The expression and localisation of membrane transporters and P450 enzymes along the longitudinal and crypt-villus axes of the rat intestine and their response to oral imatinib

Abstract

A reduction in oral bioavailability of a wide range of drug compounds may occur following efflux of drug substrates into the intestinal lumen mediated by ATP dependent ABC transporters MDR1, MRP2 and BCRP and/or by enteric metabolism of drug compounds by CYP450 enzymes. Furthermore, inter-individual variability in the expression and function of key drug disposition proteins, which may occur following a regime of drug treatment, has the potential to cause altered pharmacokinetic profiles leading to failure of therapy. This thesis aimed to establish mRNA and protein expression of key drug transporter proteins and CYP450 enzymes both along the length of the intestine and across the crypt-villus axis using NanoString mRNA technology and immunohistochemistry. Key findings include; a) evidence for cytoprotection of the crypt cells distinct from enterocytes, b) a lack of correlation between Mrp2 mRNA and protein expression in the rat small intestine, with a paucity of brush-border protein in all regions, c) using the M70 antibody, Bcrp protein was immunolocalised to the lateral cell membranes of cryptal epithelial in the duodenum, jejunum and ileum with no apparent protein expression above the crypt-villus junction. Oral administration of the tyrosine kinase inhibitor imatinib mesylate was found to cause significant induction of Cyp1a1 mRNA expression with maximal induction in ileum samples showing an average 169-fold increase in expression. Immunohistochemistry indicates an associated increase in Cyp1a1 protein levels. In addition intestinal exposure to imatinib was found to cause significant changes in mRNA expression levels of genes including Mdr1a and Bcrp. This thesis presents key differences with regards to protein localisation of ABC transporters proposed to play an important role in conferring intestinal drug resistance between rat and human tissues and highlights the high levels of complexity associated with oral drug delivery and intestinal absorption.