The blood-brain barrier (BBB) prevents entry of most drug molecules into the brain, an obstacle which adds time and cost to the discovery and development of central nervous system (CNS) drugs. The BBB possesses transporter proteins that allow biological molecules including amino acids, glucose, and hormones to cross as needed. One of these transporter proteins, the L-type amino acid transporter (LAT-1) transports specific amino acids (e.g. phenylalanine) as well as compounds that resemble them (e.g. the drug L-DOPA). This transporter offers a potential route for targeted drug delivery to the brain; moreover, it has been demonstrated that drugs chemically linked to amino acids can cross the BBB via LAT-1. It is known that some molecules are transported (substrates), while others only bind to the transporter surface (inhibitors); however, the structural requirements for either type of activity is poorly understood. We propose mapping the structure-activity relationship (SAR) for molecules that interact with LAT-1 with the aspiration of making this transporter a practical and widely used vehicle for drug delivery. Our approach involves using a computational model of the LAT-1 binding site to guide the synthesis of compounds similar in structure to natural amino acids but with unique chemical properties that could be used to design improved drug delivery agents.
Our specific aims are three-fold. First, we will synthesize amino acids chemically substituted with groups that our model predicts will form beneficial interactions with the transporter. Second, we will use cell-based assays to test these compounds. To determine whether they are transported, we will use cis-inhibition and trans-stimulation cell assays which have successfully identified previously unknown substrates. SAR from these experiments will be used to refine our computational model in order to generate hypotheses to guide further substrate optimization. The intracellular concentration of selected compounds will be analyzed by LC-MS/MS to verify our hypothesis that LAT-1 activity increases cell uptake.
A third aim of this project will involve the synthesis and testing of amino acid prodrugs of a topoisomerase I inhibitor for cancer (SN-38) to assess whether the SAR derived from optimization of LAT-1 substrates in specific aims 1 and 2 can be applied to improve prodrug cellular uptake. Additionally, the effect of the linker joining SN-38 and the amino acid promoiety, which may have a significant impact on prodrug pharmacokinetics and efficacy, will be examined. This project has broad applications for treating many different diseases both in the brain and other tissues where LAT-1 is heavily expressed, including cancer.
One of the challenges for treating brain disorders is designing drugs that are capable of crossing the blood-brain barrier (BBB). We seek to exploit one of the BBB?s natural mechanisms for transporting amino acids, the LAT-1 protein, to use it for drug delivery into the brain and other tissues (e.g. cancer) that express this transporter. We will attempt to optimize the structure and chemical properties of amino acid mimetics to provide effective drug delivery agents for brain cancer.
| Chien, Huan-Chieh; Colas, Claire; Finke, Karissa et al. (2018) Reevaluating the Substrate Specificity of the L-Type Amino Acid Transporter (LAT1). J Med Chem 61:7358-7373 |
