Jodi K. Muckelbauer

Synthesis and SAR of indole-and 7-azaindole-1,3-dicarboxamide hydroxyethylamine inhibitors of BACE-1.

Heterocyclic replacement of the isophthalamide phenyl ring in hydroxyethylamine (HEA) BACE-1 inhibitors was explored. A variety of indole-1,3-dicarboxamide HEAs exhibited potent BACE-1 enzyme inhibition, but displayed poor cellular activity. Improvements in cellular activity and aspartic protease selectivity were observed for 7-azaindole-1,3-dicarboxamide HEAs. A methylprolinol-bearing derivative (10n) demonstrated robust reductions in rat plasma Aβ levels, but did not lower rat brain Aβ due to poor central exposure. The same analog exhibited a high efflux ratio in a bidirectional Caco-2 assay and was likely a substrate of the efflux transporter P-glycoprotein. X-ray crystal structures are reported for two indole HEAs in complex with BACE-1.

Acyl guanidine inhibitors of β-secretase (BACE-1): optimization of a micromolar hit to a nanomolar lead via iterative solid- and solution-phase library synthesis.

This report describes the discovery and optimization of a BACE-1 inhibitor series containing an unusual acyl guanidine chemotype that was originally synthesized as part of a 6041-membered solid-phase library. The synthesis of multiple follow-up solid- and solution-phase libraries facilitated the optimization of the original micromolar hit into a single-digit nanomolar BACE-1 inhibitor in both radioligand binding and cell-based functional assay formats. The X-ray structure of representative inhibitors bound to BACE-1 revealed a number of key ligand:protein interactions, including a hydrogen bond between the side chain amide of flap residue Gln73 and the acyl guanidine carbonyl group, and a cation-π interaction between Arg235 and the isothiazole 4-methoxyphenyl substituent. Following subcutaneous administration in rats, an acyl guanidine inhibitor with single-digit nanomolar activity in cells afforded good plasma exposures and a dose-dependent reduction in plasma Aβ levels, but poor brain exposure was observed (likely due to Pgp-mediated efflux), and significant reductions in brain Aβ levels were not obtained.

Design, synthesis and structure-activity relationships of azole acids as novel, potent dual PPAR alpha/gamma agonists.

The design, synthesis and structure-activity relationships of a novel series of N-phenyl-substituted pyrrole, 1,2-pyrazole and 1,2,3-triazole acid analogs as PPAR ligands are outlined. The triazole acid analogs 3f and 4f were identified as potent dual PPARalpha/gamma agonists both in binding and functional assays in vitro. The 3-oxybenzyl triazole acetic acid analog 3f showed excellent glucose and triglyceride lowering in diabetic db/db mice.

Discovery of an oxybenzylglycine based peroxisome proliferator activated receptor alpha selective agonist 2-((3-((2-(4-chlorophenyl)-5-methyloxazol-4-yl)methoxy)benzyl)(methoxycarbonyl)amino)acetic acid (BMS-687453).

An 1,3-oxybenzylglycine based compound 2 (BMS-687453) was discovered to be a potent and selective peroxisome proliferator activated receptor (PPAR) alpha agonist, with an EC(50) of 10 nM for human PPARalpha and approximately 410-fold selectivity vs human PPARgamma in PPAR-GAL4 transactivation assays. Similar potencies and selectivity were also observed in the full length receptor co-transfection assays. Compound 2 has negligible cross-reactivity against a panel of human nuclear hormone receptors including PPARdelta. Compound 2 demonstrated an excellent pharmacological and safety profile in preclinical studies and thus was chosen as a development candidate for the treatment of atherosclerosis and dyslipidemia. The X-ray cocrystal structures of the early lead compound 12 and compound 2 in complex with PPARalpha ligand binding domain (LBD) were determined. The role of the crystal structure of compound 12 with PPARalpha in the development of the SAR that ultimately resulted in the discovery of compound 2 is discussed.

Discovery of azetidinone acids as conformationally-constrained dual PPARα/γ agonists

A novel class of azetidinone acid-derived dual PPARalpha/gamma agonists has been synthesized for the treatment of diabetes and dyslipidemia. The preferred stereochemistry in this series for binding and functional agonist activity against both PPARalpha and PPARgamma receptors was shown to be 3S,4S. Synthesis, in vitro and in vivo activities of compounds in this series are described. A high-yielding method for N-arylation of azetidinone esters is also described.

Tyrosine Kinase 2-mediated Signal Transduction in T Lymphocytes Is Blocked by Pharmacological Stabilization of Its Pseudokinase Domain

Background: Interleukin-23 mediates pathobiology in many autoimmune disorders. Results: A chemogenomics approach identified small molecule agents that block receptor-mediated activation or tyrosine kinase 2 (Tyk2) and downstream signaling. Compounds stabilize the pseudokinase domain of Tyk2. Conclusion: Small molecule ligands of the Tyk2 pseudokinase domain stabilize an autoinhibitory interaction with the catalytic domain. Significance: This work enables the discovery of selective therapeutics targeting Tyk2-dependent pathways critical in autoimmunity. Inhibition of signal transduction downstream of the IL-23 receptor represents an intriguing approach to the treatment of autoimmunity. Using a chemogenomics approach marrying kinome-wide inhibitory profiles of a compound library with the cellular activity against an IL-23-stimulated transcriptional response in T lymphocytes, a class of inhibitors was identified that bind to and stabilize the pseudokinase domain of the Janus kinase tyrosine kinase 2 (Tyk2), resulting in blockade of receptor-mediated activation of the adjacent catalytic domain. These Tyk2 pseudokinase domain stabilizers were also shown to inhibit Tyk2-dependent signaling through the Type I interferon receptor but not Tyk2-independent signaling and transcriptional cellular assays, including stimulation through the receptors for IL-2 (JAK1- and JAK3-dependent) and thrombopoietin (JAK2-dependent), demonstrating the high functional selectivity of this approach. A crystal structure of the pseudokinase domain liganded with a representative example showed the compound bound to a site analogous to the ATP-binding site in catalytic kinases with features consistent with high ligand selectivity. The results support a model where the pseudokinase domain regulates activation of the catalytic domain by forming receptor-regulated inhibitory interactions. Tyk2 pseudokinase stabilizers, therefore, represent a novel approach to the design of potent and selective agents for the treatment of autoimmunity.

Purine derivatives as potent Bruton's tyrosine kinase (BTK) inhibitors for autoimmune diseases.

Investigation of various heterocyclic core isosteres of imidazopyrazines 1 & 2 yielded purine derivatives 3 & 8 as potent and selective BTK inhibitors. Subsequent SAR studies of the purine series led to the discovery of 20 as a leading compound. Compound 20 is very selective when screened against a panel of 400 kinases and is a potent inhibitor in cellular assays of human B cell function including B-Cell proliferation and CD86 cell surface expression and exhibited in vivo efficacy in a mouse PCA model. Its X-ray co-crystal structure with BTK shows that the high selectivity is gained from filling a BTK specific lipophilic pocket. However, physical and ADME properties leading to low oral exposure hindered further development.

Monosubstituted γ-lactam and conformationally constrained 1,3-diaminopropan-2-ol transition-state isostere inhibitors of β-secretase (BACE)

The synthesis, evaluation, and structure-activity relationships of a class of γ-lactam 1,3-diaminopropan-2-ol transition-state isostere inhibitors of BACE are discussed. Two strategies for optimizing lead compound 1a are presented. Reducing the overall size of the inhibitors resulted in the identification of γ-lactam 1i, whereas the introduction of conformational constraint on the prime-side of the inhibitor generated compounds such as the 3-hydroxypyrrolidine inhibitor 28n. The full in vivo profile of 1i in rats and 28n in Tg 2576 mice is presented.

Design and synthesis of carbazole carboxamides as promising inhibitors of Bruton's tyrosine kinase (BTK) and Janus kinase 2 (JAK2).

Four series of disubstituted carbazole-1-carboxamides were designed and synthesised as inhibitors of Brutons tyrosine kinase (BTK). 4,7- and 4,6-disubstituted carbazole-1-carboxamides were potent and selective inhibitors of BTK, while 3,7- and 3,6-disubstituted carbazole-1-carboxamides were potent and selective inhibitors of Janus kinase 2 (JAK2).

X-Ray Crystal Structure of Bone Marrow Kinase in the X Chromosome: A Tec Family Kinase

Bone marrow kinase in the X chromosome, a member of the Tec family of tyrosine kinases, plays a role in both monocyte/macrophage trafficking as well as cytokine secretion. Although the structures of Tec family kinases Bruton’s tyrosine kinase and IL‐2‐inducible T‐cell kinase are known, the crystal structures of other Tec family kinases have remained elusive. We report the X‐ray crystal structures of bone marrow kinase in the X chromosome in complex with dasatinib at 2.4u2003Å resolution and PP2 at 1.9u2003Å resolution. The bone marrow kinase in the X chromosome structures reveal a typical kinase protein fold; with well‐ordered protein conformation that includes an open/extended activation loop and a stabilized DFG‐motif rendering the kinase in an inactive conformation. Dasatinib and PP2 bind to bone marrow kinase in the X chromosome in the ATP binding pocket and display similar binding modes to that observed in other Tec and Src protein kinases. The bone marrow kinase in the X chromosome structures identify conformational elements of the DFG‐motif that could potentially be utilized to design potent and/or selective bone marrow kinase in the X chromosome inhibitors.