Due to the emergence of drug-resistant strains and the cumulative toxicities associated with current therapies, demand remains for new inhibitors of HIV-1 replication. The HIV-1 matrix (MA) protein is an essential viral component with established roles in the assembly of the virus. Using virtual and surface plasmon resonance (SPR)-based screening, we describe the identification of the first small molecule to bind to the HIV-1 MA protein and to possess broad range anti-HIV properties.
We have identified a novel compound (7) that binds to and functions through the conserved PI(4,5)P2 binding site on HIV-MA, inhibits the production of new virus, and exhibits broad spectrum anti-HIV activity with IC50 values in the range of 7.5 to 15.6 mm for the group M isolates tested. This compound provides a novel chemical probe to investigate the roles of MA in the HIV-1 replication cycle and could serve as a good starting point for the development of a new class of HIV therapeutics through optimization by medicinal chemistry approaches.
To identify this compound, we undertook a structure-guided search for small, drug-like molecules that inhibit viral replication by binding to this highly conserved area on the HIV-1 MA protein by using a virtual screening protocol.
First, we tested whether the identified compounds could interact with MA using surface plasmon resonance (SPR). And oone of the best compounds interacts with HIV-1 MA with an equilibrium dissociation constant of 171 μM. To identify other potential hits that were not identified in the SPR-based screen, but still represent matrix-targeted inhibitors of HIV-1 replication, we evaluated the 19 compounds identified from the initial virtual screen. This led to the discovery of the novel antiviral compounds, as well as confirmation of the first compound as a genuine hit. The lead compound from this analysis, compound 7, inhibited the replication of the primary isolate HIV-189BZ167 with the lowest IC50 value 12.3 μM, showed minimal toxicity(>100 μM), and did not inhibit the replication of a panel of non-retroviruses. Compound 7 inhibited the replication of all the group M viruses tested (panel of HIV-1 clinical isolates from different geographic locations that included HIV-1 group M subtypes A–G).
To explore the interaction of compound 7 with HIV-1 MA, we employed SPR. Compound 7 interacts with HIV-1 MA with an equilibrium dissociation constant (KD) of 22.6 μM (Figure 1) This value is in remarkable agreement with the IC50 value derived from the single-round infection assay (IC50 21.2 μM).

Sensorgrams depicting the interaction of compound 7 with immobilized HIV-1lAI MA. Compound concentrations of 0, 6.25, 12.5, 25, 50, and 100 mm were tested and are represented as orange, magenta, green, blue, cyan and red lines, respectively. The insert shows the equilibrium plot used to derive the equilibrium dissociation constant (KD). Compound 7–MA interaction: KD=22.6 uM.
The effect of compound 7 on the production of both wild-type (WT) and the L21A and T81A mutant viruses was assessed by p24 content in
the supernatants after 48 h. Compound 7 decreased the amount of WT virus produced to approximately 392.5% of the control with no compound (DMSO only). However, changing residues L21 and T81 to alanine completely abrogated the effects of compound 7.
We next sought to refine the docking model and to identify the most probable binding mode of compound 7 to see whether or not the model could offer any insight into the mutational results.Exhaustive dockings to multiple conformations of HIV-1 MA resulted in the binding pose shown in Figure 2.

Refined predicted binding mode of compound 7 in the PI(4,5)P2 binding site of HIV-1 MA. Binding site residues (L21, R22, W36, R76, T81, and K98) are represented as dark sticks, whereas compound 7 is shown in green sticks. Residues highlighted in blue are thought to be critical for compound binding. Yellow dashed lines represent hydrogen bonds
The results were published in two papers:
- Bioorg Med Chem Lett. 2012 Nov 29. pii: S0960-894X(12)01493-X. doi: 10.1016/j.bmcl.2012.11.041. [Epub ahead of print] Discovery of a small-molecule antiviral targeting the HIV-1 matrix protein. Zentner I, Sierra LJ, Maciunas L, Vinnik A, Fedichev P, Mankowski MK, Ptak RG, Martín-García J, Cocklin S. [PMID:23305922]
- ChemMedChem 2013 [Epub ahead of print] Identification of a small-molecule inhibitor of HIV-1 assembly that targets the phosphatidylinositol (4,5)-bisphosphate binding site of the HIV-1 matrix protein. Zentner I, Sierra LJ, Maciunas L, Vinnik A, Fedichev P, Mankowski MK, Ptak RG, Martín-García J, Cocklin S. [Manuscript number: cmdc.201200577]





