Drug absorption evaluation, prodrug design and intestinal transporter expression by GeneChip analysis.

Abstract

The purpose is to identify transporters for drug absorption evaluation, prodrug design and <italic>in vivo/in vitro</italic> permeability correlation. Prodrug uptake and hPepT1 localization were determined in hPepT1-overexpressed cells. Drug permeability was determined by single-pass intestinal perfusion and gene expression was measured by GeneChip analysis. Prodrug valacyclovir inhibited GlySar uptake by hPepT1. Confocal microscopy showed that hPepT1 localized in Caco-2 apical membrane and was absent in either early endosome or lysosome under normal conditions. The <italic>in vivo/in vitro</italic> permeability correlated well for passive absorbed drugs, while the permeability for carrier-mediated drugs was 7-fold higher in human above the correlation of passive absorbed drugs. There were 40% of 12,559 sequences expressed in Caco-2 cells and human duodenum with 1000 genes showed 5-fold difference. The observed differences of gene expression were consistent with observed differences in drug permeability. Valacyclovir rat intestinal permeability showed 2--5-fold higher than parent drug acyclovir. The high protein diet decreased valacyclovir permeability by 2--3-fold. There were 40% of 8739 rat sequences expressed in intestine with 340--499 sequences showed 5-fold differences comparing the small intestinal regions to colon. The high protein diet changed the expression of 26--46 sequences by 5-fold. The average expression of transporters and metabolizing enzymes was decreased along the GI tract by 2--3-fold with the exception that the expression of transporters including mdr1, MLP2, MCT1 increased along the GI tract by 5--25-fold from duodenum to colon. Cluster analysis showed duodenum and jejunum exhibited a similar expression pattern, while ileum and colon showed a similar expression pattern. Valacyclovir permeability <italic>in vivo</italic> correlated with expression of sodium/purine transporter (rCNT2), and an EST sequence similar to acyl carrier, an ER ATPase as well as 7 other proteins including tubulin. In conclusion, the 2--5-fold higher permeability of valacyclovir than its parent drug is consistent with their 2--3-fold difference in human bioavailability. Although valacyclovir inhibited GlySar uptake by hPepT1 <italic> in vitro</italic>, its permeability did not correlate with PepT1 mRNA level <italic> in vivo</italic>. The correlation of valacyclovir permeability with the expression of CNT2 and other sequences indicated that more than one transporter is involved in valacyclovir intestinal absorption.