Discovery of novel PCFT-targeted agents

Wayne State University, Detroit MI

Investigators

Linked publications & trials

Abstract

DESCRIPTION (provided by applicant): In recent years, there has been renewed emphasis on folate-based therapeutics for cancer based on capacities for tumor-selective membrane transport. This application explores the therapeutic potential for the human proton-coupled folate transporter (hPCFT), a recently discovered membrane transport system for folates and antifolates which is functionally and (to some degree) anatomically distinct from the ubiquitously expressed reduced folate carrier (RFC), the major tissue folate transporter. We hypothesize that hPCFT represents a novel and selective means for therapeutic targeting small molecule cytotoxins that are not transported by RFC. This concept is based on frequent and high level hPCFT expression in many human solid tumors, the acidic pH optimum for hPCFT vis ` vis RFC which parallels the pH microenvironments of solid tumors, and, most significantly, identification of the first small molecule cytotoxins that are selectively transported by hPCFT but not RFC. For analogs AG17, AG71, and AG94, in vitro cytotoxicities resulted from hPCFT transport and inhibition of de novo purine biosynthesis at glycinamide ribonucleotide formyltransferase. Another cytotoxic hPCFT substrate, AG112, was a potent inhibitor of thymidylate synthase. AG71 was tested in vivo against subcutaneous human hepatoma cells in SCID mice and showed potent hPCFT-targeted activity without significant toxicity. The goal of this project is to develop a new generation of potent tumor-targeted chemotherapy agents based on their selective capacities for cellular uptake by hPCFT over RFC. In aim 1, we will synthesize novel bicyclic and tricyclic analogs from 22 series of compounds, based on structure-activity profiles for AG17, AG71, AG94 and AG112, and molecular modeling with modifications of the ring systems, the linker domain, and the terminal glutamate. In aim 2, we test compounds from aim 1 for cytotoxicity in isogenic hamster and human tumor cell line models with established patterns of RFC and hPCFT expression, identify molecular targets by nucleoside protection, in situ metabolic labeling and analysis of intracellular metabolites, and studies with isolated enzymes. Additional studies will characterize transport properties of the novel analogs with hPCFT vis a vis RFC, and metabolism to polyglutamates. Finally, in aim 3, we will evaluate in vivo efficacies of the most potent hPCFT-targeted analogs by in vivo toxicity and efficacy trials in hPCFT-expressing human tumors implanted into SCID mice. Our results will define a comprehensive structure-activity relationship for transport by hPCFT vis a vis RFC and afford optimized analogs with potent and selective antitumor activities against hPCFT-expressing tumors in vitro and in vivo. This study will define mechanisms of action of the novel hPCFT-targeted cytotoxic analogs and potentially provide agents to be used clinically, albeit with a different spectrum of antitumor activities and reduced toxicities than those currently in use.