HIV infection remains a major world health issue requiring new methods to treat AIDS. Integrase (IN) is a virus-
encoded enzyme that is essential for retroviral replication and is an established target for the development of
drugs to treat HIV/AIDS. Five FDA-approved drugs that target the active site are in clinical use to treat AIDS
patients, but cross-resistant virus variants have been well documented. To combat resistance to existing IN
active site inhibitors, we need drugs that utilize a mechanism of action targeting other required viral protein
functions, such as multimerization or interaction with viral and cellular co-factors. A promising new approach is
centered on developing inhibitors targeting non-active site (or “allosteric”) locations, but none of these have been
approved for clinical use.
For proper function, integrase must form multimers competent to perform the coordinated insertion of two viral
DNA ends into the host target DNA, called concerted integration. We will test the potential for inhibiting proper
IN multimerization as an effective antiviral strategy. In addition to the well-characterized catalytic core dimer
interface, our studies have revealed a dimer interface mediated by interactions between the N-terminal and
catalytic core domains, which is also observed in the recent cryo-EM structure of HIV intasomes. The proposed
research targets this novel protein-protein interaction and proposes to characterize compounds that disrupt this
interface required for IN activity.
As a unique approach to identify allosteric inhibitors that target the protein-protein interactions required for
functional integrase multimerization, we designed a FRET-based assay that specifically detects formation of the
N-terminal domain-catalytic core domain dimer. In the first version of this assay, we identified several compounds
that also inhibited IN catalytic activities, and importantly two of these compounds inhibited HIV infection of cells
in culture. However, toxicity concerns limited the usefulness of these early hits. We have developed an improved,
second-generation screen and have identified 25 new hit compounds from specialized chemical libraries
designed to target protein-protein interactions. Eight of these new candidate compounds were deemed
acceptable by strict standards of drug-like qualities and possess strong medicinal chemistry potential for
advancement. The purpose of the proposed work is to: Aim 1 - Validate these hit compounds including a detailed
biochemical characterization; and Aim 2 - Test compounds for cell toxicity and the ability to block virus infection.
The information gained from the experiments in this application will have the potential for major impact in
development of anti-AIDS drugs that function as effective allosteric inhibitors of HIV IN, for use in combination
with existing drugs to prevent the emergence of resistant virus strains.