Coupling of an Acyl Migration Prodrug Strategy with Bio-activation To Improve Oral Delivery of the HIV-1 Protease Inhibitor Atazanavir.

HIV-1 protease inhibitors (PIs), which include atazanavir (ATV, 1), remain important medicines to treat HIV-1 infection. However, they are characterized by poor oral bioavailability and a need for boosting with a pharmacokinetic enhancer, which results in additional drug-drug interactions that are sometimes difficult to manage. We investigated a chemo-activated, acyl migration-based prodrug design approach to improve the pharmacokinetic profile of 1 but failed to obtain improved oral bioavailability over dosing the parent drug in rats.

Applications of 2, 2, 2 trifluoroethanol as a versatile co-solvent in supercritical fluid chromatography for purification of unstable boronate esters, enhancing throughput, reducing epimerization, and for additive free purifications.

Analysis and purification of boronic acid pinacol esters by RPLC is very challenging due to their degradation in aqueous and alcoholic solvents. These compounds are difficult to purify by SFC too as they are equally sensitive to traditional co-solvents like methanol, ethanol, and 2-propanol. 2,2,2 trifluoroethanol (TFE), which has been reported for the purification of a few alcohol sensitive compounds, was evaluated as a co-solvent in this study for the purification of chiral and achiral boronate esters by SFC.

Impact of CO-solvent separators on the degradation of benzyl-2,3-dihydroxypiperidine-1-carboxylate during preparative supercritical fluid chromatographic (SFC) purification.

During a preparative separation of the cis enantiomeric pair of benzyl-2,3-dihydroxypiperidine-1-carboxylate using supercritical-fluid chromatography (SFC) with methanol modifier, significant degradation of the products in the collected fractions was observed when a Waters SFC-350 (Milford, MA, USA) was used, but same was not observed when a Waters SFC-80q (Milford, MA, USA) was used. Through a systematic investigation, we discovered that the compound degraded over time under an acidic condition created by the formation of methyl carbonic acid from methanol and CO The extent of the product degradation was dependent on the time and the concentration of CO remained in the product fraction, which was governed by the efficiency of CO-methanol separation during the fraction collection. Hence, we demonstrated that the different designs of CO-solvent separator (high pressurized cyclone in Waters SFC-350 and low-pressurized vortexing separator in Waters SFC-80q) had a significant impact on the degradation of an acid-sensitive compound.

Use of Hybrid Capillary Tube Apparatus on 400 MHz NMR for Quantitation of Crucial Low-Quantity Metabolites Using aSICCO Signal.

Metabolites of new chemical entities can influence safety and efficacy of a molecule and often times need to be quantified in preclinical studies. However, synthetic standards of metabolites are very rarely available in early discovery. Alternate approaches such as biosynthesis need to be explored to generate these metabolites.

Integrating a post-column makeup pump into preparative supercritical fluid chromatography systems to address stability and recovery issues during purifications.

Purification of many pharmaceutical compounds by supercritical fluid chromatography (SFC) has always been challenging because of degradation of compound during the isolation step in the presence of acidic or basic modifiers in the mobile phase. Stability of such acid or base-sensitive compounds could be improved by post-column addition of a solvent containing base or acid modifier as counter ion through a make-up pump respectively to neutralize the compound fraction without affecting the resolution. One such case study has been presented in this work where the stability of a base-sensitive compound was addressed by the addition of acidic co-solvent through the make-up pump.

Identification of RO4597014, a Glucokinase Activator Studied in the Clinic for the Treatment of Type 2 Diabetes.

To resolve the metabolite redox cycling associated with our earlier clinical compound 2, we carried out lead optimization of lead molecule 1. Compound 4 showed improved lipophilic ligand efficiency and demonstrated robust glucose lowering in diet-induced obese mice without a liability in predictive preclinical drug safety studies. Thus, it was selected as a clinical candidate and further studied in type 2 diabetic patients.

Repair of diverse diabetic defects of β-cells in man and mouse by pharmacological glucokinase activation.

Glucokinase activators (GKAs) are being developed and clinically tested for potential antidiabetic therapy. The potential benefits and limitations of this approach continue to be intensively debated. To contribute to the understanding of experimental pharmacology and therapeutics of GKAs, we have tested the efficacy of one of these agents (Piragliatin) in isolated islets from humans with type 2 diabetes mellitus (T2DM), from mice with glucokinase (GK) mutations induced by ethyl-nitroso-urea (ENU) as models of Maturity Onset Diabetes of the Young linked to GK and Permanent Neonatal Diabetes Mellitus linked to GK (PNDM-GK) and finally of islets rendered glucose insensitive by treatment with the sulphonyl urea compound glyburide in organ culture.

Discovery of piragliatin--first glucokinase activator studied in type 2 diabetic patients.

Glucokinase (GK) activation as a potential strategy to treat type 2 diabetes (T2D) is well recognized. Compound 1, a glucokinase activator (GKA) lead that we have previously disclosed, caused reversible hepatic lipidosis in repeat-dose toxicology studies. We hypothesized that the hepatic lipidosis was due to the structure-based toxicity and later established that it was due to the formation of a thiourea metabolite, 2.

Thermal stability of glucokinase (GK) as influenced by the substrate glucose, an allosteric glucokinase activator drug (GKA) and the osmolytes glycerol and urea.

We investigated how glycerol, urea, glucose and a GKA influence kinetics and stability of wild-type and mutant GK. Glycerol and glucose stabilized GK additively. Glycerol barely affected the TF spectra of all GKs but decreased k(cat), glucose S(0.

Glucokinase activation repairs defective bioenergetics of islets of Langerhans isolated from type 2 diabetics.

It was reported previously that isolated human islets from individuals with type 2 diabetes mellitus (T2DM) show reduced glucose-stimulated insulin release. To assess the possibility that impaired bioenergetics may contribute to this defect, glucose-stimulated respiration (Vo(2)), glucose usage and oxidation, intracellular Ca(2+), and insulin secretion (IS) were measured in pancreatic islets isolated from three healthy and three type 2 diabetic organ donors. Isolated mouse and rat islets were studied for comparison.

Mutational analysis of allosteric activation and inhibition of glucokinase.

GK (glucokinase) is activated by glucose binding to its substrate site, is inhibited by GKRP (GK regulatory protein) and stimulated by GKAs (GK activator drugs). To explore further the mechanisms of these processes we studied pure recombinant human GK (normal enzyme and a selection of 31 mutants) using steady-state kinetics of the enzyme and TF (tryptophan fluorescence). TF studies of the normal binary GK-glucose complex corroborate recent crystallography studies showing that it exists in a closed conformation greatly different from the open conformation of the ligand-free structure, but indistinguishable from the ternary GK-glucose-GKA complex.

Research and development of glucokinase activators for diabetes therapy: theoretical and practical aspects.

Glucokinase Glucokinase (GK GK ; EC 2.7.1.

Discovery of benzothiazole-based adenosine A2B receptor antagonists with improved A2A selectivity.

The highly potent but modestly selective N-(2-amino-4-methoxy-benzothiazol-7-yl)-N-ethyl-acetamide derivative 2 was selected as the starting point for the design of novel selective A(2B) antagonists, due to its excellent potency, and good drug-like properties. A series of compounds containing nonaromatic amides or ureas of five- or six-membered rings, and also bearing an m-trifluoromethyl-phenyl group (shown to impart superior potency) was prepared and evaluated for their selectivity against the A(2A) and A(1) receptors. This work resulted in the identification of compound 30, with excellent potency and high selectivity against both A(2A) and A(1) receptors.

Novel glucokinase activators: a patent review (2008 - 2010).

Importance Of The Field: Small molecule glucokinase activators (GKAs) continue to represent a potential strategy to treat type 2 diabetes (T2D). Glucokinase (GK) primarily exerts its effect through modulatory actions in pancreatic β-cells and hepatocytes. It couples insulin secretion in the pancreas with plasma glucose concentration and improves glucose utilization in the liver, thus, affecting two key aspects of glucose homeostasis.

2,3-Disubstituted acrylamides as potent glucokinase activators.

The phenylacetamide 1 represents the archtypical glucokinase activator (GKA) in which only the R-isomer is active. In order to probe whether the chiral center could be replaced, we prepared a series of olefins 2 and show in the present work that these compounds represent a new class of GKAs. Surprisingly, the SAR of the new series paralleled that of the saturated derivatives with the exception that there was greater tolerance for larger alkyl and cycloalkyl groups at R(2) region in comparison to the phenylacetamides.

4-Substituted-7-N-alkyl-N-acetyl 2-aminobenzothiazole amides: drug-like and non-xanthine based A2B adenosine receptor antagonists.

7-N-Acetamide-4-methoxy-2-aminobenzothiazole 4-fluorobenzamide (compound 1) was chosen as a drug-like and non-xanthine based starting point for the discovery of A(2B) receptor antagonists because of its slight selectivity against A(1) and A(2A) receptors and modest A(2B) potency. SAR exploration of compound 1 described herein included modifications to the 7-N-acetamide group, substitution of the 4-methoxy group by halogens as well as replacement of the p-flouro-benzamide side chain. This work culminated in the identification of compound 37 with excellent A(2B) potency, modest selectivity versus A(2A) and A(1) receptors, and good rodent PK properties.

Discovery, structure-activity relationships, pharmacokinetics, and efficacy of glucokinase activator (2R)-3-cyclopentyl-2-(4-methanesulfonylphenyl)-N-thiazol-2-yl-propionamide (RO0281675).

Glucokinase (GK) is a glucose sensor that couples glucose metabolism to insulin release. The important role of GK in maintaining glucose homeostasis is illustrated in patients with GK mutations. In this publication, identification of the hit molecule 1 and its SAR development, which led to the discovery of potent allosteric GK activators 9a and 21a, is described.

Glucokinase activators for the potential treatment of type 2 diabetes.

The search for innovative and clinically-differentiated medicines for the treatment of type 2 diabetes is an active area of research for pharmaceutical companies. The discovery of allosteric Glucokinase (GK) activators in 2003 represents the first time a pharmaceutical agent was used to directly augment the actions of an enzyme by increasing its maximal velocity and substrate affinity. This discovery, coupled with translational medicine which has shown that inactivating and activating GK mutations cause glycemic diseases, has triggered an intensive medicinal chemistry effort in the field of glucokinase activators (GKAs).

Targeting glucokinase activation for the treatment of type 2 diabetes--a status review.

Glucokinase (GK) plays a key role in glucose homeostasis. Developments over the past decade, such as the determination of the function of GK regulatory protein, the discovery of GK mutations related to maturity onset of diabetes of the young, permanent neonatal diabetes mellitus and persistent hyperinsulinemia hypoglycemia of infancy, and the discovery of novel GK activators (GKAs) and their X-ray co-crystal structures with GK, have significantly enhanced our understanding of GK structure and function. This review discusses key publications on GKAs that report full characterization, key compound disclosures from patents, and a current hypothesis on the mechanism of GK activation based on the co-crystal structures of GK-GKA complexes.

Metabolic diseases drug discovery world summit. July 28-29, 2003, San Diego, CA, USA.

In Type 2 diabetes, glucose homeostasis is impaired due to either a decrease in insulin secretion or insulin action. In this symposium, molecular targets that could have an impact on either or both of these defects were discussed and data related to specific compounds were presented. Protein tyrosine phosphatase 1B inhibitors that relieve the negative control on insulin action and are active in cell assays, dipeptidyl peptidase IV inhibitors that raise postprandial glucagon-like peptide 1 levels in animals and humans, and pyruvate dehydrogenase kinase inhibitors that increase the levels of pyruvate dehydrogenase, which in turn improve insulin sensitivity, were all discussed.

Allosteric activators of glucokinase: potential role in diabetes therapy.

Glucokinase (GK) plays a key role in whole-body glucose homeostasis by catalyzing the phosphorylation of glucose in cells that express this enzyme, such as pancreatic beta cells and hepatocytes. We describe a class of antidiabetic agents that act as nonessential, mixed-type GK activators (GKAs) that increase the glucose affinity and maximum velocity (Vmax) of GK. GKAs augment both hepatic glucose metabolism and glucose-induced insulin secretion from isolated rodent pancreatic islets, consistent with the expression and function of GK in both cell types.

Oxazole- and imidazole-based Ser-Leu dipeptide mimetics in potent inhibitors of antigen presentation by MHC class II DR molecules.

Imidazole and oxazole derivatives 1 to 4 were designed and prepared as dipeptide mimetics to replace the Ser-Leu dipeptide sequence of Ro-25-9980 (Ac-(Cha)-RAMA-S-L-NH2), a peptidic inhibitor of antigen binding to major histocompatibility complex (MHC) class II DR molecules linked to rheumatoid arthritis (RA). The most potent analog in binding assays (IC50 = 30 nM in DRB1*0401 binding; 1.6 times as potent as Ro 25-9980) was 16, Ac-(Cha)RAMA-(S)S-psi(oxazole)-L-NH2.

Peptide and peptide mimetic inhibitors of antigen presentation by HLA-DR class II MHC molecules. Design, structure-activity relationships, and X-ray crystal structures.

Molecular features of ligand binding to MHC class II HLA-DR molecules have been elucidated through a combination of peptide structure-activity studies and structure-based drug design, resulting in analogues with nanomolar affinity in binding assays. Stabilization of lead compounds against cathepsin B cleavage by N-methylation of noncritical backbone NH groups or by dipeptide mimetic substitutions has generated analogues that compete effectively against protein antigens in cellular assays, resulting in inhibition of T-cell proliferation. Crystal structures of four ternary complexes of different peptide mimetics with the rheumatoid arthritis-linked MHC DRB10401 and the bacterial superantigen SEB have been obtained.