Effective in vitro evaluation of the risk of histamine release related to valemetostat tosylate using MRGPRX2-expressing cells.

Mas-related G-protein-coupled receptor X2 (MRGPRX2), expressed on mast cells, is associated with drug-induced pseudo-allergic reactions. Although it is well known that there are differences of sensitivity between species in the pseudo-allergic reactions, no platform for evaluating a human risk of the pseudo-allergic reactions observed in nonclinical studies has been established. Valemetostat tosylate, developed as an anti-cancer drug, induced histamine release in a nonclinical study with dogs. The purpose of the current study was to identify the mechanism and assess the human risk of valemetostat-tosylate-induced histamine release using dog and human MRGPRX2-expressing cells. In an experiment with human or dog MRGPRX2-expressing cells, valemetostat tosylate caused activation of human and dog MRGPRX2. Importantly, the EC50 for dog MRGPRX2 was consistent with the Cmax value at which histamine release was observed in dogs. Furthermore, the EC50 for human MRGPRX2 was ca. 27-fold higher than that for dog MRGPRX2, indicating a species difference in histamine-releasing activity. In a clinical trial, histamine release was not observed in patients receiving valemetostat tosylate. In conclusion, an in vitro assay using human and animal MRGPRX2-expressing cells would be an effective platform to investigate the mechanism and predict the human risk of histamine release observed in nonclinical studies.

Embryofetal development study of vismodegib, a hedgehog pathway inhibitor, in rats.

Vismodegib (Erivedge) is a first-in-class small-molecule hedgehog pathway inhibitor for the treatment of adults with advanced basal-cell carcinoma. Because this pathway is known to play key roles in patterning and growth during vertebrate development, vismodegib was anticipated to be embryotoxic. To support marketing applications, an embryofetal development study was completed in which a limited number of pregnant rats (n = 6/group) was administered vismodegib by oral gavage on gestation days 6 to 17. When vismodegib was administered at ≥60 mg/kg/day, doses associated with evidence of pharmacologic activity in previous rat toxicity studies, all conceptuses were resorbed at an early embryonic stage in the absence of significant maternal toxicity. When administered at 10 mg/kg/day, corresponding to an exposure (AUC0-24h ) approximately 15% of the median in patients at steady state, a variety of malformations were observed, including absent/fused digits in the hindlimb of multiple fetuses, multiple craniofacial abnormalities in one fetus, and an anorectal defect in one fetus. In addition, the incidence of variations, including dilated renal pelvis or ureter and incompletely or unossified skeletal elements, was significantly greater when compared with the controls. These results confirmed that vismodegib is likely to be embryotoxic at clinically relevant maternal exposures, and doses ≥60 mg/kg/day resulted in a 100% incidence of embryolethality that likely resulted from severe defects in early embryonic development. In contrast, craniofacial defects typically associated with hedgehog pathway inhibition were only observed in one fetus at the low dose of 10 mg/kg/day, which likely reflected minimal or intermittent pathway inhibition at low exposures.

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. Subsequent SAR studies of 1 led to the identification of a pyrazine-based lead analogue 3, lacking the thiazole moiety. In vivo metabolite identification studies, followed by the independent synthesis and profiling of the cyclopentyl keto- and hydroxyl- metabolites of 3, led to the selection of piragliatin, 4, as the clinical lead. Piragliatin was found to lower pre- and postprandial glucose levels, improve the insulin secretory profile, increase ß-cell sensitivity to glucose, and decrease hepatic glucose output in patients with T2D.

Liarozole markedly increases all trans-retinoic acid toxicity in mouse limb bud cell cultures: a model to explain the potency of the aromatic retinoid (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthylenyl)-1-propenyl] benzoic acid.

The remarkable toxicity of (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthylenyl)-1-propenyl] benzoic acid (TTNPB) compared to all trans-retinoic acid (tRA) is due to multiple factors, including reduced affinities for cytosolic binding proteins (CRABPs), resistance to metabolism, and prolonged nuclear receptor activation. To further investigate the role of half-life in retinoid toxicity, experiments were performed to determine whether, and to what extent, inhibition of tRA metabolism by liarozole increased its toxicity comparable to that of TTNPB in the mouse limb bud system. Liarozole is a known inhibitor of tRA 4-hydroxylation (CYP26). In the absence of liarozole, the IC50 for inhibition of chondrogenesis by tRA was 140 nM compared to 0.3 nM for TTNPB, a 467-fold difference. Following the addition of liarozole (10(-6) M) to limb bud cultures, the potency of tRA to inhibit chondrogenesis was increased approximately 14-fold (IC50 of 9.8 nM). Although liarozole markedly increased toxicity of tRA in mouse limb bud micromass cultures, tRA metabolism was inhibited only about 10%. These results indicate that a relatively minor decrease in the metabolism of tRA in the mouse limb bud system is associated with a marked enhancement of toxicity that is likely related to the prolongation of tRA half-life during a critical period of development. Thus, the prolonged half-life of TTNPB is the most significant factor contributing to the remarkable teratogenicity of this synthetic aromatic retinoid.