324 – Novel Strategy to Inhibit HIV by Targeting Integrase/INI1 Interaction Based on the TAR RNA Mimicry

Background

Targeting host-virus interactions is an attractive strategy to overcome the problem of HIV-1 drug resistance. INI1/SMARCB1 is an HIV-1 integrase (IN)-interacting host factor required for assembly and particle production. Studies from our laboratory has indicated that IN binding Rpt1 domain of INI1 (INI1-Rpt1) and viral TAR RNA structurally mimic each other and bind to the same surface on IN (Dixit et al. Nat. Comm. 2021). IN mutants defective for binding to INI1 are impaired for virion morphogenesis and infection. Because of the INI1-Rpt1 mimicry to TAR, we hypothesize that drugs that target IN-INI1 interaction will be “dual acting”, disrupt both IN-INI1 and IN-TAR interactions, and induce particle morphogenesis defects to inhibit HIV-1 replication.

Methods

We used structural information on INI1-Rpt1/IN-(C-terminal domain) CTD complex to generate hydrocarbon stapled peptides derived from interface α1 helix of INI1-Rpt1. These peptides were tested for their effect on IN-INI1, INI-RNA interactions using Alpha Assay and on HIV-1 replication using T-cell lines, PBMC and primary macrophages. The mechanism was tested by assessing their effect on late events and particle morphogenesis, using virology and transmission electron microscopy (TEM) analyses. 

Results

One of the stapled peptides, SP38, inhibited both IN-INI1 and IN-RNA interactions both in vitro (IC₅₀ of ~0.3 µM) and in cells. SP38 potently inhibited HIV-1 replication in T-cell line (by 4-5 logs), PBMCs and primary macrophages. SP38 was not cytotoxic and had no effect on expression, proteolytic processing of viral proteins or particle production. However, particles produced in the presence of SP38 were defective for infection, and were impaired for early and late reverse transcription, nuclear localization and integration. TEM analysis indicated that SP38 treatment resulted in morphologically defective “eccentric” virus particles. Furthermore, SP38 inhibited spread of the reactivated virus from latent cells.

Conclusions

INI1 α1 helix-derived stapled peptide SP38 dually targets IN-INI1 and IN-RNA interactions and potently inhibits HIV-1 replication by inducing defective particle morphology. These results validate our hypothesis, and provide a proof-of-concept for developing drugs derived from INI1 to simultaneously target IN/INI1 and IN/RNA interactions. SP38 represents a novel first-in-class HIV-1 inhibitor that targets host-virus interactions to inhibit HIV-1 particle morphogenesis.

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