Discovery of potent, selective, and orally bioavailable quinoline-based dipeptidyl peptidase IV inhibitors targeting Lys554

Dipeptidyl peptidase IV (DPP-4) inhibition is a validated therapeutic option for type 2 diabetes, exhibiting multiple antidiabetic effects with little or no risk of hypoglycemia. In our studies involving non-covalent DPP-4 inhibitors, a novel series of quinoline-based inhibitors were designed based on the co-crystal structure of isoquinolone 2 in complex with DPP-4 to target the side chain of Lys554. Synthesis and evaluation of designed compounds revealed 1-[3-(aminomethyl)-4-(4-methylphenyl)-2-(2-methylpropyl)quinolin-6-yl]piperazine-2,5-dione (1) as a potent, selective, and orally active DPP-4 inhibitor (IC??=1.3 nM) with long-lasting ex vivo activity in dogs and excellent antihyperglycemic effects in rats. A docking study of compound 1 revealed a hydrogen-bonding interaction with the side chain of Lys554, suggesting this residue as a potential target site useful for enhancing DPP-4 inhibition.     

1-((3S,4S)-4-amino-1-(4-substituted-1,3,5-triazin-2-yl) pyrrolidin-3-yl)-5,5-difluoropiperidin-2-one inhibitors of DPP-4 for the treatment of type 2 diabetes

A 3-amino-4-substituted pyrrolidine series of dipeptidyl peptidase IV (DPP-4) inhibitors was rapidly developed into a candidate series by identification of a polar valerolactam replacement for the lipophilic 2,4,5-trifluorophenyl pharmacophore. The addition of a gem-difluoro substituent to the lactam improved overall DPP-4 inhibition and an efficient asymmetric route to 3,4-diaminopyrrolidines was developed. Advanced profiling of a subset of analogs identified 5o with an acceptable human DPP-4 inhibition profile based on a rat PK/PD model and a projected human dose that was suitable for clinical development.     

New dipeptidyl peptidase 4 inhibitors among adamantane derivatives

Fifteen adamantane derivatives were synthesized. Preliminary evaluation of their potential as dipeptidyl peptidase 4 (DPP-4) inhibitors was performed in silico by the Microcosm informational technology, PASS system, and docking in AutoDock Vina. The DPP-4 inhibition was studied in vitro. The selectivity of action of the most active compounds was studied by the direct inhibition of human plasma DPP-4 and recombinant human DPP-8. The highest activity was found for the compounds containing a nitrogen atom in the β-position of the side chain, namely, derivatives of adamantane carboxylic acid and N-(3-adamantyl-allyl) thiourea. We demonstrated that the most active compound of the series, 3,5-dimethyladamantane 1-carboxamide, was a selective DPP-4 inhibitor with IC₅₀ 53.94 μM.     

Discovery of DA-1229: a potent, long acting dipeptidyl peptidase-4 inhibitor for the treatment of type 2 diabetes

A series of β-amino amide containing substituted piperazine-2-one derivatives was synthesized and evaluated as inhibitors of dipeptidyl pepdidase-4 (DPP-4) for the treatment of type 2 diabetes. As results of intensive SAR study of the series, (R)-4-[(R)-3-amino-4-(2,4,5-trifluorophenyl)-butanoyl]-3-(t-butoxymethyl)-piperazin-2-one (DA-1229) displayed potent DPP-4 inhibition pattern in several animal models, was selected for clinical development.     

Synthesis and biological evaluation of azobicyclo[3.3.0] octane derivatives as dipeptidyl peptidase 4 inhibitors for the treatment of type 2 diabetes

A series of novel azobicyclo[3.3.0]octane derivatives were synthesized and evaluated as dipeptidyl peptidase 4 (DPP-4) inhibitors. The effort resulted in the discovery of inhibitor 2a, which exhibited excellent efficacies in an oral glucose tolerance test. Introduction of methyl group (2j) could prolong the inhibition of serum DPP-4 activity.     

4-arylcyclohexylalanine analogs as potent, selective, and orally active inhibitors of dipeptidyl peptidase IV

A novel series of 4-arylcyclohexylalanine DPP-4 inhibitors was synthesized and tested for inhibitory activity as well as selectivity over the related proline-specific enzymes DPP-8 and DPP-9. Optimization of this series led to 28 (DPP-4 IC(50)=4.8 nM), which showed an excellent pharmacokinetic profile across several preclinical species. Evaluation of 28 in an oral glucose tolerance test demonstrated that this compound effectively reduced glucose excursion in lean mice.     

Synthesis and evaluation of [(1R)-1-amino-2-(2,5-difluorophenyl)ethyl]cyclohexanes and 4-[(1R)-1-amino-2-(2,5-difluorophenyl)ethyl]piperidines as DPP-4 inhibitors

A series of 4-amino cyclohexanes and 4-substituted piperidines were prepared and evaluated for inhibition of DPP-4. Analog 20q displayed both good DPP-4 potency and selectivity against other proteases, while derivative 20k displayed long half life and modest oral bioavailability in rat. The most potent analog, 3-(5-aminocarbonylpyridyl piperidine 53j, displayed excellent DPP-4 activity with good selectivity versus other proline enzymes.     

Synthesis and biological evaluation of novel benzyl-substituted (S)-phenylalanine derivatives as potent dipeptidyl peptidase 4 inhibitors

A series of novel benzyl-substituted (S)-phenylalanine derivatives were synthesized and evaluated for their dipeptidyl peptidase 4 (DPP-4) inhibitory activity and selectivity. It was found that most synthesized target compounds were potent DPP-4 inhibitors with IC₅₀ values in 3.79–25.52 nM, which were significantly superior to that of the marketed drug sitagliptin. Furthermore, the 4-fluorobenzyl substituted phenylalanine derivative 6g not only displayed the potent DPP-4 inhibition with an IC₅₀ value of 3.79 nM, but also showed better selectivity against DPP-4 over other related enzymes including DPP-7, DPP-8, and DPP-9. In an oral glucose tolerance test (OGTT) in normal Sprague Dawley rats, compound 6g reduced blood glucose excursion in a dose-dependent manner.     

Effects of novel chalcone derivatives on α-glucosidase, dipeptidyl peptidase-4, and adipocyte differentiation in vitro

Chana series are new chalcone derivatives. To evaluate the possibility of Chana series as therapeutic agents of type 2 diabetes, the inhibitory effects of Chana series on the activities of α-glucosidase and DPP-4 were investigated using in vitro enzyme assays, and their effects on adipocyte differentiation were investigated in C3H10T1/2 cells. Chana 1 and Chana 7 among the Chana series showed significant inhibition of α-glucosidase activity. In DPP-4 enzyme assay, Chana 1 exhibited the highest inhibitory activity while Chana 7 did not. In MTT assay, Chana 1 did not show significant cytotoxicity up to a concentration of 250 μM, whereas cytotoxicity was observed with Chana 7 at a concentration of 300 μM. In addition, Chana 1 induced adipocyte differentiation. Therefore, Chana 1 showed inhibitory effects on α-glucosidase and DPP-4 as well as a stimulatory effect on adipocyte differentiation, suggesting that Chana 1 may be a potential beneficial agent for the treatment of type 2 diabetes.     

Medicinal chemistry approaches to the inhibition of dipeptidyl peptidase-4 for the treatment of type 2 diabetes

Emerging as an epidemic of the 21st century type 2 diabetes has become a major health problem throughout the globe. The number of deaths attributable to diabetes reflects the insufficient glycemic control achieved with the treatments used in recent past. DPP-4 inhibitors have been investigated as a new therapy with novel mechanisms of action and improved tolerability. DPP-4, a protease that specifically cleaves dipeptides from proteins and oligopeptides after a penultimate N-terminal proline or alanine, is involved in the degradation of a number of neuropeptides, peptide hormones and cytokines, including the incretins GLP-1 and GIP. As soon as released from the gut in response to food intake, GLP-1 and GIP exert a potent glucose-dependent insulinotropic action, thereby playing a key role in the maintenance of post-meal glycemic control. Consequently, inhibiting DPP-4 prolongs the action of GLP-1 and GIP, which in turn improves glucose homeostasis with a low risk of hypoglycemia and potential for disease modification. Indeed, clinical trials involving diabetic patients have shown improved glucose control by administering DPP-4 inhibitors, thus demonstrating the benefit of this promising new class of antidiabetics. Intense research activities in this area have resulted in the launch of sitagliptin and vildagliptin (in Europe only) and the advancement of a few others into preregistration/phase 3, for example, saxagliptin, alogliptin and ABT-279. Achieving desired selectivity for DPP-4 over other related peptidases such as DPP-8 and DPP-9 (inhibition of which was linked to toxicity in animal studies) and long-acting potential for maximal efficacy (particularly in more severe diabetic patients) were the major challenges. Whether these goals are achieved with the present series of inhibitors in the advanced stages of clinical development is yet to be confirmed. Nevertheless, treatment of this metabolic disorder especially in the early stages of the disease via DPP-4 inhibition has been recognized as a validated principle and a large number of inhibitors are presently in various stage of pre-clinical/clinical development. Sitagliptin is a new weapon in the arsenal of oral antihyperglycemic agents. This review will focus on the journey of drug discovery of DPP-4 inhibitors for oral delivery covering a brief scientific background and medicinal chemistry approaches along with the status of advanced clinical candidates.     

4-aminophenylalanine and 4-aminocyclohexylalanine derivatives as potent, selective, and orally bioavailable inhibitors of dipeptidyl peptidase IV

A novel series of 4-aminophenylalanine and 4-aminocyclohexylalanine derivatives were designed and evaluated as inhibitors of dipeptidyl peptidase IV (DPP-4). The phenylalanine series afforded compounds such as 10 that were potent and selective (DPP-4, IC(50)=28nM), but exhibited limited oral bioavailability. The corresponding cyclohexylalanine derivatives such as 25 afforded improved PK exposure and efficacy in a murine OGTT experiment. The X-ray crystal structure of 25 bound to the DPP-4 active site is presented.     

Macrocarpal C isolated from Eucalyptus globulus inhibits dipeptidyl peptidase 4 in an aggregated form

Dipeptidyl peptidase 4 (DPP-4) inhibitors are used for the treatment of type-2 diabetes mellitus. Various synthetic inhibitors have been developed to date, and plants containing natural DPP-4 inhibitors have also been identified. Here, 13 plant samples were tested for their DPP-4 inhibitory activity. Macrocarpals A–C were isolated from Eucalyptus globulus through activity-guided fractionation and shown to be DPP-4 inhibitors. Of these, macrocarpal C showed the highest inhibitory activity, demonstrating an inhibition curve characterised by a pronounced increase in activity within a narrow concentration range. Evaluation of macrocarpal C solution by turbidity, nuclear magnetic resonance spectroscopy and mass spectrometry indicated its aggregation, which may explain the characteristics of the inhibition curve. These findings will be valuable for further study of potential small molecule DPP-4 inhibitors.     

DPP-4 inhibition improves glucose tolerance and increases insulin and GLP-1 responses to gastric glucose in association with normalized islet topography in mice with beta-cell-specific overexpression of human islet amyloid polypeptide

Inhibition of dipeptidyl peptidase-4 (DPP-4) is currently explored as a novel therapy of type 2 diabetes. The strategy has been shown to improve glycemia in most, but not all, rodent forms of glucose intolerance. In this study, we explored the effects of DPP-4 inhibition in mice with beta-cell overexpression of human islet amyloid polypeptide (IAPP). We therefore administered the orally active and highly selective DPP-4 inhibitor, vildagliptin (3 micromol/mouse daily) to female mice with beta-cell overexpression of human IAPP. Controls were given plain water, and a series of untreated wildtype mice was also included. After five weeks, an intravenous glucose tolerance test showed improved glucose disposal and a markedly enhanced insulin response in mice treated with vildagliptin. After eight weeks, a gastric tolerance test showed that vildagliptin improved glucose tolerance and markedly (approximately ten-fold) augmented the insulin response in association with augmented (approximately five-fold) levels of intact glucagon-like peptide-1 (GLP-1). Furthermore, after nine weeks, islets were isolated. Islets from vildagliptin-treated mice showed augmented glucose-stimulated insulin response and a normalization of the islet insulin content, which was reduced by approximately 50% in transgenic controls versus wildtype animals. Double immunostaining of pancreatic islets for insulin and glucagon revealed that transgenic islets displayed severely disturbed intra-islet topography with frequently observed centrally located alpha-cells. Treatment with vildagliptin restored the islet topography. We therefore conclude that DPP-4 inhibition improves islet function and islet topography in mice with beta-cell specific transgenic overexpression of human IAPP.     

Natural compounds with DPP-4 inhibitory effects: Implications for the treatment of diabetes

Dipeptidyl peptidase-4 (DPP-4) inhibitors are antidiabetes agents that decrease blood glucose by preventing the degradation of endogenous glucagon-like peptide-1. The first DPP-4 was sitagliptin followed by several other agents in the class introduced to manage diabetes. Recent studies have suggested that naturally occurring compounds can exert an antidiabetes effect through DPP-4 inhibition. Such compounds may have a place in the treatment of diabetes within the diet; however, while DPP-4 inhibition alone is not associated with hypoglycemia, in combinations with other medication hypoglycemia can result, therefore, it is critical to know what herbal or food-based compounds may have these activities in the management of diabetes patients. In this review, we have outlined the compounds that have DPP-4 inhibition that may have utility in the treatment of diabetes.     

Synthesis and biological evaluation of pyrrolidine-2-carbonitrile and 4-fluoropyrrolidine-2-carbonitrile derivatives as dipeptidyl peptidase-4 inhibitors for the treatment of type 2 diabetes

A novel series of pyrrolidine-2-carbonitrile and 4-fluoropyrrolidine-2-carbonitrile derivatives was designed, synthesized, and found to act as dipeptidyl peptidase-4 (DPP-4) inhibitors. From this series of compounds, compound 17a was identified as an efficacious, safe, and selective inhibitor of DPP-4. In vivo studies in ICR and KKAy mice showed that administration of this compound resulted in decreased blood glucose in these mice after an oral glucose challenge. Compound 17a showed high DPP-4 inhibitory activity (IC₅₀ = 0.017 μM), moderate selectivity against DPP-4 (selective ratio: DPP-8/DPP-4 = 1324; DPP-9/DPP-4 = 1164), and good efficacy in oral glucose tolerance tests in ICR and KKAy mice. These in vivo anti-diabetic properties and its desirable pharmacokinetic profile in Sprague–Dawley rats demonstrate that compound 17a is a promising candidate for development as an anti-diabetic agent.     

Discovery of triazole-based uracil derivatives bearing amide moieties as novel dipeptidyl peptidase-IV inhibitors

Dipeptidyl peptidase-IV (DPP-4) is a validated target for T2DM treatment. We previously reported a novel series of triazole-based uracil derivatives bearing aliphatic carboxylic acids with potent DPP-4 inhibitory activities in vitro, but these compounds showed poor hypoglycemic effects in vivo. Herein we further optimized the triazole moiety by amidation of the carboxylic acid to improve in vivo activities. Two series of compounds 3a-f and 4a-g were designed and synthesized. By screening in DPP-4, compound 4c was identified as a potent DPP-4 inhibitor with the IC₅₀ value of 28.62 nM. Docking study revealed compound 4c has a favorable binding mode and interpreted the SAR of these analogs. DPP-8 and DPP-9 tests indicated compound 4c had excellent selectivity over DPP-8 and DPP-9. Further in vivo evaluations revealed that compound 4c showed more potent hypoglycemic activity than its corresponding carboxylic acid in ICR mice and dose-dependently reduced glucose levels in type 2 diabetic C57BL/6 mice. The overall results have shown that compound 4c could be a promising lead for further development of novel DPP-4 agents treating T2DM.     

Novel tetrahydropyran analogs as dipeptidyl peptidase IV inhibitors: Profile of clinical candidate (2R,3S,5R)-2-(2,5-difluorophenyl)-5-[2-(methylsulfonyl)-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl]tetrahydro-2H-pyran-3-amine (23)

A series of novel tri-2,3,5-substituted tetrahydropyran analogs were synthesized and evaluated as inhibitors of dipeptidyl peptidase IV (DPP-4) for the treatment of type 2 diabetes. Optimization of the series provided inhibitors with good DPP-4 potency and selectivity over other peptidases (QPP, DPP8, and FAP). Compound 23, which is very potent, selective, efficacious in the diabetes PD model, and has an excellent pharmacokinetic profile, is selected as a clinical candidate.     

(R)-3-Amino-1-((3aS,7aS)-octahydro-1H-indol-1-yl)-4-(2,4,5-trifluorophenyl)butan-1-one derivatives as potent inhibitors of dipeptidyl peptidase-4: Design,synthesis, biological evaluation,and molecular modeling

A series of (R)-3-amino-1-((3aS,7aS)-octahydro-1H-indol-1-yl)-4-(2,4,5-trifluorophenyl)butan-1-one derivatives was designed, synthesized, and evaluated as novel inhibitors of dipeptidyl peptidase-4 (DPP-4) for the treatment of type 2 diabetes. Most of the synthesized compounds demonstrated good inhibition activities against DPP-4. Among these, compounds 3e, 4c, 4l, and 4n exhibited prominent inhibition activities against DPP-4, with IC₅₀s of 0.07, 0.07, 0.14, and 0.17 μM, respectively. The possible binding modes of compounds 3e and 4n with dipeptidyl peptidase-4 were also explored by molecular docking simulation. These potent DPP-4 inhibitors were optimized for the absorption, distribution, metabolism, and excretion (ADME) properties, and compound 4n displayed an attractive pharmacokinetic profile (F = 96.3%, t_1/2 = 10.5 h).     

Discovery of new binding elements in DPP-4 inhibition and their applications in novel DPP-4 inhibitor design

Probing with tool molecules, and by modeling and X-ray crystallography the binding modes of two structurally distinct series of DPP-4 inhibitors led to the discovery of a rare aromatic fluorine H-bond and the spatial requirement for better biaryl binding in the DPP-4 enzyme active site. These newly found binding elements were successfully incorporated into novel DPP-4 inhibitors.     

Fluoroolefins as amide bond mimics in dipeptidyl peptidase IV inhibitors

The synthesis, selectivity, rat pharmacokinetic profile, and drug metabolism profiles of a series of potent fluoroolefin-derived DPP-4 inhibitors (4) are reported. A radiolabeled fluoroolefin 33 was shown to possess a high propensity to form reactive metabolites, thus revealing a potential liability for this class of DPP-4 inhibitors.