Discovery of potent inhibitors of the lysophospholipase autotaxin.

The autotaxin-lysophosphatidic acid (ATX-LPA) axis has been implicated in several disease conditions including inflammation, fibrosis and cancer. This makes ATX an attractive drug target and its inhibition may lead to useful therapeutic agents. Through a high throughput screen (HTS) we identified a series of small molecule inhibitors of ATX which have subsequently been optimized for potency, selectivity and developability properties. This has delivered drug-like compounds such as 9v (CRT0273750) which modulate LPA levels in plasma and are suitable for in vivo studies. X-ray crystallography has revealed that these compounds have an unexpected binding mode in that they do not interact with the active site zinc ions but instead occupy the hydrophobic LPC pocket extending from the active site of ATX together with occupying the LPA 'exit' channel.

Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration.

Autotaxin (ATX) is a secreted lysophospholipase D (lysoPLD) that binds to integrin adhesion receptors. We dissected the roles of integrin binding and lysoPLD activity in stimulation of human breast cancer and mouse aortic vascular smooth muscle cell migration by ATX. We compared effects of wild-type human ATX, catalytically inactive ATX, an integrin binding-defective ATX variant with wild-type lysoPLD activity, the isolated ATX integrin binding N-terminal domain, and a potent ATX selective lysoPLD inhibitor on cell migration using transwell and single-cell tracking assays. Stimulation of transwell migration was reduced (18 or 27% of control, respectively) but not ablated by inactivation of integrin binding or inhibition of lysoPLD activity. The N-terminal domain increased transwell migration (30% of control). ATX lysoPLD activity and integrin binding were necessary for a 3.8-fold increase in the fraction of migrating breast cancer cell step velocities >0.7 µm/min. ATX increased the persistent directionality of single-cell migration 2-fold. This effect was lysoPLD activity independent and recapitulated by the integrin binding N-terminal domain. Integrin binding enables uptake and intracellular sequestration of ATX, which redistributes to the front of migrating cells. ATX binding to integrins and lysoPLD activity therefore cooperate to promote rapid persistent directional cell migration.

IOA-244 is a Non-ATP-competitive, Highly Selective, Tolerable PI3K Delta Inhibitor That Targets Solid Tumors and Breaks Immune Tolerance.

PI3K delta (PI3Kδ) inhibitors are used to treat lymphomas but safety concerns and limited target selectivity curbed their clinical usefulness. PI3Kδ inhibition in solid tumors has recently emerged as a potential novel anticancer therapy through the modulation of T-cell responses and direct antitumor activity. Here we report the exploration of IOA-244/MSC2360844, a first-in-class non-ATP-competitive PI3Kδ inhibitor, for the treatment of solid tumors. We confirm IOA-244's selectivity as tested against a large set of kinases, enzymes, and receptors. IOA-244 inhibits the in vitro growth of lymphoma cells and its activity correlates with the expression levels of PIK3CD, suggesting cancer cell-intrinsic effects of IOA-244. Importantly, IOA-244 inhibits regulatory T cell proliferation while having limited antiproliferative effects on conventional CD4+ T cells and no effect on CD8+ T cells. Instead, treatment of CD8 T cells with IOA-244 during activation, favors the differentiation of memory-like, long-lived CD8, known to have increased antitumor capacity. These data highlight immune-modulatory properties that can be exploited in solid tumors. In CT26 colorectal and Lewis lung carcinoma lung cancer models, IOA-244 sensitized the tumors to anti-PD-1 (programmed cell death protein 1) treatment, with similar activity in the Pan-02 pancreatic and A20 lymphoma syngeneic mouse models. IOA-244 reshaped the balance of tumor-infiltrating cells, favoring infiltration of CD8 and natural killer cells, while decreasing suppressive immune cells. IOA-244 presented no detectable safety concerns in animal studies and is currently in clinical phase Ib/II investigation in solid and hematologic tumors. Significance: IOA-244 is a first-in-class non-ATP-competitive, PI3Kδ inhibitor with direct antitumor in vitro activity correlated with PI3Kδ expression. The ability to modulate T cells, in vivo antitumor activity in various models with limited toxicity in animal studies provides the rationale for the ongoing trials in patients with solid tumors and hematologic cancers.

Optimization of novel acyl pyrrolidine inhibitors of hepatitis C virus RNA-dependent RNA polymerase leading to a development candidate.

Optimization of a pyrrolidine-based template using structure-based design and physicochemical considerations has provided a development candidate 20b (3082) with submicromolar potency in the HCV replicon and good pharmacokinetic properties.

Discovery of further pyrrolidine trans-lactams as inhibitors of human neutrophil elastase (HNE) with potential as development candidates and the crystal structure of HNE complexed with an inhibitor (GW475151).

Described herein is a modern approach to the rapid preparation and evaluation of compounds as potential back-up drug candidates. GW311616A, 1, a derivative of pyrrolidine trans-lactams, has previously been described as a potent, orally active inhibitor of human neutrophil elastase (HNE) for the treatment of respiratory disease. These properties made it a suitable candidate for development. Described here is the discovery of three further derivatives of pyrrolidine trans-lactams, which fulfill the criteria required for back-up candidates 28, 29, and 32. These include increased activity in inhibiting HNE in human whole blood (HWB) and comparable pharmacokinetic properties, in particular clearance, in two species. To provide a rapid assessment of clearance, cassette dosing in dog was used. Modern array techniques, including the synthesis of mixtures, were used to synthesize compounds rapidly. Having selected three potential compounds as back-up candidates, they were prepared as single enantiomers and profiled in in vitro and in vivo assays and evaluated pharmacokinetically in rat and dog. These compounds are highly potent and selective HNE inhibitors, with a prolonged pharmacodynamic action. Pharmacokinetically, these compounds are comparable with 1 while they are more potent in HWB. Compound 28, however, has a higher clearance. One of these compounds, 32, was cocrystallized with HNE, and features of this structure are described and compared with the cocrystal structure of 1 in porcine pancreatic elastase.

Applications of the amino-Cope rearrangement: synthesis of tetrahydropyran, delta-lactone and piperidine targets.

We report a novel approach to some chiral tetrahydropyran and delta-lactone targets that utilizes the asymmetric amino-Cope rearrangement as a key synthetic step. Products of amino-Cope rearrangement chemistry have also been applied to access piperidine targets, further demonstrating the potential of the methodology.