Identification of an imidazopyridine scaffold to generate potent and selective TYK2 inhibitors that demonstrate activity in an in vivo psoriasis model.

Herein we report identification of an imidazopyridine class of potent and selective TYK2 inhibitors, exemplified by prototype 6, through constraint of the rotatable amide bond connecting the pyridine and aryl rings of compound 1. Further optimization led to generation of compound 30 that potently inhibits the TYK2 enzyme and the IL-23 pathway in cells, exhibits selectivity against cellular JAK2 activity, and has good pharmacokinetic properties. In mice, compound 30 demonstrated dose-dependent reduction of IL-17 production in a PK/PD model as well as in an imiquimod-induced psoriasis model. In this efficacy model, the IL-17 decrease was accompanied by a reduction of ear thickness indicating the potential of TYK2 inhibition as a therapeutic approach for psoriasis patients.

Preclinical assessment of novel BRAF inhibitors: integrating pharmacokinetic-pharmacodynamic modelling in the drug discovery process.

The objective of these studies were to determine the preclinical disposition of the two BRAF inhibitors, G-F and G-C, followed by pharmacokinetic (PK)-pharmacodynamic (PD) modelling to characterize the concentration-efficacy relationship of these compounds in the Colo205 mouse xenograft model. With G-F, the relationship of pERK inhibition to concentration was also characterized. Compounds G-F and G-C were administered to mice, rats and dogs and the pharmacokinetics of G-F and G-C was determined. In addition, using indirect response models the concentration-efficacy relationship was described. The clearance of G-F was low; 0.625 and 4.65 mL/min/kg in rat and dog respectively. Similarly, the clearance of G-C was low in rat and dog, 0.490 and 4.43 mL/min/kg, respectively. Both compounds displayed low volumes of distribution (0.140-0.267 L/kg), resulting in moderate half-lives across species (~2.5 to 4 h). Bioavailability was formulation dependent and decreased with increasing dose. Using the indirect response models, the KC(50) (50% K(max); maximal response) value for tumor growth inhibition for G-F and G-C were 84.5 and 19.2 µM, respectively. The IC(50) for pERK inhibition in Colo205 tumors by G-F was estimated to be 29.2 µM. High exposures of G-F and G-C were required for efficacy. Despite good PK properties of low CL and moderate half-life, limitations in obtaining exposures adequate for safety testing in rat and dog resulted in development challenges.

Lead optimization of a 4-aminopyridine benzamide scaffold to identify potent, selective, and orally bioavailable TYK2 inhibitors.

Herein we report our lead optimization effort to identify potent, selective, and orally bioavailable TYK2 inhibitors, starting with lead molecule 3. We used structure-based design to discover 2,6-dichloro-4-cyanophenyl and (1R,2R)-2-fluorocyclopropylamide modifications, each of which exhibited improved TYK2 potency and JAK1 and JAK2 selectivity relative to 3. Further optimization eventually led to compound 37 that showed good TYK2 enzyme and interleukin-12 (IL-12) cell potency, as well as acceptable cellular JAK1 and JAK2 selectivity and excellent oral exposure in mice. When tested in a mouse IL-12 PK/PD model, compound 37 showed statistically significant knockdown of cytokine interferon-γ (IFNγ), suggesting that selective inhibition of TYK2 kinase activity might be sufficient to block the IL-12 pathway in vivo.

Lead identification of novel and selective TYK2 inhibitors.

A therapeutic rationale is proposed for the treatment of inflammatory diseases, such as psoriasis and inflammatory bowel diseases (IBD), by selective targeting of TYK2. Hit triage, following a high-throughput screen for TYK2 inhibitors, revealed pyridine 1 as a promising starting point for lead identification. Initial expansion of 3 separate regions of the molecule led to eventual identification of cyclopropyl amide 46, a potent lead analog with good kinase selectivity, physicochemical properties, and pharmacokinetic profile. Analysis of the binding modes of the series in TYK2 and JAK2 crystal structures revealed key interactions leading to good TYK2 potency and design options for future optimization of selectivity.

Determination of loperamide in mdr1a/1b knock-out mouse brain tissue using matrix-assisted laser desorption/ionization mass spectrometry and comparison with quantitative electrospray-triple quadrupole mass spectrometry analysis.

Recently matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) imaging has been used to analyze small molecule pharmaceutical compounds directly on tissue sections to determine spatial distribution within target tissue and organs. The data presented to date usually indicate relative amounts of drug within the tissue. The determination of absolute amounts is still done using tissue homogenization followed by traditional liquid chromatography-tandem mass spectrometry (LC-MS/MS). In this study, the quantitative determination of loperamide, an antidiarrheal agent and a P-glycoprotein substrate, in mdr1a/1b (-/-) mouse brain tissue sections using MALDI MS on a quadrupole time-of-flight mass spectrometry is described. 5 mg/mL α-cyano-4-hydroxycinnamic acid in 50% acetonitrile with 0.1% trifluoroacetic acid and 0.5 µM reserpine was used as the MALDI matrix. The calibration curve constructed by the peak intensities of standard samples from MALDI MS was linear from 0.025 to 0.5 µM with r² = 0.9989. The accuracy of calibration curve standards was 78.3-105.9% and the percent deviation was less than 25%. Comparison between direct MALDI tissue analysis and conventional tissue analysis using homogenization followed by electrospray LC-MS/MS was also explored.