layers are dried over sodium sulfate and the solvent is removed. The residue is taken up in dichloromethane (50 mL) and pyridine (9.4 mL), acetic anhydride (9.3 mL) and 4-dimethylaminopyridine (0.5 g) are added in succession to the solution. The solution is stirred for 1.5 h at ambient temperature and then diluted with dichloromethane. This solution is washed twice with 1 M hydrochloric acid and dried over sodium sulfate. After the solvent is removed, the residue is recrystallized from ethanol to furnish the product as a colorless solid.

[0308] Yield: 6.78 g (60% of theory)

[0309] Mass spectrum (ESI.sup.+): m/z=610/612 (Br) [M+NH.sub.4].sup.+

Preparation of 2-(phenoxymethyl)-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-benzonitrile

[0310] ##STR00025##

[0311] A flask charged with zinc cyanide (1.0 g), zinc (30 mg), Pd.sub.2(dibenzylideneacetone).sub.3*CHCl.sub.3 (141 mg) and tri-tert-butylphosphonium tetrafluoroborate (111 mg) is flushed with argon. Then a solution of 1-bromo-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-2-(phenoxymethyl)-benzene (5.4 g) in NMP (12 mL) is added and the resulting mixture is stirred at room temperature for 18 h. After dilution with ethyl acetate, the mixture is filtered and the filtrate is washed with aqueous sodium hydrogen carbonate solution. The organic phase is dried (sodium sulfate) and the solvent is removed. The residue is recrystallized from ethanol.

[0312] Yield: 4.10 g (84% of theory)

[0313] Mass spectrum (ESI.sup.+): m/z=557 [M+NH.sub.4].sup.+

[0314] Alternatively, the compound described above is synthesized starting from 1-bromo-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-2-(phenoxymethyl)-benzene using copper(I) cyanide (2 equivalents) in NMP at 210° C.

Preparation of 2-bromomethyl-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-benzonitrile

[0315] ##STR00026##

[0316] A 33% solution of hydrobromic acid in acetic acid (15 mL) is added to a solution of 2-phenyloxymethyl-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-benzonitrile (0.71 g) and acetic anhydride (0.12 mL) in acetic acid (10 ml). The resulting solution is stirred at 55° C. for 6 h and then cooled in an ice-bath. The reaction mixture is neutralized with chilled aqueous potassium carbonate solution, and the resultant mixture is extracted with ethyl acetate. The combined organic extracts are dried over sodium sulfate and the solvent is removed under reduced pressure. The residue is taken up in ethyl acetate/cyclohexane (1:5), and the precipitate is separated by filtration and dried at 50° C. to give the pure product.

[0317] Yield: 0.52 g (75% of theory)

[0318] Mass spectrum (ESI.sup.+): m/z=543/545 (Br) [M+NH.sub.4].sup.+

Preparation of 4-cyclopropyl-phenylboronic acid

[0319] ##STR00027##

[0320] 2.5 M solution of nButyllithium in hexane (14.5 mL) is added dropwise to 1-bromo-4-cyclopropyl-benzene (5.92 g) dissolved in THF (14 mL) and toluene (50 mL) and chilled to −70° C. The resultant solution is stirred at −70° C. for 30 min before triisopropyl borate (8.5 mL) is added. The solution is warmed to −20° C. and then treated with 4 M aqueous hydrochloric acid (15.5 mL). The reaction mixture is further warmed to room temperature and then the organic phase is separated. The aqueous phase is extracted with ethyl acetate and the combined organic phases are dried (sodium sulfate). The solvent is evaporated and the residue is washed with a mixture of ether and cyclohexane to give the product as a colorless solid.

[0321] Yield: 2.92 g (60% of theory)

[0322] Mass spectrum (ESP): m/z=207 (Cl) [M+HCOO].sup.−

Preparation of 1-cyano-2-(4-cyclopropyl-benzyl)-4-(β-D-glucopyranos-1-yl)-benzene

[0323] ##STR00028##

[0324] An Ar filled flask is charged with 2-bromomethyl-4-(2,3,4,6-tetra-O-acetyl-D-glucopyranos-1-yl)-benzonitrile (1.60 g), 4-cyclopropyl-phenylboronic acid (1.0 g), potassium carbonate (1.85 g) and a degassed 3:1 mixture of acetone and water (22 mL). The mixture is stirred at room temperature for 5 min, before it is cooled in an ice-bath. Then palladium dichloride (30 mg) is added and the reaction mixture is stirred for 16 h at ambient temperature. The mixture is then diluted with brine and extracted with ethyl acetate. The combined extracts are dried over sodium sulfate and the solvent is removed under reduced pressure. The residue is dissolved in methanol (20 mL) and treated with 4 M aqueous potassium hydroxide solution (4 mL). The resulting solution is stirred at ambient temperature for 1 h and then neutralized with 1 M hydrochloric acid. The methanol is evaporated, and the residue is diluted with brine and extracted with ethyl acetate. The organic extracts collected are dried over sodium sulfate, and the solvent is removed. The residue is chromatographed on silica gel (dichloromethane/methanol 1:0->8:1).

[0325] Yield: 0.91 g (76% of theory)

[0326] Mass spectrum (ESI.sup.+): m/z=413 [M+NH.sub.4].sup.+

Preparation of a Crystalline Complex (1:1) of Compound A with L-Proline

[0327] L-proline (0.34 g) dissolved in 2.1 mL of a mixture of ethanol and water (volume ratio 10:1) is added to a solution of 1-cyano-2-(4-cyclopropyl-benzyl)-4-(β-D-glucopyranos yl)-benzene (1.17 g, obtained as described above) dissolved in 2 mL ethanol. The resulting solution is allowed to stand at ambient temperature. After about 16 h the crystalline complex is isolated as white crystals by filtration. If necessary the crystallization may be initiated by scratching with a glass rod or metal spatula for example or by inoculating with seed crystals. Residual solvent is removed by storing the crystals at slightly elevated temperature (30 to 50° C.) under vacuum for about 4 h to yield 1.27 g of the crystalline 1:1 complex of L-proline and 1-cyano-2-(4-cyclopropyl-benzyl)-4-(β-D-glucopyranos-1-yl)-benzene.

[0328] Several batches of the crystalline complex according to the above preparation are obtained. The X-ray powder diffraction patterns coincide. The melting points are determined via DSC and evaluated as onset-temperature. Examples of melting points are approximately 89° C., 90° C., 92° C., 101° C. and 110° C. The X-ray powder diffraction pattern as contained in Table 1 and as depicted in FIG. 11 and the DSC and TG diagram in FIG. 12 correspond to a batch with a melting point of approximately 90° C.

[0329] The X-ray powder diffraction pattern of the crystalline complex of the compound A and L-proline (peaks up to 30° in 2 Θ) is provided above in Table 1.

Example 13 Formulations

[0330] Some examples of formulations are described in which the term “active substance” denotes an SGLT2 inhibitor or pharmaceutically acceptable form thereof, e.g. a prodrug or a crystalline form, for use according to the invention. In the case of a combination with one or additional active substances, the term “active substance” may also include the additional active sub stance.

Tablets Containing 100 mg of Active Substance

[0331] Composition:

1 tablet contains:

TABLE-US-00003 active substance 100.0 mg Lactose 80.0 mg corn starch 34.0 mg Polyvinylpyrrolidone 4.0 mg magnesium stearate 2.0 mg 220.0 mg

[0332] Method of Preparation:

[0333] The active substance, lactose and starch are mixed together and uniformly moistened with an aqueous solution of the polyvinylpyrrolidone. After the moist composition has been screened (2.0 mm mesh size) and dried in a rack-type drier at 50° C. it is screened again (1.5 mm mesh size) and the lubricant is added. The finished mixture is compressed to form tablets.

Weight of tablet: 220 mg
Diameter: 10 mm, biplanar, facetted on both sides and notched on one side.

Tablets Containing 150 mg of Active Substance

[0334] Composition:

1 tablet contains:

TABLE-US-00004 active substance 150.0 mg powdered lactose 89.0 mg corn starch 40.0 mg colloidal silica 10.0 mg Polyvinylpyrrolidone 10.0 mg magnesium stearate 1.0 mg 300.0 mg

[0335] Preparation:

[0336] The active substance mixed with lactose, corn starch and silica is moistened with a 20% aqueous polyvinylpyrrolidone solution and passed through a screen with a mesh size of 1.5 mm. The granules, dried at 45° C., are passed through the same screen again and mixed with the specified amount of magnesium stearate. Tablets are pressed from the mixture. [0337] Weight of tablet: 300 mg [0338] die: 10 mm, flat

Hard Gelatin Capsules Containing 150 mg of Active Substance

[0339] Composition:

1 capsule contains:

TABLE-US-00005 active substance 150.0 mg corn starch (dried) approx. 180.0 mg lactose (powdered)  approx. 87.0 mg magnesium stearate  3.0 mg approx. 420.0 mg

[0340] Preparation:

[0341] The active substance is mixed with the excipients, passed through a screen with a mesh size of 0.75 mm and homogeneously mixed using a suitable apparatus. The finished mixture is packed into size 1 hard gelatin capsules. [0342] Capsule filling: approx. 320 mg [0343] Capsule shell: size 1 hard gelatin capsule.

Suppositories Containing 150 mg of Active Substance

[0344] Composition:

1 suppository contains:

TABLE-US-00006 active substance 150.0 mg polyethylene glycol 1500 550.0 mg polyethylene glycol 6000 460.0 mg polyoxyethylene sorbitan monostearate 840.0 mg 2,000.0 mg.sup. 

[0345] Preparation:

[0346] After the suppository mass has been melted the active substance is homogeneously distributed therein and the melt is poured into chilled molds.

Ampoules Containing 10 mg Active Substance

[0347] Composition:

TABLE-US-00007 active substance 10.0 mg 0.01N hydrochloric acid/NaCl q.s. double-distilled water ad 2.0 ml

[0348] Preparation:

[0349] The active substance is dissolved in the necessary amount of 0.01 N HCl, made isotonic with common salt, filtered sterile and transferred into 2 ml ampoules.

Ampoules Containing 50 mg of Active Substance

[0350] Composition:

TABLE-US-00008 active substance 50.0 mg 0.01N hydrochloric acid/NaCl q.s. double-distilled water ad 10.0 ml

[0351] Preparation:

[0352] The active substance is dissolved in the necessary amount of 0.01 N HCl, made isotonic with common salt, filtered sterile and transferred into 10 ml ampoules.

REFERENCES

[0353] 1) Curry et al., Comp Biochem Physiol. 1982. 72A(2): 333-338 [0354] 2) EP 1 213 296 [0355] 3) EP 1 354 888 [0356] 4) EP 1 344 780 [0357] 5) EP 1 489 089 [0358] 6) Hoenig, Mol Cell Endocrinol 2002, 197(1-2): 221-229 [0359] 7) Hoenig et al., Am J Physiol, 2011, 301(6):R1798-1807 [0360] 8) NADA 141-236 Freedom of Information VETSULIN® [0361] 9) Palm C A et al., Vet Clin Small Anim 2013, 43: 407-415 [0362] 10) Reusch C E et al., Schweizer Archiv fuer Tierheilkunde 2011, 153811): 495-500 [0363] 11) Tanaka et al., Vet Res Commun. 2005, 29(6):477-485 [0364] 12) Verbrugghe et al., Crit Rev Food Sci Nutr. 2012; 52(2):172-182 [0365] 13) WO 01/27128 [0366] 14) WO 03/099836 [0367] 15) WO 2004/007517 [0368] 16) WO 2004/080990 [0369] 17) WO 2005/012326 [0370] 18) WO 2005/092877 [0371] 19) WO 2006/034489 [0372] 20) WO 2006/064033 [0373] 21) WO 2006/117359 [0374] 22) WO 2006/117360 [0375] 23) WO 2006/120208 [0376] 24) WO 2007/025943 [0377] 25) WO 2007/028814 [0378] 26) WO 2007/031548 [0379] 27) WO 2007/093610 [0380] 28) WO 2007/114475 [0381] 29) WO 2007/128749 [0382] 30) WO 2007/140191 [0383] 31) WO 2008/002824 [0384] 32) WO 2008/013280 [0385] 33) WO 2008/042688 [0386] 34) WO 2008/049923 [0387] 35) WO 2008/055870 [0388] 36) WO 2008/055940 [0389] 37) WO 2008/069327 [0390] 38) WO 2008/116179 [0391] 39) WO 2009/014970 [0392] 40) WO 2009/022008 [0393] 41) WO 2009/022020 [0394] 42) WO 2009/035969 [0395] 43) WO 2010/023594 [0396] 44) WO 2011/039107 [0397] 45) WO 2011/039108 [0398] 46) WO 2011/117295 [0399] 47) WO 2014/016381

[0400] The above references are incorporated by reference.