Differential inhibition of HIV-1 preintegration complexes and purified integrase protein by small molecules.

Abstract

To replicate, HIV-1 must integrate a cDNA copy of the viral RNA genome into a chromosome of the host. The integration system is a promising target for antiretroviral agents, but to date no clinically useful integration inhibitors have been identified. Previous screens for integrase inhibitors have assayed inhibition of reactions containing HIV-1 integrase purified from an Escherichia coli expression system. Here we compare action of inhibitors in vitro on purified integrase and on subviral preintegration complexes (PICs) isolated from lymphoid cells infected with HIV-1. We find that many inhibitors active against purified integrase are inactive against PICs. Using PIC assays as a primary screen, we have identified three new anthraquinone inhibitors active against PICs and also against purified integrase. We propose that PIC assays are the closest in vitro match to integration in vivo and, as such, are particularly appropriate for identifying promising integration inhibitors.Following binding of HIV-1 to a sensitive cell, the viral and cellular membranes fuse and the viral core particle is released into the cytoplasm. There the viral genomic RNA is reverse transcribed, yielding a double-stranded DNA copy of the viral RNA genome. Next, the complex of viral cDNA and proteins, the "preintegration complex" (PIC), migrates to the nucleus and covalently attaches the viral cDNA to a chromosome of the host. Integration completes the formation of a provirus, which contains all the information necessary to direct the synthesis of the viral RNAs and proteins required for the formation of new virions (for reviews, see refs. 1 and 2). Fig. 1A summarizes the DNA breaking and joining reactions involved in integration in vivo. The blunt ends of the linear product of reverse transcription are cleaved to remove two nucleotides from each 3' end. The recessed 3' ends are then joined to protruding 5' ends of breaks made in the target DNA to yield an integration intermediate (Fig. 1A, II). The points of joining of the two DNA strands are staggered by five base pairs. The DNA between the two points of joining then melts, yielding gaps at each junction between host and viral DNA.

A Novel Aspochalasin with HIV-1 Integrase Inhibitory Activity from Aspergillus flavipes

“…Whilst the cytochalasins identified in this study are able to inhibit the activity of integrase in in vitro enzyme assays, in their current forms they are not able to inhibit the replication of HIV-1. This is not uncommon amongst integrase enzyme inhibitors with many unable to inhibit the enzyme within a preintegration complex or within the cell [18,19].…”

Triazine Dyes Inhibit HIV‐1 Entry by Binding to Envelope Glycoproteins

“…Furthermore, Evans blue was found to partially inhibit gp120-CD4 interaction, and Chicago blue sky was shown to impede virus infection by inhibiting the virus from cell interaction (5,8,19). It was also described that CB3GA did not inhibit integrase activity of subviral preintegration complexes, though it inhibited the activity of purified enzymes (9).…”

HIV-1 integrase inhibitors that compete with the target DNA substrate define a unique strand transfer conformation for integrase

“…Integrase inhibitors that disrupt earlier steps in integration (e.g., assembly) have thus far not proved to be effective inhibitors of preintegration complexes or of viral replication (34)(35)(36). The DKAs are the first inhibitors of integrase whose antiviral activity clearly results from inhibition of integration (17).…”