P450-mediated O-demethylated metabolite is responsible for rat hepatobiliary toxicity of pyridyltriazine-containing PI3K inhibitors

The dysregulation of phosphatidylinositol 3-kinase (PI3K)-dependent pathways is implicated in several human cancers making it an attractive target for small molecule PI3K inhibitors. A series of potent pyridyltriazine-containing inhibitors of class Ia PI3Ks were synthesized and a subset of compounds was evaluated in exploratory repeat-dose rat toxicology studies. Daily oral dosing of compound 1: in Sprague Dawley rats for four consecutive days was associated with hepatobiliary toxicity that included biliary epithelial hyperplasia and hypertrophy, periductular edema, biliary stasis, and acute peribiliary inflammatory infiltrates. These histological changes were associated with clinical pathology changes that included increased serum liver enzymes, total bile acids, and bilirubin. The predominant clearance pathway of 1: was shown in vitro and in a bile-duct cannulated rat (14)C-ADME study to be P450-mediated oxidative metabolism. An O-demethylated pyridine metabolite, M3: , was identified as a candidate proximal metabolite that caused the hepatotoxicity. Co-administration of the pan-P450 inhibitor 1-aminobenzotriazole with 1: to rats significantly reduced the formation of M3: and prevented liver toxicity, whereas direct administration of M3: reproduced the toxicity. Structural changes were introduced to 1: to make the methoxypyridine ring less susceptible to P450 oxidation (compound 2: ), and addition of a methyl group to the benzylic carbon (compound 3: ) improved the pharmacokinetic profile. These changes culminated in the successful design of a clinical candidate 3: (AMG 511) that was devoid of liver toxicity in a 14-day rat toxicity study. Herein, we describe how a metabolism-based structure-activity relationship analysis allowed for the successful identification of a PI3K inhibitor devoid of off-target toxicity.