Research Project
8523398
DESCRIPTION (provided by applicant): Following systemic administration, most anticancer drugs exhibit very limited uptake into the brain. The blood- brain barrier (BBB) is the single most important factor for restricting drug delivery to the brain. The BBB is formed by a continuous monolayer of cerebrovascular endothelial cells joined together by tight junctions and lacks fenestrations. To cross the BBB, solutes must diffuse across the lipid endothelial cell membrane or be carried across either by receptor-mediated transport or by membrane transport. The capillary endothelial cells of the BBB express an array of uptake and efflux transporters. Regulated expression and function of these transporters govern access of essential nutrients and therapeutic drugs to the central nervous system (CNS). Successful drug delivery to brain tumors includes not only penetration of the BBB, but also delivery to and uptake into tumor cells. The Large Amino Acid Transporter 1 (LAT1) fulfills all necessary requirements - expression at both sides of the BBB, high expression in almost all tumor cells regardless of the tissue of origin, and very low endogenous expression in non-proliferative cells. Furthermore, it is characterized by an appreciable capacity, which can be exploited to deliver therapeutically meaningful amounts of chemotherapeutic agents. LAT1 activity correlates with cell proliferation, tumor growth, and is inversely related to disease prognosis and patient survival. Quadriga BioSciences will use NIH/NCI funding to engineer novel anti-cancer agents by combining the properties of cytotoxic agents with LAT1-mediated delivery to cancerous cells. The aims of Phase 1 of the proposed SBIR research are geared to determine whether LAT1-transported anticancer agents are a viable option for the development of chemotherapeutics to treat brain tumors. Success of our strategy is defined as activity and high specificity for LAT1. In Aim 1 a set of compounds will be synthesized that are hybrid compounds in which a known cytotoxic moiety is chemically fused to a LAT1- specific recognition element. These compounds will be tested for LAT1 activity and specificity in Aim 2. LAT1 transport will be determined in vitro using [3H]-gabapentin uptake competition in HEK cells and direct uptake coupled with LC/MS/MS detection. The specificity for LAT1 vs. other amino acid transporters will be tested by [3H]-phenylalanine uptake competition in cells expressing those transporters. Possible BBB transport of the compounds will be assessed by and in vitro transwell transport assay. In Aim 3, cytotoxicity (LD50 and LD100) of compounds toward cancer cell lines and bone marrow progenitor cells is evaluated. To exclude that test compounds enter the cell via means other than LAT1, the effect of LAT1 inhibitors on the cytotoxicity will be determined. A successful compound will be cytotoxic to LAT1-expressing cancer cell lines through selective accumulation and exhibit no cytotoxicity to bone marrow progenitor cells. In projected Phase 2 studies, successful compounds will be tested in vivo in validated xenograft models for various brain tumors.
