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.

A restricted role for TYK2 catalytic activity in human cytokine responses revealed by novel TYK2-selective inhibitors.

TYK2 is a JAK family protein tyrosine kinase activated in response to multiple cytokines, including type I IFNs, IL-6, IL-10, IL-12, and IL-23. Extensive studies of mice that lack TYK2 expression indicate that the IFN-α, IL-12, and IL-23 pathways, but not the IL-6 or IL-10 pathways, are compromised. In contrast, there have been few studies of the role of TYK2 in primary human cells. A genetic mutation at the tyk2 locus that results in a lack of TYK2 protein in a single human patient has been linked to defects in the IFN-α, IL-6, IL-10, IL-12, and IL-23 pathways, suggesting a broad role for TYK2 protein in human cytokine responses. In this article, we have used a panel of novel potent TYK2 small-molecule inhibitors with varying degrees of selectivity against other JAK kinases to address the requirement for TYK2 catalytic activity in cytokine pathways in primary human cells. Our results indicate that the biological processes that require TYK2 catalytic function in humans are restricted to the IL-12 and IL-23 pathways, and suggest that inhibition of TYK2 catalytic activity may be an efficacious approach for the treatment of select autoimmune diseases without broad immunosuppression.

Discovery of highly potent and efficacious MC4R agonists with spiroindane N-Me-1,2,4-triazole privileged structures for the treatment of obesity.

We report an SAR study of MC4R analogs containing spiroindane heterocyclic privileged structures. Compound 26 with N-Me-1,2,4-triazole moiety possesses exceptional potency at MC4R and potent anti-obesity efficacy in a mouse model. However, the efficacy is not completely mediated through MC4R. Additional SAR studies led to the discovery of compound 32, which is more potent at MC4R. Compound 32 demonstrates MC4R mediated anti-obesity efficacy in rodent models.

Discovery of a spiroindane based compound as a potent, selective, orally bioavailable melanocortin subtype-4 receptor agonist.

We report the design, synthesis and properties of spiroindane based compound 1, a potent, selective, orally bioavailable, non-peptide melanocortin subtype-4 receptor agonist. Compound 1 shows excellent erectogenic activity in the rodent models.

Spiroindane based amides as potent and selective MC4R agonists for the treatment of obesity.

We report a series of potent and selective MC4R agonists based on spiroindane amide privileged structures for potential treatments of obesity. Among the synthetic methods used, Method C allows rapid synthesis of the analogs. The series of compounds can afford high potency on MC4R as well as good rodent pharmacokinetic profiles. Compound 1r (MK-0489) demonstrates MC4R mediated reduction of food intake and body weight in mouse models. Compound 1r is efficacious in 14-day diet-induced obese (DIO) rat models.

The synthesis and structure-activity relationship of substituted N-phenyl anthranilic acid analogs as amyloid aggregation inhibitors.

It is believed that beta-amyloid aggregation is an important event in the development of Alzheimer's disease. In the course of our studies to identify beta-amyloid aggregation inhibitors, a series of N-phenyl anthranilic acid analogs were synthesized and studied for beta-amyloid inhibition activity. The synthesis, structure-activity relationship, and in vivo activity of these analogs are discussed.

Effect of residue substitution via site-directed mutagenesis on activity and steroselectivity of transaminase BpTA from Bacillus pumilus W3 for sitafloxacin hydrate intermediate.

Aminotransferases are widely employed as biocatalysts for the asymmetric synthesis of biologically active pharmaceuticals. Transaminase BpTA from Bacillus pumilus W3 can accept a broad spectrum of sterically demanding substrates, but it does not process the key five-membered ring intermediate of sitafloxacin. In the present study, we rationally constructed numerous single-point mutants and six multi-point mutants by combining the structural characteristics of transaminase and its substrates. Biochemical characteristics of wild-type and mutant enzymes were initially analyzed, and mutants I215M, I215F, and Y32L displayed increased catalytic efficiency, K155A, I215V and T252A completely lost enzyme activity. Residues K155 and T252 had a particularly strong influence on catalytic activity. Four multi-point mutants (L212M/I215M, Y32L/S190A/L212M/I215M, Y32L/Y159F/T252A and Y32W/Y159F/I215M/T252A) possess potential for industrial production of the key five-membered ring intermediate of sitafloxacin. Furthermore, mutants Y32L/Y159F/T252A and Y32W/Y159F/I215M/T252A can catalyze conversion of (R)-α-phenethylamine, albeit at an extremely low rate (<8%). In summary, mutants L212M/I215M and Y32L/S190A/L212M/I215M are more suitable for industrial production of the antibiotic, sitafloxacin, via an enzymatic approach.

Mining of aminotransferase gene ota3 from Bacillus pumilus W3 via genome analysis, gene cloning and expressing for compound bioamination.

Aminotransferases are widely employed as biocatalysts to produce chiral amines and biologically active pharmaceuticals via asymmetric synthesis. In this study, transaminase genes in the Bacillus pumilus W3 genome were analysed, and gene ota3 encoding a putative (R)-selective transaminase was identified. The sequence of ota3 shares highest sequence identity (24.7%) with the first (R)-selective aminotransferase from Arthrobacter sp. KNK 168. Amino acid sequence and conserved domains analyses indicated that ω-BPAT encoded by ota3 belonged to the pyridoxal 5'-phosphate-dependent class IV (PLPDE_IV) superfamily. Both native and codon-optimised ω-BPAT genes were recombinantly expressed, and the purified proteins had a molecular mass of ~33.4 kDa. Furthermore, enantioselectivity tests with (S)- and (R)-α-phenethylamine revealed its (R)-selectivity. The optimal conditions for catalytic reaction were 45 °C and pH 7.0, and ω-BPAT retained stability at 20 °C and pH 7.0. Thus, ω-BPAT is a novel (R)-selective aminotransferase with great potential as a universal biocatalyst.

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.

Melanocortin subtype 4 receptor agonists: structure-activity relationships about the 4-alkyl piperidine core.

SAR about the piperidine core in a series of MC4R agonists is described. A number of alkyl substituents that furnish compounds with good affinity and functional potency are reported.