2025 Volume 48 Issue 1 Pages 65-74
Pharmacological activation of G protein-coupled receptor 119 (GPR119) produces pleiotropic beneficial effects, including the promotion of insulin secretion from pancreatic β-cells, enhancement of glucagon-like peptide (GLP)-1 secretion from intestinal L cells, glucose-dependent insulin secretion, and food intake and body weight gain suppression. Thus, GPR119 has attracted attention as a promising new target for type 2 diabetes mellitus (T2DM) treatment. Here, we identified a new small GPR119 agonist, NCP-322. This compound showed potent enhancing effects on insulin and GLP-1 secretion, which played a role in pancreatic β-cells and intestinal L cells. In the oral glucose tolerance test, NCP-322 administration reduced glycemic excursions that were only exhibited during hyperglycemia. Furthermore, NCP-322 administration did not induce hypoglycemia, the main side effect of antidiabetic drugs. These results suggest the promising therapeutic potential of NCP-322 for T2DM treatment.
The prevalence of type 2 diabetes mellitus (T2DM) has markedly increased worldwide. T2DM treatment involves the combined use of multiple hypoglycemic agents with different mechanisms, including stimulating insulin secretion, improving insulin resistance, and regulating glucose absorption and excretion, in addition to exercise and dietary therapy. Sulfonylurea antidiabetic drugs, which are insulin secretion stimulants, exhibit potent hypoglycemic effects; however, severe hypoglycemia is the main issue.1–3) Moreover, the long-term administration of sulfonylurea antidiabetic drugs decreases drug efficacy.4,5) To improve insulin resistance, biguanides and thiazolidinediones have been frequently used, and as glucose absorption and excretion regulators, alpha-glucosidase inhibitors have also been frequently employed in the treatment strategy for T2DM. However, these medications can cause side effects, including gastrointestinal symptoms (diarrhea, vomiting, abdominal distension, and constipation), weight gain, and edema.6–11) SGLT2 inhibitors have been recently used as a new treatment for T2DM for regulating glucose excretion; however, these inhibitors also have side effects, including urinary tract infections and nocturia.12–14) Incretin-based therapies, including glucagon-like peptide (GLP)-1 mimetic and dipeptidyl peptidase 4 (DPP-IV) inhibitors, have attracted attention for T2DM treatment as they are less likely to cause hypoglycemia and are expected to cause weight loss due to their appetite-suppressing effects. However, not all incretin-based therapies can cause weight loss, and side effects, including gastrointestinal symptoms (nausea and diarrhea), have been reported.15,16) Thus, the current treatments have drawbacks, and, most importantly, the loss of responsiveness of insulin secretion stimulants, including sulfonylurea antidiabetic drugs, which occur over time.17) Over the last 2 decades, despite the development of several new classes of drugs for T2DM treatment, an unmet need remains.
G protein-coupled receptor 119 (GPR119), which was initially identified in 2003 as an orphan 7-transmembrane spanning G protein-coupled receptor (GPCR), has been increasingly recognized as a novel target in T2DM treatment.18) GPR119 is expressed in islet β-cells, entero-endocrine K and L cells, and the brain.19,20) GPR119 is activated by lipid amides, including oleoylethanolamide and lysophosphatidylcholine,20,21) resulting in increased intracellular cAMP accumulation and glucose-dependent insulin secretion. Reportedly, oleoylethanolamide can suppress obesity, which increases the risk of diabetes development, owing to its appetite-suppressing effects.21) Furthermore, its GPR119 agonistic activity is reportedly more than 10-fold greater than that of lysophosphatidylcholine, which was initially reported as an endogenous ligand for GPR119.22) Additionally, synthetic GPR119 agonists, AR231453 and PSN632408, have been shown to increase intracellular cAMP levels, glucose-dependent insulin secretion, and food intake and body weight in a rat model.21,23) Moreover, reportedly, GPR119 agonists stimulate GLP-1 secretion in human and mouse intestinal L-cell models.24,25)
We here newly identified a potent GPR119 agonist, NCP-322. By analyzing the pharmacological effects of GLP-1 on insulin secretion and hypoglycemia, along with a comparison of the efficacy against previously developed GPR119 agonists using various in vitro and in vivo experiments, we demonstrated that NCP-322 promotes insulin and GLP-1 secretion at low doses without causing marked side effects, including hypoglycemia, which have been issues with previous therapeutic drugs.
Male C57BL/6J mice (Charles River Laboratories Japan, Inc., Yokohama, Japan) (8- to 11-weeks old at the start of the experiments) were used. They were housed under controlled conditions (temperature, 21–25°C; humidity, 4–70%). The room was lit from 7:00 to 19:00 h. Food and water were available ad libitum. The Committee for Animal Experiments at Discovery Research Laboratories, Nippon Chemiphar Co., Ltd. (Saitama, Japan) approved this study.
DrugsNCP-322 (Isopropyl 4-[3-fluoro-2-[7-fluoro-5-(tetrazol-1-yl)indol-1-ylmethyl]pyridin-5-yl]piperidine-1-carboxylate) and MBX-2982 were synthesized at Discovery Research Laboratories, Nippon Chemiphar Co., Ltd. (Saitama, Japan). The chemical structure of NCP-322 is shown in Fig. 1.
GPR119-expressing cell lines were generated using the FIp-In T-REx293 tetracycline-inducible gene expression system (Invitrogen, Carlsbad, CA, U.S.A. #R780-07). The human GPR119 gene (NM_178471) was purchased from the American Type Culture Collection (ATCC, #10807349). Mouse GPR119 gene (NM_181751) was amplified by performing PCR on the NIT-1 cell line (ATCC, #CRL-2055, β-cell line isolated from the islet of Langerhans) purchased from ATCC. The forward primer with an added BamHI site (human GPR119: TCCTGGATCCatggaatcatctttctcatt, mouse GPR119: TCCTGGATCCatggagtcatccttctcatt), and the reverse primer with an added ApaI site (human GPR119: TCCTGGGCCCttagccatcaaactctgagc, mouse GPR119: TCCTGGGCCCttagccatcgagctccggat) were designed. The target gene was amplified by PCR using a KOD-Plus-Ver.2 (TOYOBO #KOD-211). PCR was repeated 3 steps (human GPR119: 98°C for 10 s, 55°C for 30 s, 68°C for 1 min 15 s; mouse GPR119: 98°C for 10 s, 50°C for 30 s, 68°C for 1 min 30 s) in 35 cycles. The amplified PCR product was inserted into the pcDNA5/FRT/TO vector (Invitrogen, #V6520-20), and the FIp-In T-REx293 tetracycline-inducible gene expression system was used to create a stable cell line that expressed the target gene when induced with tetracycline. These cell lines were cultured in a culture medium comprising the following components: Dulbecco’s modified Eagle medium (DMEM; Invitrogen, #11965-092), 10% fetal bovine serum (FBS) (Bio-West, Inc., France. #S1560-500), 1% penicillin–streptomycin (Invitrogen, #15070-063), 1 mmol/L (mM) sodium pyruvate (Invitrogen, #11360-070), 100 μg/mL of hygromycin B (Invitrogen, #R220-05), and 15 μg/mL of blasticidin (Invitrogen, #R210-01).
For the cell-based cAMP assays, cells were plated on 96-half well white plates (Greiner Bio-One Co., Ltd., Tokyo, Japan. #675074) at a density of approximately 1000 cells/well in an assay medium (DMEM, 10% FBS, 1% penicillin–streptomycin, and 1 mM sodium pyruvate). To induce GPR119 expression, the assay medium containing tetracycline (Invitrogen, #Q10019) was added to the cells after 24 h and incubated for another 24 h at 37°C.
Following incubation, the assay medium was aspirated; subsequently, an assay buffer containing the test compound was added to the GPR119-expressed cells. After incubation for 30 min at 37°C, the intracellular cAMP concentration was determined using the HITHUNTER® cAMP kit (DiscoveRX, Fremont, CA, U.S.A. #90-0075A) following the manufacturer’s protocol. The plate was read using a Fluostar optima plate reader (BMG LABTECH JAPAN Ltd., Saitama, Japan).
Glucose-Stimulated Insulin Secretion AssessmentC57BL/6J mice aged 9- to 11-week old were euthanized by cervical dislocation, and the pancreas was quickly removed. The pancreatic islets were isolated as previously described.23) Subsequently, the islets were handpicked under a stereomicroscope, and the isolated islets were digested up to β-cells using 0.04% ethylenediaminetetraacetic acid (EDTA) and 1000 PU of dispase (GODO SHUSEI CO., LTD. # GD 81060).26) β-Cells were resuspended and plated on a 48-well plate at a density of approximately 5000 cells/well in a culture medium (DMEM, 10% FBS, and 1% penicillin–streptomycin). After a 2-d culture, β-cells were washed with a HEPES-balanced Krebs–Ringer bicarbonate buffer containing 3.3 mM glucose and 0.1% bovine serum albumin (BSA) and preincubated for 30 min at 37°C with 3.3 mM glucose containing the test compound or vehicle (final concentration 1% dimethyl sulfoxide). Following preincubation, β-cells were incubated for 30 min at 37°C with 25 mM glucose containing the test compound or vehicle. Insulin levels secreted into the supernatant were measured using an enzyme-linked immunosorbent assay (ELISA) kit (Shibayagi Co., Ltd., Gunma, Japan. #AKRIN-011H).
Effects of Stimulating GLP-1 SecretionC57BL/6J mice aged 8- to 11-week old were fasted overnight and subsequently orally administered with a vehicle (1% carboxymethyl cellulose [CMC] containing 2% Tween 80], NCP-322 (0.1–3 mg/kg), or MBX-2982 (0.1–3 mg/kg). The DPP-IV inhibitor, sitagliptin (10 mg/kg), was pretreated orally at 40 min before administering the test compounds. For the determination of GLP-1 (active form), portal vein blood was collected in the tubes containing the DPP-IV inhibitor (Millipore Corp., Boston, MA, U.S.A. #DPP4) at 30 min following treatment with the test compounds. Plasma GLP-1 (active form) levels were determined using an ELISA kit (Millipore, #EGLP-35K).
Oral Glucose Tolerance TestC57BL/6J mice aged 9- to 11-week old were fasted for 16 h and subsequently orally administered with a vehicle (1% CMC containing 2% Tween 80), glibenclamide (1 or 3 mg/kg), NCP-322 (0.1–30 mg/kg), or MBX-2982 (0.1–30 mg/kg). After 30 min, glucose was orally administered at 3 g/kg dose. Tail vein blood was collected using the heparinized capillary (calibrated pipet) before administering the test compounds (defined as –30 min value), immediately before the glucose challenge (0 min), or at 30, 60, and 120 min following glucose ingestion. Blood glucose levels were determined using a Glucose CII Test Wako (Wako Pure Chemical Corporation, Osaka, Japan. # 439-90901). Glycemic excursions for plasma glucose were presented as absolute values. Additionally, after subtracting the glucose concentration before the glucose challenge, the absolute areas under the glycemic excursions were calculated between 0 and 120 min after the glucose challenge and were referred to as the area under the curve (AUC).
GLP-1 and Insulin Secretion Pharmacodynamic AnalysisC57BL/6J mice aged 9- to 11-week old were fasted overnight and subsequently orally administered with a vehicle (1% CMC containing 2% Tween 80) or NCP-322 (1 mg/kg). After 30 min, a glucose challenge (3 g/kg) was administered orally. Abdominal vein blood was collected at 0 (just before glucose administration) and 5, 10, 20, 30, and 40 min after the glucose challenge, and the plasma was obtained for glucose and insulin level determination using a Glucose CII Test Wako and an ELISA kit, respectively. For GLP-1 (active) determination, portal vein blood was collected in tubes containing a DPP-IV inhibitor at the same time points. Plasma GLP-1 (active) levels were determined using an ELISA kit.
Statistical AnalysisStatistical analyses were performed using GraphPad Prism version 5.0.3 (GraphPad Software, Inc., San Diego, CA, U.S.A.). All data were presented as means and standard errors of the mean. Statistical significance was analyzed using Student’s t-test or one-way ANOVA followed by Dunnett’s multiple comparison test. A p-value of <0.05 was considered statistically significant. Using the Smirnov–Grubbs test, a method for outlier detection assuming a normal data distribution, outliers were identified and eliminated.
The chemical structure of NCP-322 is depicted in Fig. 1. In human or mouse GPR119-expressing cell lines, we examined the agonistic effects of NCP-322 compared with AR231453 (a GPR119 agonist previously developed by Arena27)) and confirmed that NCP-322 increased the intracellular cAMP levels in a concentration-dependent manner (human GPR119: EC50 = 9.4 nmol/L, Emax = 95% of AR231453 response; mouse GPR119: EC50 = 68 nmol/L, Emax = 100% of AR231453 response, respectively) (Table 1). NCP-322 and AR231453 did not increase the cAMP levels in the control host cell line without GPR119 expression (Fig. 2). Furthermore, the Emax value of NCP-322 for human GPR119 was comparable to that of another GPR119 agonist, MBX-2982, developed by Metabolex,28) and NCP-322 had a 31% higher Emax value in mouse GPR119 (Table 1).
| Compound | EC50: nmol/L (Emax: %AR231453) | ||
|---|---|---|---|
| Human | Mouse | CAT | |
| NCP-322 | 9.4 (95%) | 68.0 (100%) | N.D. |
| MBX-2982 | 13.7 (92%) | 45.9 (69%) | N.T. |
EC50 values of NCP-322 and MBX-2982 on intracellular cAMP production in human and mouse GPR119-expressing FIp-In T-REx293 cells and in CAT-expressing FIp-In T-REx293 cells. Emax is calculated as the percentage of the maximum response of AR231453. EC50 is the concentration producing 50% of Emax. N.D.: not detected; N.T.: not tested.
NCP-322 showing a potent agonist activity comparable to AR231453 in human and mouse GPR119-expressing FIp-In T-REx293 cells. Effects of NCP-322 on intracellular cAMP production in human (A) and mouse (B) GPR119-expressing FIp-In T-REx293 cells and in CAT-expressing FIp-In T-REx293 cells (C) (n = 3–6 wells).
A previous study has demonstrated that the insulinotropic effect of GPR119 agonists in the islets is glucose dependent and that the GPR119 agonist stimulates insulin release in isolated mouse islets at glucose concentrations of 8–17 mM.23) We examined the effects of NCP-322 compared with MBX-2982 on glucose-stimulated insulin secretion from isolated mouse β-cells. In the presence of high glucose levels (25 mM), NCP-322 dose-dependently enhanced insulin secretion from mouse β-cells (Fig. 3A); however, MBX-2982 showed no significant effects in this experimental condition (Fig. 3B). Although glibenclamide, a sulfonylurea antidiabetic agent that stimulates insulin secretion in a glucose-independent manner, markedly enhanced insulin secretion from mouse β-cells in the presence of low glucose levels (3.3 mM), NCP-322 had no effect (Fig. 3C). These results indicate that the insulinotropic effect of NCP-322 is produced under high-glucose conditions.
β-Cells are washed with HEPES balanced Krebs–Ringer bicarbonate buffer containing 3.3 mM glucose and 0.1% BSA and preincubated for 30 min at 37°C with 3.3 mM glucose containing the test compound (NCP-322 or glibenclamide [Gliben.]) or vehicle (final concentration 1% dimethyl sulfoxide). Following preincubation, β-Cells are incubated for 30 min at 37°C with 25 mM glucose containing the test compound (NCP-322, MBX-2982, or Gliben.) or vehicle. Data are presented as means ± S.E.M. (n = 2–7 wells). * p < 0.05, ** p < 0.01, N.S.: no significant versus vehicle (one-way ANOVA Dunnett’s multiple comparison test). S.E.M.: standard error of the mean.
GPR119 activation stimulates GLP-1 secretion from intestinal L cells.24,25) Therefore, we next evaluated the effect of NCP-322 compared with MBX-2982 on GLP-1 secretion in an in vivo model. As GLP-1 has a very short half-life in vivo, we used mice that had been pretreated with the DPP-IV inhibitor, sitagliptin, to prevent GLP-1 degradation. Compared with the nontreated control group, the oral administration of sitagliptin (10 mg/kg) alone (vehicle group) significantly increased the plasma GLP-1 levels (Fig. 4A). NCP-322 (0.3–3 mg/kg) oral administration significantly increased the plasma GLP-1 levels in sitagliptin-pretreated mice (Fig. 4A). By contrast, MBX-2982 increased the plasma GLP-1 levels only at high doses (Fig. 4B).
Vehicle (1% CMC containing 2% Tween 80), NCP-322, or MBX-2982 is administered to C57BL/6J mice under fasting conditions. The DPP-IV inhibitor, sitagliptin (10 mg/kg), is pretreated orally at 40 min before the administration of the vehicle or test compounds (NCP-322 or MBX-2982). The plasma GLP-1 concentration is measured for 30 min following test compound administration (A, NCP-322; B, MBX-2982). Data are presented as means ± S.E.M. (n = 5–7 mice). ## p < 0.01 versus the control group (Student’s t-test), * p < 0.05, ** p < 0.01, and *** p < 0.001 versus vehicle (one-way ANOVA Dunnett’s multiple comparison test). CMC: carboxymethyl cellulose; S.E.M.: standard error of the mean; DPP-IV: dipeptidyl peptidase 4.
Using low doses of NCP-322 (0.1–3 mg/kg), we performed oral glucose tolerance test (OGTT) and observed that NCP-322 oral administration significantly reduced glycemic excursions following oral glucose challenge in mice (Fig. 5A), resulting in a significantly decreased AUC at 120 min following the glucose challenge (Fig. 5C). MBX-2982 (3 mg/kg) oral administration significantly reduced glycemic excursions (Supplementary Fig. 2A) and dose-dependently decreased the AUC at 120 min following the glucose challenge (Supplementary Fig. 2C). Glibenclamide (1 mg/kg, orally) markedly reduced glycemic excursions; however, compared with the control group, it significantly reduced plasma glucose levels at 0 min (Fig. 5A, Supplementary Fig. 2A). At high doses of NCP-322 (3–30 mg/kg), its oral administration significantly reduced glycemic excursions (Fig. 5B) and resulted in a significantly decreased AUC (3–30 mg/kg) at 120 min following glucose ingestion (Fig. 5D). Moreover, MBX-2982 oral administration at high doses (30 mg/kg) markedly reduced the plasma glucose levels at 30 min following the oral glucose challenge but significantly increased the plasma glucose levels at 120 min following the oral glucose challenge (Supplementary Fig. 2B). Even at high doses, both NCP-322 and MBX-2982 did not induce hypoglycemia. Furthermore, to evaluate the hypoglycemic risk of NCP-322, the following study aimed to confirm that NCP-322 does not exhibit a glucose-lowering effect when blood glucose levels are low by monitoring the changes in plasma glucose levels at 150 min following a single treatment in fasted mice. NCP-322 (3 mg/kg) single administration caused similar changes in the plasma glucose levels to those of the vehicle group; however, glibenclamide (1 and 3 mg/kg) caused a significant reduction from 30 to 150 min after administration (Fig. 5E). These results also indicate that NCP-322 does not have a risk of hypoglycemia in vivo, at least in the doses tested.
Vehicle (1% CMC containing 2% Tween 80) or NCP-322 is administered to C57BL/6J mice under fasting conditions. Blood samples are collected from the tail vein 30 min before (defined as -30 min value), immediately before (0 min), and 60 and 120 min after the challenge of glucose or water (control group). Time course of the plasma glucose concentration at low (A) or high (B) NCP-322 doses. The area under the curve (AUC) at 120 min following the glucose challenge is calculated (C, low doses; D, high doses of NCP-322). Vehicle, glibenclamide (Gliben.), or NCP-322 is administered to C57BL/6J mice under fasting conditions. Blood samples are collected from the tail vein immediately before (0 min) and at 30, 60, 90, and 150 min following test compound administration. Time course of the plasma glucose concentration on the vehicle, NCP-322, or glibenclamide (E). Data are presented as means ± S.E.M. (A–D) n = 6–10 mice; #p < 0.05 versus control. * p < 0.05, ** p < 0.01, and *** p < 0.001 versus vehicle (one-way ANOVA Dunnett’s multiple comparison test). (E) n = 7–8 mice; ** p < 0.01, *** p < 0.001 versus vehicle (one-way ANOVA Dunnett’s multiple comparison test). CMC: carboxymethyl cellulose; S.E.M.: standard error of the mean.
We performed GLP-1 and insulin pharmacodynamic analysis in mice to investigate whether the improved glucose tolerance was related to the enhanced glucose-dependent insulin and GLP-1 secretion. In the vehicle-treated group, the plasma GLP-1 and insulin levels increased following the glucose challenge and peaked at 10 and 20 min, respectively. NCP-322 (1 mg/kg) oral administration significantly increased the plasma GLP-1 levels at 0, 5, and 10 min following the glucose challenge. Following the increased GLP-1 level, the plasma insulin level significantly increased compared with those of the vehicle-treated group at 20 min following the glucose challenge (Fig. 6). NCP-322 (1 mg/kg) significantly reduced the glucose levels at 40 min following the oral glucose challenge. These results indicate that the effects of NCP-322 on glycemic control involve glucose-dependent insulin secretion enhancement following the potent, long-lasting elevation of the GLP-1 level following NCP-322 oral administration.
Vehicle (1% CMC containing 2% Tween 80) or NCP-322 is administered to C57BL/6J mice under fasting conditions. After 30 min, glucose (3 g/kg, orally) is challenged. Blood is collected from the abdominal vein at 0 min (just before the glucose challenge) and 5, 10, 20, 30, and 40 min following the glucose challenge. For GLP-1 determination, portal vein blood is collected in tubes containing the DPP-IV inhibitor at the same time points. Data are presented as means ± S.E.M. (n = 9–13 mice). * p < 0.05, ** p < 0.01, N.D.: not detected versus vehicle (Student’s t-test). GLP-1: glucagon-like peptide 1; CMC: carboxymethyl cellulose; S.E.M.: standard error of the mean; DPP-IV: dipeptidyl peptidase 4.
To date, drugs for T2DM treatment with different mechanisms have been developed; however, significant unmet medical needs, including sustained glycemic control over a long period and side effects, remain. Here we identified the newly synthesized small molecule GPR119 agonist, NCP-322, which has a therapeutic potential for T2DM and overcomes the drawbacks caused by treatment using previously developed GPR119 agonists.
Gs protein-coupled GPCR has an agonist-stimulated effect on increasing intracellular cAMP levels and may mediate glucose-dependent insulin secretion, incretin secretion from the gastrointestinal tract, and food intake and weight gain suppression.21,23–25) GPR119 is coupled with Gs protein and is expressed in the pancreatic β-cells, gastrointestinal tract, and brain.19,20) In a rat model, GPR119 agonists have been shown to increase intracellular cAMP levels, glucose-dependent insulin secretion, food intake, and body weight,21,23) as well as stimulate GLP-1 secretion,24,25) attracting attention as a therapeutic target for T2DM. This study demonstrated that the newly developed NCP-322 exhibited a potent GPR119 agonistic activity. Off-target tests confirmed its high selectivity, wherein binding assays for several receptors, ion channels, and transporters (76 types), enzymes (76 types), and cell functions (4 types) were evaluated. Although NCP-322 can bind some receptors and inhibit the activity of enzymes (Supplementary Table 1), almost all of these effects were weak (approximately 50%) even at high concentrations (10μmol/L). In fact, the IC50 values of these off-target effects were approximately 1000-fold higher than the EC50 values of NCP-322 for activating GPR119. We further demonstrated that NCP-322 exhibited a concentration-dependent glucose-dependent insulin secretion effect in murine-isolated pancreatic β-cells and, from a low dose of 0.3 mg/kg, a dose-dependent GLP-1 secretagogue effect in vivo. Furthermore, in an OGTT in mice, reduced glycemic excursions were demonstrated from a low dose of 0.1 mg/kg. Therefore, the pharmacological effects of NCP-322 obtained from the in vitro and in vivo experiments in this study are believed to be mediated by GPR119 but not by other off-target effects observed in high NCP-322 concentrations.
Furthermore, we showed that the potency and efficacy of NCP-322 for activating GPR119 were equivalent to those of the previously developed GPR119 agonists (MBX-2982 and AR231453). Regarding mouse GPR119, NCP-322 had a 31% higher Emax value than MBX-2982. The high Emax values reflected the amount of cAMP produced, which was associated with a higher pharmacological response. As previously reported in a case of DS-8500, a GPR119 agonist that has a higher Emax value to increase intracellular cAMP levels and has a therapeutic effect of more potent glucose-lowering effects in diabetic models,29) the pharmacological effects of NCP-322 were more potent than those of MBX-2982. Another notable feature of NCP-322 was its few side effects. The most common side effect when using therapeutic drugs in diabetes treatment is hypoglycemia. Symptoms include cold sweats, facial pallor, and palpitations; however, a continued hypoglycemic state may cause more serious symptoms, including disturbance of consciousness and convulsions. In insulin secretion evaluation using pancreatic β-cells, the sulfonylurea, glibenclamide, exhibited an insulin secretory effect even at low glucose concentrations, whereas NCP-322 showed no effect. Additionally, when orally administered to mice following an overnight fast, glibenclamide demonstrated a hypoglycemic effect, whereas NCP-322 showed a blood glucose profile similar to that of the vehicle group. These findings indicate that NCP-322 does not have the ability to induce hypoglycemia, at least at the doses tested. In this study, MBX-2982 did not reduce plasma glucose levels before the glucose challenge in the OGTT, similar to NCP-322. Additionally, AS1535907 (a GPR119 agonist previously developed by Astellas Pharma Inc.) reportedly does not promote insulin secretion under low glucose conditions but enhances glucose-dependent insulin secretion only under high glucose conditions.30) Based on these findings, we believe that the characteristic of fewer hypoglycemic side effects is not unique to NCP-322 but is a feature of GPR119 agonists. However, a recent report suggested that GPR119 agonists suppress hypoglycemia by promoting glucagon secretion from islet a-cells only during hypoglycemia.31) Although that study did not examine concentration dependence, it confirmed the specificity of the action against GPR119 in KO mice. Therefore, it is presumed that the inhibition of hypoglycemia induction depends on the GPR119 agonistic activity and pharmacokinetic profile. In addition, GPR119 agonists, which promote GLP-1 secretion, are expected to enhance glucose-lowering effects when combined with DPP-IV inhibitors, which prevent GLP-1 degradation. In fact, we have confirmed in a previous study that the combination of compound 9j (a GPR119 agonist previously developed by Arena32)) and sitagliptin shows synergistic hypoglycemic effects (data not shown). On the other hand, many diabetes medications may increase the risk of hypoglycemia when used in combination with these drugs compared to their use as single agents.31) However, as mentioned above, GPR119 agonists have been suggested to suppress hypoglycemia, and the combination of a GPR119 agonist and a DPP-IV inhibitor may be a promising treatment option, reducing the risk of hypoglycemia when used together.
In conclusion, this study demonstrated that our newly developed compound, NCP-322, has abilities to promote insulin and GLP-1 secretion at low doses without causing noticeable side effects, including hypoglycemia, which have been the drawbacks caused by previous therapeutic drugs; furthermore, this study showed beneficial effects of NCP-322 for T2DM treatment. To date, 4 GPR119 agonists (GSK1292263A, MBX-2982, PSN821, and DS-8500) have progressed to Phase II clinical trials; however, the development of GSK1292263A had been abandoned owing to the attenuated drug efficacy with repeated dosing observed during the clinical trial. By contrast, PSN821 and DS-8500 have yielded favorable results.33–37) Therefore, determining whether the therapeutic effects of NCP-322 are maintained by repeated dosing using disease models will be a significant subject for future research.
The authors are grateful to Noriko Kanakubo for compound supply, Toru Ogawa, Sawako Kanda, and Mai Ito for their expertly conducted experiments, and Sachiko Ebata and Kazuaki Shimada for their animal care.
This work was supported by Nippon Chemiphar Co., Ltd., Tokyo, Japan.
Conceptualization, H.N.; Methodology, H.N.; Software, H.N.; Validation, H.N.; Formal analysis, H.N.; Investigation, H.N. and T.E.; Resources, H.N.; Data curation, H.N.; Writing—original draft preparation, H.N.; Writing—review and editing, H.N. and M.T.; Visualization, H.N.; Supervision, H.N.; Project administration, H.N.; Funding acquisition, H.N. All authors have read and agreed to the published version of the manuscript.
This work was funded by Nippon Chemiphar Co., Ltd. H.N. and T.E. are employees of Nippon Chemiphar Co., Ltd. M.T. has no conflict of interest.
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