A biosensor for detection of HIV fusion inhibitors
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: The development of highly active antiretroviral therapy (HAART) for the treatment of HIV-infected patients has greatly improved the outcome and quality of life for these individuals. However, severe side effects in some patients and the development of multi-drug resistant strains of HIV are challenging us to develop alternative therapies which target different regions of the virus. In this proposal, we target the fusion mechanism of HIV, whereby it injects its genetic material into the host cell. Currently there is a single fusion inhibitor on the market called Enfuvirtide®, which is used as a salvage therapy in cases where patients do not respond well to HAART medications. As a peptide drug, Enfuvirtide is of limited use because it is very expensive and must be given by injection. The goal of our project is to identify novel classes of fusion inhibitors by developing a method to rapidly screen chemical databases for molecules with fusion inhibition properties. In particular, we will look for molecules which have the ability to bind to the same viral target as Enfuvirtide-like peptides. This target is a rod-like structure consisting of three helical domains wrapped into a supercoiled helix bundle. We will devise a novel strategy for our screen using protein immobilized on a chip as a biosensor for detecting fusion inhibition properties among drug candidates. Our protein will be a specially designed HIV-1 fusion domain which is able to recognize both peptide and non-peptide fusion inhibitors. It consists of a three-stranded coiled coil structure, capped by a metal binding site which promotes the correct folding of the protein and introduces redox properties to the system. The protein will be coated on a gold-plated quartz crystal, and measured using an electrochemical quartz crystal microbalance. Mass changes, redox changes and / or electron transfer changes associated with the binding of known peptide inhibitors will be used to calibrate the system for the eventual detection of as yet unknown inhibitors from a library. The specific aims of this proposal are 1) to generate a biosensor device by developing a stable protein film of the metallopeptide coiled coil structure oriented on a gold electrode chip; 2) to measure the binding event by immersing the biosensor in a solution containing known binders and detecting accompanying mass and electrochemical changes to the system. The sensitivity and range of the resulting assay will be calibrated using peptides of varying affinities to establish positive and negative controls, and the limits of detection. This technology may be used for rapid or “high-throughput” screening of chemical libraries. Hundreds of thousands of chemicals are publicly available for screening at NIH and at various screening centers around the country including the Bay Area Screening Center at the California Institute for Quantitative Biomedical Research.