R01AI170206

RNA-Mimicry to Guide the Intra-Cellular Targeting of Host Virus Protein and Viral RNA-Protein Interactions to Inhibit HIV Replication.

Abstract:
The long-term goal of this application is to characterize host-virus interaction interface as a novel drug target and to develop inhibitors to disrupt intracellular protein-protein interactions (PPI) between the host and the virus to curb HIV-1 replication. It has been established that perturbing IN without affecting its enzymatic activity can inhibit late stages of HIV-1 replication such as assembly, particle production, and/or particle morphogenesis. Such class II IN mutations and allosteric inhibitors of IN (ALLINI) inhibit late events, and they do so by perturbing IN/IN multimerization, IN/host factor interaction, or IN/RNA interactions.

We have observed that such defects in particle morphogenesis can also be observed in IN mutants defective for interaction with a host factor, INI1/HSNF5, an IN-binding host factor that is selectively incorporated into HIV-1 virions. INI1 is required for HIV-1 late events. Expression of a minimal-IN-binding domain of INI1 (INI1183-292) termed S6 disrupts IN/INI1 interaction in vivo and potently inhibits HIV-1 particle production. Knocking down INI1 and use of INI1-/- cell lines inhibit HIV-1 particle production. INI1-binding defective IN mutants lead to the production of morphologically defective particles, indicating that targeting IN/INI1 interaction is an effective strategy to inhibit HIV-1 particle production. Lack of structure of INI1 and IN/INI1 complex has precluded our ability to develop inhibitors to target this interaction.

New research from our laboratory, including the NMR structure of the IN-binding repeat 1 (RPT1) domain of INI1, and molecular docking of IN/INI1 interaction, have helped overcome this knowledge gap. We found that the IN-binding domain of INI1, termed RPT1, and trans activating response element (TAR) of HIV-1 genomic RNA structurally mimic each other, a novel finding. Both RPT1 and TAR bind to the same surface of IN C-terminal domain (CTD) and compete for binding to IN with an identical IC50 value (0.005 μM). Furthermore, INI1-interaction-defective mutants of IN cause impairment of particle morphogenesis. We hypothesize that peptidomimetics and small molecules derived from RPT1 have dual activity and inhibit both IN/INI1 and IN/TAR interactions. As a proof of principle, we have developed a stapled peptide derived from interface A-1 helix of RPT1 that potently disrupts IN/INI1 and IN/RNA interactions, inhibits particle morphogenesis, and in vivo HIV-1 replication.

In this proposal, we will characterize IN/INI1 interface as an outstanding drug target by carrying out:
I) Genetic analysis to understand the mechanism of INI1 influence on assembly/particle production via trans-complementation and "synthetic rescue" experiments;
II) Development of a novel class of stapled peptides and small molecules with dual activity in targeting IN/INI1 and IN/RNA interactions by SAR and virtual-ligand screening; and determine the NMR structure of INI1-stapled peptide complexes with IN-C-terminal domain; and
III) Understanding the mechanism by which INI1-derived stapled peptides and small molecules inhibit HIV-1 replication and target identification by screening for viral escape mutants.

These studies will establish IN/INI1 as a novel drug target and provide new lead compounds to inhibit HIV-1 late events.

Albert Einstein College Of Medicine

NOT APPLICABLE

93.855 - Allergy and Infectious Diseases Research

National Institute of Allergy and Infectious Diseases (NIAID) [HHS - NIH]

Bronx, New York 10461 United States

Single Zip Code

NIH Research Project Grant (Parent R01 Clinical Trial Not Allowed) (PA-20-185)

Amendment Since initial award the total obligations have increased 420% from $614,093 to $3,190,267.