Development of a Novel Therapeutic Strategy Against HIV-1 by Simultaneously Targeting IN/INI1 and IN/RNA Interactions

About This Grant

Project Summary/Abstract This proposal seeks to develop a new class of HIV-1 inhibitors that disrupt critical intracellular host–virus protein– protein and viral protein–RNA interactions. Anti-HIV therapeutics targeting essential host–virus interactions are expected to reduce drug resistance relative to drugs targeting viral proteins alone, as host factors mutate less readily and resistant viral mutants may lose host binding capacity, thus becoming non-infectious. The long-term goal of this project is to develop novel antiretrovirals that target interactions between HIV-1 integrase (IN) and the host factor Integrase Interactor-1 (INI1), which may also dually disrupt IN interactions with viral RNA due to structural mimicry between the IN-binding domain of INI1 (Rpt1) and TAR RNA. IN is essential for viral cDNA integration and also regulates late stages of replication through interactions with TAR RNA and host proteins including INI1. Cells lacking INI1 fail to produce HIV-1 particles, and viruses encoding IN mutants defective for INI1 or RNA binding produce morphologically defective, noninfectious virions. Structural studies reveal that the IN-binding domain of INI1 mimics TAR RNA, explaining shared binding interfaces and phenotypes. We hypothesize that inhibitors disrupting IN/INI1 interactions will also disrupt IN/RNA binding, leading to defective particle formation and inhibition of HIV-1 replication. As a proof-of-concept, we developed a hydrocarbon-stapled helical peptide derived from the INI1 helix-1 that blocks both IN/INI1 and IN/RNA interactions in vitro, potently inhibits HIV-1 replication, and induces the production of morphologically defective, noninfectious particles. Building on this premise, we aim to identify small-molecule inhibitors of IN/INI1 and IN/RNA interactions. High-throughput virtual screening of the NCI therapeutic library for compounds that dock onto IN/INI1 interface and identified glycyrrhizic acid (GL) as a top candidate. GL is a natural compound with reported anti-HIV-1 activity in vitro and in patients, previously implicated in inhibiting viral entry via membrane modulation. We hypothesize that GL inhibits HIV-1 replication through dual mechanisms: disruption of IN/INI1 and IN/RNA interactions during late events and inhibition of viral entry during early events. In Aim 1, we will characterize GL-mediated inhibition of IN/INI1 and IN/RNA interactions in vitro and in vivo. In Aim 2, we will define GL’s mechanism of action during early and late replication stages of HIV-1 by analyzing particle production, Gag processing, virion morphology, viral entry, reverse transcription, nuclear import, and integration. We will also isolate and characterize GL- resistant viruses and assess GL’s activity against ART-resistant strains. Completion of this project will establish GL as a lead compound for a novel class of dual-targeting HIV-1 inhibitors with the potential to reduce resistance while providing comprehensive training in virology, molecular biology, mechanistic analysis, and antiviral drug discovery.