title: Discovery tactics to mitigate toxicity risks due to reactive metabolite formation with 2-(2-hydroxyaryl)-5-(trifluoromethyl)pyrido[4,3-d]pyrimidin-4(3h)-one derivatives, potent calcium-sensing receptor antagonists and clinical candidate(s) for the treatment of osteoporosis.
abstract: The synthesis and structure-activity relationship studies on 5-trifluoromethylpyrido[4,3-d]pyrimidin-4(3H)-ones as antagonists of the human calcium receptor (CaSR) have been recently disclosed [ Didiuk et al. ( 2009 ) Bioorg. Med. Chem. Lett. 19 , 4555 - 4559 ). On the basis of its pharmacology and disposition attributes, (R)-2-(2-hydroxyphenyl)-3-(1-phenylpropan-2-yl)-5-(trifluoromethyl)pyrido[4,3-d]pyrimidin-4(3H)-one (1) was considered for rapid advancement to first-in-human (FIH) trials to mitigate uncertainty surrounding the pharmacokinetic/pharmacodynamic (PK/PD) predictions for a short-acting bone anabolic agent. During the course of metabolic profiling, however, glutathione (GSH) conjugates of 1 were detected in human liver microsomes in an NADPH-dependent fashion. Characterization of the GSH conjugate structures allowed insight(s) into the bioactivation pathway, which involved CYP3A4-mediated phenol ring oxidation to the catechol, followed by further oxidation to the electrophilic ortho-quinone species. While the reactive metabolite (RM) liability raised concerns around the likelihood of a potential toxicological outcome, a more immediate program goal was establishing confidence in human PK predictions in the FIH study. Furthermore, the availability of a clinical biomarker (serum parathyroid hormone) meant that PD could be assessed side by side with PK, an ideal scenario for a relatively unprecedented pharmacologic target. Consequently, progressing 1 into the clinic was given a high priority, provided the compound demonstrated an adequate safety profile to support FIH studies. Despite forming identical RMs in rat liver microsomes, no clinical or histopathological signs prototypical of target organ toxicity were observed with 1 in in vivo safety assessments in rats. Compound 1 was also devoid of metabolism-based mutagenicity in in vitro (e.g., Salmonella Ames) and in vivo assessments (micronuclei induction in bone marrow) in rats. Likewise, metabolism-based studies (e.g., evaluation of detoxicating routes of clearance and exhaustive PK/PD studies in animals to prospectively predict the likelihood of a low human efficacious dose) were also conducted, which mitigated the risks of idiosyncratic toxicity to a large degree. In parallel, medicinal chemistry efforts were initiated to identify additional compounds with a complementary range of human PK predictions, which would maximize the likelihood of achieving the desired PD effect in the clinic. The back-up strategy also incorporated an overarching goal of reducing/eliminating reactive metabolite formation observed with 1. Herein, the collective findings from our discovery efforts in the CaSR program, which include the incorporation of appropriate derisking steps when dealing with RM issues are summarized.
authors.affiliation: Pharmacokinetics, Dynamics and Metabolism Department and Department of Medicinal Chemistry, Pfizer Global Research and Development, Groton, Connecticut 06340, USA. amit.kalgutkar@pfizer.com
authors.forename: Amit S
authors.initials: AS
authors.lastname: Kalgutkar
authors.forename: David A
authors.initials: DA
authors.lastname: Griffith
authors.forename: Tim
authors.initials: T
authors.lastname: Ryder
authors.forename: Hao
authors.initials: H
authors.lastname: Sun
authors.forename: Zhuang
authors.initials: Z
authors.lastname: Miao
authors.forename: Jonathan N
authors.initials: JN
authors.lastname: Bauman
authors.forename: Mary T
authors.initials: MT
authors.lastname: Didiuk
authors.forename: Kosea S
authors.initials: KS
authors.lastname: Frederick
authors.forename: Sabrina X
authors.initials: SX
authors.lastname: Zhao
authors.forename: Chandra
authors.initials: C
authors.lastname: Prakash
authors.forename: John R
authors.initials: JR
authors.lastname: Soglia
authors.forename: Scott W
authors.initials: SW
authors.lastname: Bagley
authors.forename: Bruce M
authors.initials: BM
authors.lastname: Bechle
authors.forename: Ryan M
authors.initials: RM
authors.lastname: Kelley
authors.forename: Kenneth
authors.initials: K
authors.lastname: Dirico
authors.forename: Michael
authors.initials: M
authors.lastname: Zawistoski
authors.forename: Jianke
authors.initials: J
authors.lastname: Li
authors.forename: Robert
authors.initials: R
authors.lastname: Oliver
authors.forename: Angel
authors.initials: A
authors.lastname: Guzman-Perez
authors.forename: Kevin K C
authors.initials: KK
authors.lastname: Liu
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Walker
authors.forename: John W
authors.initials: JW
authors.lastname: Benbow
authors.forename: Joel
authors.initials: J
authors.lastname: Morris
mesh_terms: D045930:Anabolic Agents / Q000009:adverse effects / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D018360:Crystallography, X-Ray; D006801:Humans; D010024:Osteoporosis / Q000188:drug therapy*; D011725:Pyridines / Q000009:adverse effects / Q000737:chemistry* / Q000378:metabolism*; D011744:Pyrimidinones / Q000009:adverse effects / Q000737:chemistry* / Q000378:metabolism*; D051381:Rats; D044169:Receptors, Calcium-Sensing / Q000037:antagonists & inhibitors*
chemical_list: D045930:Anabolic Agents; D011725:Pyridines; D011744:Pyrimidinones; D044169:Receptors, Calcium-Sensing