The goal of the proposed research is to determine the biochemical and molecular genetic mechanisms underlying the gastrointestinal absorption of peptoid-type drugs and to develop approaches to improving the oral delivery of drugs in humans. Rapid pharmacological screening methodologies combined with combinatorial synthetic approaches has produced a diverse array of compounds of potential therapeutic utility. However, the therapeutic utility particularly for peptide and peptide-type drugs is severely limited by their low oral absorption. The proposed research will take advantage of new molecular biological results and methodologies to advance our understanding of peptide transport. These approaches will be combined with the determination of intestinal membrane permeabilities in cell, animal and human systems allowing for cross- correlations between species including humans. Finally, this proposal will develop extensions of a prodrug strategy utilizing the peptide transporter found to be successful for improving the oral systemic delivery of polar compounds.
The specific aims of the proposed project are: A. Determine the structural requirements for intestinal and hepatic transport of peptoid-type compounds and extend the mechanistic correlations between cell, tissue, animal and human absorption. B. Extend the structure-transport relationship for epithelial cell and hepatocyte transport as a basis for the design of prodrugs for oral drug delivery and evaluate the effectiveness of this strategy in improving oral delivery. C. Identified genes mediating peptide transport and compare with the cloned gene from human colon carcinoma HT-29 cells identified in our laboratories and establish transfected cell lines for mechanistic studies of peptoid transport. D. Determine the subcellular distribution of the peptide transporter genes and functional expression at the cell surface in cell culture, intestine, and liver tissue through immunohistochemistry and transport assays and determine the requirements for functional expression of the transporter at the plasma membrane surface. E. Identify the functional domains of the peptide transporters using sequence analysis and site-directed mutagenesis, and develop a structural model to understand substrate binding and transport at the molecular level. F. Develop a nonlinear absorption model incorporating intestinal and hepatic permeability, including site dependent permeability, nonlinear absorption, enzyme hydrolysis and transit rate that can be used to predict human oral absorption. Thus the proposed research project will combine molecular biological approaches with integrated cellular, animal and human studies to establish the mechanistic basis for peptide type drug absorption. It will develop molecular and kinetic information and utilize this knowledge to develop and test integrated approaches to estimating and improving the bioavailability of peptoid drugs. This research collaboration allows us to work at the very frontier of current research methodologies and provides an opportunity to truly impact oral drug delivery of peptide-type drugs.
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