Defining HIV Env protein expression in latently infected cells

点击翻译按钮获取中文摘要

Project Summary/Abstract Advances in B cell biology and molecular virology have enabled the discovery, characterization, and commercial development of broadly neutralizing antibodies (bNAbs). They are a promising immunotherapy that can be incorporated in many strategies for the treatment and/or cure of HIV-1. Antibodies have many effector functions beyond neutralization and their most important mechanism of action for treatment/cure may be their ability to opsonize infected cells, tagging them for antibody-dependent cellular cytotoxicity or phagocytosis (ADCC/ADCP) by immune effector cells. It is not clear that bNAbs are mediating clearance of infected cells in clinical trials when they are passively infused into chronically infected people living with HIV (PLWH). An obvious reason for this lack of efficacy of bNAbs may be a lack of Env protein expression while on suppressive ART. Our lack of knowledge about the level of Env expression during HIV latency and latency reversal prevents effective use of bNAbs as therapeutics. To address this gap, the ability to detect low level Env protein expression in cells infected with different subtypes of HIV-1 is needed. The Bosque lab has published an ultrasensitive method to detect p24 Gag protein down to the fg/ml level and has preliminary data for a newly developed Env assay. Here we propose to use these assays to detect low levels of Env and Gag protein in a well-described model of latency and compare the data to Env surface expression as measured by flow cytometry (Aim 1). Within these assays, we have the flexibility to test Env detection using bNAbs targeting different epitopes to probe for Env conformation (i.e., trimer, monomer, etc) and extend these analyses to diverse HIV isolates from different subtypes to measure differences in Env expression and latency attributable to genetically diverse virus (Aim 2). Knowing these crucial factors about Env and Gag expression in a carefully controlled model of latency will allow us to further investigate HIV protein translation in future studies using cohorts of PLWH. These studies will improve the ability of therapeutics using Env-targeting strategies to target latently infected cells.