A small molecule HIV-1 inhibitor that targets the HIV-1 envelope and inhibits CD4 receptor binding

Abstract

BMS-378806 is a recently discovered small molecule HIV-1 inhibitor that blocks viral entrance to cells. The compound exhibits potent inhibitory activity against a panel of R5-(virus using the CCR5 coreceptor), X4-(virus using the CXCR4 coreceptor), and R5/X4 HIV-1 laboratory and clinical isolates of the B subtype (median EC50 of 0.04 microM) in culture assays. BMS-378806 is selective for HIV-1 and inactive against HIV-2, SIV and a panel of other viruses, and exhibits no significant cytotoxicity in the 14 cell types tested (concentration for 50% reduction of cell growth, >225 microM). Mechanism of action studies demonstrated that BMS-378806 binds to gp120 and inhibits the interactions of the HIV-1 envelope protein to cellular CD4 receptors. Further confirmation that BMS-378806 targets the envelope in infected cells was obtained through the isolation of resistant variants and the mapping of resistance substitutions to the HIV-1 envelope. In particular, two substitutions, M426L and M475I, are situated in the CD4 binding pocket of gp120. Recombinant HIV-1 carrying these two substitutions demonstrated significantly reduced susceptibility to compound inhibition. BMS-378806 displays many favorable pharmacological traits, such as low protein binding, minimal human serum effect on anti-HIV-1 potency, good oral bioavailability in animal species, and a clean safety profile in initial animal toxicology studies. Together, the data show that BMS-378806 is a representative of a new class of HIV inhibitors that has the potential to become a valued addition to our current armamentarium of antiretroviral drugs.

Fig. 1.

(A) The structure of BMS-378806. (B) Time-of-addition assay. HeLa CD4 CCR5 cells were infected with HIV-1_JRFL pseudotyped virions, encoding a luciferase reporter gene. At the times indicated, either 50 nM BMS-378806 (▪) or 1 μM nevirapine (▴) (10× EC₅₀) was added. At 4 h, the virus was removed and only the media and compounds were put back. Antiviral activity was measured after 48 h by monitoring the luciferase activity.

Fig. 2.

Mode-of-action studies on BMS-378806. (A) Inhibition of gp120/CD4 binding by BMS-378806. The dose-dependent inhibition of sCD4 binding to gp120 by BMS-378806 gave an IC₅₀ of ≈100 nM, indicating that the compound interferes with gp120/CD4 binding. (B and C) Gel filtration analysis of [³H]BMS-378806 binding to gp120. [³H]BMS-378806 was mixed with the proteins shown and then centrifuged through a Micro BioSpin 6 gel filtration column. Protein-bound radiolabel was quantitated in the eluent by liquid scintillation counting. (B) Samples contained no protein (Blank), 500 nM gp120, 3 μM CD4, 500 nM gp120 and 3 μM CD4, or 25 μM BSA. (C) Samples contained 0–500 nM gp120. The amount of [³H]BMS-378806 bound was proportional to the amount of gp120 added.

Fig. 3.

Susceptibility histogram for subtype B clinical isolates and laboratory strains. BMS-378806 susceptibility was plotted against 42 clinical isolates and 11 laboratory strains of the HIV-1 B subtype. The x axis indicates EC₅₀ (in μM), with the left boundary of each bar listing the lower limit and the right boundary showing the upper limit of EC₅₀. The y axis represents the number of isolates that falls within a particular range of EC₅₀. The median EC₅₀ is 0.04 μM.