Kenji Arakawa

Effect of canagliflozin on renal threshold for glucose, glycemia, and body weight in normal and diabetic animal models.

Background Canagliflozin is a sodium glucose co-transporter (SGLT) 2 inhibitor in clinical development for the treatment of type 2 diabetes mellitus (T2DM). Methods 14C-alpha-methylglucoside uptake in Chinese hamster ovary-K cells expressing human, rat, or mouse SGLT2 or SGLT1; 3H-2-deoxy-d-glucose uptake in L6 myoblasts; and 2-electrode voltage clamp recording of oocytes expressing human SGLT3 were analyzed. Graded glucose infusions were performed to determine rate of urinary glucose excretion (UGE) at different blood glucose (BG) concentrations and the renal threshold for glucose excretion (RTG) in vehicle or canagliflozin-treated Zucker diabetic fatty (ZDF) rats. This study aimed to characterize the pharmacodynamic effects of canagliflozin in vitro and in preclinical models of T2DM and obesity. Results Treatment with canagliflozin 1 mg/kg lowered RTG from 415±12 mg/dl to 94±10 mg/dl in ZDF rats while maintaining a threshold relationship between BG and UGE with virtually no UGE observed when BG was below RTG. Canagliflozin dose-dependently decreased BG concentrations in db/db mice treated acutely. In ZDF rats treated for 4 weeks, canagliflozin decreased glycated hemoglobin (HbA1c) and improved measures of insulin secretion. In obese animal models, canagliflozin increased UGE and decreased BG, body weight gain, epididymal fat, liver weight, and the respiratory exchange ratio. Conclusions Canagliflozin lowered RTG and increased UGE, improved glycemic control and beta-cell function in rodent models of T2DM, and reduced body weight gain in rodent models of obesity.

Intestinal Sodium Glucose Cotransporter 1 Inhibition Enhances Glucagon-Like Peptide-1 Secretion in Normal and Diabetic Rodents.

The sodium glucose cotransporter (SGLT) 1 plays a major role in glucose absorption and incretin hormone release in the gastrointestinal tract; however, the impact of SGLT1 inhibition on plasma glucagon-like peptide-1 (GLP-1) levels in vivo is controversial. We analyzed the effects of SGLT1 inhibitors on GLP-1 secretion in normoglycemic and hyperglycemic rodents using phloridzin, CGMI [3-(4-cyclopropylphenylmethyl)-1-(β-d-glucopyranosyl)-4-methylindole], and canagliflozin. These compounds are SGLT2 inhibitors with moderate SGLT1 inhibitory activity, and their IC50 values against rat SGLT1 and mouse SGLT1 were 609 and 760 nM for phloridzin, 39.4 and 41.5 nM for CGMI, and 555 and 613 nM for canagliflozin, respectively. Oral administration of these inhibitors markedly enhanced and prolonged the glucose-induced plasma active GLP-1 (aGLP-1) increase in combination treatment with sitagliptin, a dipeptidyl peptidase-4 (DPP4) inhibitor, in normoglycemic mice and rats. CGMI, the most potent SGLT1 inhibitor among them, enhanced glucose-induced, but not fat-induced, plasma aGLP-1 increase at a lower dose compared with canagliflozin. Both CGMI and canagliflozin delayed intestinal glucose absorption after oral administration in normoglycemic rats. The combined treatment of canagliflozin and a DPP4 inhibitor increased plasma aGLP-1 levels and improved glucose tolerance compared with single treatment in both 8- and 13-week-old Zucker diabetic fatty rats. These results suggest that transient inhibition of intestinal SGLT1 promotes GLP-1 secretion by delaying glucose absorption and that concomitant inhibition of intestinal SGLT1 and DPP4 is a novel therapeutic option for glycemic control in type 2 diabetes mellitus.

Canagliflozin, a sodium glucose cotransporter 2 inhibitor, attenuates obesity-induced inflammation in the nodose ganglion, hypothalamus, and skeletal muscle of mice

&NA; Chronic inflammation in systemic organs, such as adipose tissue, nodose ganglion, hypothalamus, and skeletal muscles, is closely associated with obesity and diabetes mellitus. Because sodium glucose cotransporter 2 (SGLT2) inhibitors exert both anti‐diabetic and anti‐obesity effects by promoting urinary excretion of glucose and subsequent caloric loss, we investigated the effect of canagliflozin, an SGLT2 inhibitor, on obesity‐induced inflammation in neural tissues and skeletal muscles of mice. High‐fat diet (HFD)‐fed male C57BL/6J mice were treated with canagliflozin for 8 weeks. Canagliflozin attenuated the HFD‐mediated increases in body weight, liver weight, and visceral and subcutaneous fat weight. Additionally, canagliflozin decreased blood glucose as well as the fat, triglyceride, and glycogen contents of the liver. Along with these metabolic corrections, canagliflozin attenuated the increases in the mRNA levels of the proinflammatory biomarkers Iba1 and Il6 and the number of macrophages/microglia in the nodose ganglion and hypothalamus. In the skeletal muscle of HFD‐fed obese mice, canagliflozin decreased inflammatory cytokine levels, macrophage accumulation, and the mRNA level of the specific atrophic factor atrogin‐1. Canagliflozin also increased the mRNA level of insulin‐like growth factor 1, protected against muscle mass loss, and restored the contractile force of muscle. These findings suggested that SGLT2 inhibition disrupts the vicious cycle of obesity and inflammation, not only by promoting caloric loss, but also by suppression of obesity‐related inflammation in both the nervous system and skeletal muscle.

Beneficial Effects of Canagliflozin in Combination with Pioglitazone on Insulin Sensitivity in Rodent Models of Obese Type 2 Diabetes

Background Despite its insulin sensitizing effects, pioglitazone may induce weight gain leading to an increased risk of development of insulin resistance. A novel sodium glucose co-transporter 2 (SGLT2) inhibitor, canagliflozin, provides not only glycemic control but also body weight reduction through an insulin-independent mechanism. The aim of this study was to investigate the combined effects of these agents on body weight control and insulin sensitivity. Methods Effects of combination therapy with canagliflozin and pioglitazone were evaluated in established diabetic KK-Ay mice and prediabetic Zucker diabetic fatty (ZDF) rats. Results In the KK-Ay mice, the combination therapy further improved glycemic control compared with canagliflozin or pioglitazone monotherapy. Furthermore, the combination significantly attenuated body weight and fat gain induced by pioglitazone and improved hyperinsulinemia. In the ZDF rats, early intervention with pioglitazone monotherapy almost completely prevented the progressive development of hyperglycemia, and no further improvement was observed by add-on treatment with canagliflozin. However, the combination significantly reduced pioglitazone-induced weight gain and adiposity and improved the Matsuda index, suggesting improved whole-body insulin sensitivity. Conclusions Our study indicates that combination therapy with canagliflozin and pioglitazone improves insulin sensitivity partly by preventing glucotoxicity and, at least partly, by attenuating pioglitazone-induced body weight gain in two different obese diabetic animal models. This combination therapy may prove to be a valuable option for the treatment and prevention of obese type 2 diabetes.

Analysis of the Effect of Canagliflozin on Renal Glucose Reabsorption and Progression of Hyperglycemia in Zucker Diabetic Fatty Rats

Sodium–glucose cotransporter 2 (SGLT2) plays a major role in renal glucose reabsorption. To analyze the potential of insulin-independent blood glucose control, the effects of the novel SGLT2 inhibitor canagliflozin on renal glucose reabsorption and the progression of hyperglycemia were analyzed in Zucker diabetic fatty (ZDF) rats. The transporter activity of recombinant human and rat SGLT2 was inhibited by canagliflozin, with 150- to 12,000-fold selectivity over other glucose transporters. Moreover, in vivo treatment with canagliflozin induced glucosuria in mice, rats, and dogs in a dose-dependent manner. It inhibited apparent glucose reabsorption by 55% in normoglycemic rats and by 94% in hyperglycemic rats. The inhibition of glucose reabsorption markedly reduced hyperglycemia in ZDF rats but did not induce hypoglycemia in normoglycemic animals. The change in urinary glucose excretion should not be used as a marker to predict the glycemic effects of this SGLT2 inhibitor. In ZDF rats, plasma glucose and HbA1c levels progressively increased with age, and pancreatic β-cell failure developed at 13 weeks of age. Treatment with canagliflozin for 8 weeks from the prediabetic stage suppressed the progression of hyperglycemia, prevented the decrease in plasma insulin levels, increased pancreatic insulin contents, and minimized the deterioration of islet structure. These results indicate that selective inhibition of SGLT2 with canagliflozin controls the progression of hyperglycemia by inhibiting renal glucose reabsorption in ZDF rats. In addition, the preservation of β-cell function suggests that canagliflozin treatment reduces glucose toxicity via an insulin-independent mechanism.

The effect of combined treatment with canagliflozin and teneligliptin on glucose intolerance in Zucker diabetic fatty rats.

To assess the impact of concomitant inhibition of sodium-glucose cotransporter (SGLT) 2 and dipeptidyl peptidase IV (DPP4) for the treatment of type 2 diabetes mellitus (T2DM), the effect of combined treatment with canagliflozin, a novel SGLT2 inhibitor, and teneligliptin, a DPP4 inhibitor, on glucose intolerance was investigated in Zucker diabetic fatty (ZDF) rats. Canagliflozin potently inhibited human and rat SGLT2 and moderately inhibited human and rat SGLT1 activities but did not affect DPP4 activity. In contrast, teneligliptin inhibited human and rat DPP4 activities but not SGLT activities. A single oral treatment of canagliflozin and teneligliptin suppressed plasma glucose elevation in an oral glucose tolerance test in 13 week-old ZDF rats. This combination of agents elevated plasma active GLP-1 levels in a synergistic manner, probably mediated by intestinal SGLT1 inhibition, and further improved glucose intolerance. In the combination-treated animals, there was no pharmacokinetic interaction of the drugs and no further inhibition of plasma DPP4 activity compared with that in the teneligliptin-treated animals. These results suggest that the inhibition of SGLT2 and DPP4 improves glucose intolerance and that combined treatment with canagliflozin and teneligliptin is a novel therapeutic option for glycemic control in T2DM.

Changes in glucose-induced plasma active glucagon-like peptide-1 levels by co-administration of sodium–glucose cotransporter inhibitors with dipeptidyl peptidase-4 inhibitors in rodents

We investigated whether structurally different sodium-glucose cotransporter (SGLT) 2 inhibitors, when co-administered with dipeptidyl peptidase-4 (DPP4) inhibitors, could enhance glucagon-like peptide-1 (GLP-1) secretion during oral glucose tolerance tests (OGTTs) in rodents. Three different SGLT inhibitors-1-(β-d-Glucopyranosyl)-4-chloro-3-[5-(6-fluoro-2-pyridyl)-2-thienylmethyl]benzene (GTB), TA-1887, and canagliflozin-were examined to assess the effect of chemical structure. Oral treatment with GTB plus a DPP4 inhibitor enhanced glucose-induced plasma active GLP-1 (aGLP-1) elevation and suppressed glucose excursions in both normal and diabetic rodents. In DPP4-deficient rats, GTB enhanced glucose-induced aGLP-1 elevation without affecting the basal level, whereas metformin, previously reported to enhance GLP-1 secretion, increased both the basal level and glucose-induced elevation. Oral treatment with canagliflozin and TA-1887 also enhanced glucose-induced aGLP-1 elevation when co-administered with either teneligliptin or sitagliptin. These data suggest that structurally different SGLT2 inhibitors enhance plasma aGLP-1 elevation and suppress glucose excursions during OGTT when co-administered with DPP4 inhibitors, regardless of the difference in chemical structure. Combination treatment with DPP4 inhibitors and SGLT2 inhibitors having moderate SGLT1 inhibitory activity may be a promising therapeutic option for improving glycemic control in patients with type 2 diabetes mellitus.

Involvement of heparan sulfate in the renoprotective effects of imidapril, an angiotensin-converting enzyme inhibitor, in diabetic db/db mice

Abstract We investigated the renoprotective effects of imidapril hydrochloride ((-)-(4 S)-3-[(2 S)-2-[[(1 S)-1-ethoxycarbonyl-3-phenylpropyl] amino] propionyl]-1-methyl-2-oxoimidazolidine-4-carboxylic acid hydrochloride, imidapril), an angiotensin-converting enzyme inhibitor, in a diabetic animal model. We used BKS.Cg-+Leprdb/+Leprdb (db/db) mice, a genetic animal model of obese type 2 diabetes. Diabetic db/db mice suffered from glomerular hyperfiltration, albuminuria and hypoalbuminemia. Oral administration of 5 mg/kg/day of imidapril for 3 weeks suppressed renal hyperfiltration, reduced albuminuria and normalized hypoalbuminemia. Imidapril did not influence body weights, blood pressure or blood glucose concentrations in db/db mice. Urinary excretion of heparan sulfate (HS) in non-treated 11-week-old db/db mice was significantly lower than that in age-matched non-diabetic db/+m mice. HS is a component of HS proteoglycans, which are present in glomerular basement membranes and glycocalyx of cell surfaces. Reduced urinary HS excretion indicated glomerular HS loss in db/db mice. Imidapril increased urinary excretion of HS to concentrations observed in db/+m mice, indicating that imidapril prevented the loss of renal HS. These results suggest that imidapril ameliorates renal hyperfiltration and loss of renal contents of HS. Improvement of filtration function and maintenance of HS, which is an important structural component of glomeruli, may contribute to renoprotective effects of imidapril.

Effect of Canagliflozin on Urinary Albumin Excretion in Japanese Patients with Type 2 Diabetes Mellitus and Microalbuminuria: A Pilot Study

Abstract Background: Albuminuria characterizes the progression of kidney injury. The effect of canagliflozin on the excretion of microalbumin was assessed for investigating its renoprotective potential in Japanese patients with type 2 diabetes mellitus (T2DM). Patients and Methods: Twenty Japanese patients with T2DM and microalbuminuria were enrolled and administered with 100 mg of canagliflozin once a day for 12 weeks. These subjects were admitted to the clinic at the start and end of the treatment period for 24-h urine collection. The primary endpoint was the percentage change in geometric mean 24-h urinary albumin excretion from baseline to week 12. Results: The urinary albumin level decreased by 42.0% (95% confidence interval: 21.9–57.0; P = 0.0011) after 12 weeks of canagliflozin treatment. A number of blood and urinary parameters also significantly decreased, including hemoglobin A1c, fasting plasma glucose, estimated glomerular filtration rate, and creatinine clearance, while hematocrit was elevated. Among the biomarkers associated with kidney injury and inflammation, the urinary level of the oxidative stress marker 8-hydroxy-2′-deoxyguanosine was also decreased. There were no meaningful correlations noted between changes in urinary albumin excretion and other parameters/biomarkers. No severe adverse events were reported over the 12-week treatment period. Conclusions: The results of this study indicate that canagliflozin decreases microalbuminuria in Japanese patients with T2DM. Albuminuria could be reduced as a result of changes in various physiological pathways; therefore, it is imperative that future, large-scale, studies attempt to determine the detailed mechanisms involved. Canagliflozin may offer a novel therapeutic option for Japanese patients with T2DM and incipient nephropathy.