Graduation Date

Fall 12-15-2017

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Pharmaceutical Sciences

Abstract

All biomedical research is conducted in animal models first. In addition, the Food and Drug Administration requires extrapolation from animal data to predict human responses. There are ongoing scientific and regulatory challenges translating interspecies comparisons and predictions. Metabolic pathways are a cornerstone to understanding drug metabolism and toxicities and the liver is a key organ in this process. Bile acids (BAs) play a central role in the hepatobiliary toxicities of chemicals, toxins, and biological reagents. BAs have many physiological functions including regulation of genes involved in cholesterol and glucose metabolism and BA homeostasis. However, BAs also have several pathological effects including carcinogenicity and liver toxicity. Maintenance of bile acid (BA) homeostasis is essential to achieve their physiologic functions and avoid their toxic effects. Several metabolic pathways including sulfation by sulfotransferase (SULT), glucuronidation by UDP-glucuronosyltransferases (UGTs), and oxidation by Cytochrome-P450 (CYP450) enzymes participate in the direct detoxification, enhance the elimination of BAs, and help maintain their homeostasis. In addition, influx and efflux transporters at both the sinusoidal and basolateral membranes play an important role in determining intracellular BA concentration, and therefore their hepatotoxicity. There are known species differences in BA metabolism and transporter. There are known species differences in the composition of the BA pool, the toxicity of BAs, and drug-induced hepatotoxicity related to BAs. These species differences can prevent extrapolation of toxicity profiles of xenobiotics between species causing a serious disconnect between preclinical safety findings in rodent and canine animal models and safety finding at clinical stages. In this thesis, we compare the metabolic profile of representative BAs between several species including humans, chimpanzee, monkeys, minipigs, hamster, rabbits, dogs, rats, and mice. The metabolic profile was characterized by the identification of BA metabolites and by quantifying the kinetics of their formation in hepatocyte S9 in-vitro system. The relative contributions of individual metabolic pathways were determined. LC-MS/MS was used for the qualitative and quantitative analysis of BAs and their metabolites. A mixture of stable-isotope labeled (2H4) and unlabeled BAs were used to facilitate the identification of all minor and major metabolites. Major species differences were found in the metabolism of BAs. Amidation with taurine and glycine was the major pathway in all species. Sulfation was predominant in humans, whereas oxidation and glucuronidation were predominant in rodents and dogs, respectively. Glucuronidation and amidation of BAs are exclusive, where glucuronidation only takes place for unamidated BAs. In vitro-In vivo extrapolation (IVIVE) is performed to establish the correlation of the in vitro results and the bile acid profile in vivo. These results explain, at least in part, the dissociation between preclinical toxicity data in various in vitro and in vivo models and toxicities observed in humans. Furthermore, more relevant species are suggested based on the similarity to the human BA metabolism. In addition, we also screened different bile acids as a potential biomarker for transporter activity using cynomolgus monkey as preclinical model. This resulted in identification of key bile acid sulfates as biomarker for transporter mediated drug-drug interactions.

DMD45-721_THAKA_170911.pdf (54 kB)
copyright permission-ASPET

Biomedical Chromatography.pdf (1329 kB)
copyright permission-Biomedical Chromatography

JPBA.pdf (1924 kB)
copyright permission-JPBA

Toxicological Sciences-I.pdf (638 kB)
copyright permission-Tox Sci

Toxicological Sciences-II.pdf (629 kB)
copyright permission-Tox Sci-II

Available for download on Sunday, December 01, 2019

Share

COinS