Structure-Based Design, Synthesis, and Characterization of Dual Hotspot Small-Molecule HIV-1 Entry Inhibitors

Abstract

Cellular infection by HIV-1 is initiated with a binding event between the viral envelope glycoprotein gp120 and the cellular receptor protein CD4. The CD4–gp120 interface is dominated by two hotspots: a hydrophobic gp120 cavity capped by Phe43CD4 and an electrostatic interaction between residues Arg59CD4 and Asp368gp120. The CD4 mimetic small-molecule NBD-556 (1) binds within the gp120 cavity; however, 1 and related congeners demonstrate limited viral neutralization breadth. Herein, we report the design, synthesis, characterization, and X-ray structures of gp120 in complex with small molecules that simultaneously engage both binding hotspots. The compounds specifically inhibit viral infection of 42 tier 2 clades B and C viruses and are shown to be antagonists of entry into CD4-negative cells. Dual hotspot design thus provides both a means to enhance neutralization potency of HIV-1 entry inhibitors and a novel structural paradigm for inhibiting the CD4–gp120 protein–protein interaction.

CD4 mimetics sensitize HIV-1-infected cells to ADCC

“…sCD4 and the miniprotein M48U1 were produced and purified as previously described (26,52). The CD4-mimetic small molecules JP-III-48 and DMJ-I-228 were synthesized as described previously (20,21). The compounds were analyzed, dissolved in dimethyl sulfoxide (DMSO) at a stock concentration of 10 mM, aliquoted, and stored at −20°C.…”

“…These epitopes become exposed on virions only on the interaction of Env trimers with host CD4, indicating that binding membrane-anchored CD4 provides an additional energy component that is not provided by sCD4 (35). Rationally designed CD4mc (JP-III-48, DMJ-I-228) engage gp120 within the Phe-43 cavity (22) and can act as CD4 agonists, inducing thermodynamic changes in the Env trimer more similar to those observed during membrane CD4 binding (20,24). Importantly, compounds of this class have been shown to sensitize HIV-1 particles to neutralization by CD4i and V3 nonneutralizing vaccine-elicited Abs (25).…”

“…The prototypes of such compounds, NBD-556 and NBD-557, were discovered in a screen for inhibitors of gp120-CD4 interaction (19). These small-molecule ∼337-Da compounds and recent derivatives (DMJ-I-228, JP-III-48) bind in the Phe-43 cavity (20)(21)(22), a highly conserved ∼150-Å 3 pocket in the gp120 glycoprotein located at the interface of the inner domain, outer domain, bridging sheet, and CD4 receptor (23). CD4mc block gp120-CD4 interaction and induce thermodynamic changes in gp120 similar to those observed during CD4 or soluble CD4 (sCD4) binding (24).…”

Short Communication: Synergism Between a CD4-Mimic Peptide and Antibodies Elicited by a Constrained V3 Peptide

“…8,[12][13][14] However, the low potencies of small organic CD4-mimetics, problems associated with administering CD4-mimetic peptides, and the potential of enhancing infection of CD4 -, CCR5 + , or CXCR4 + cells has thus far considerably limited enthusiasm for this approach.…”

“…18,19 Soluble CD4-mimetic compounds such as CD4M33 and the small organic molecule NBD-556 and improved variants of these compounds have been considered as potential anti-HIV-1 therapeutics or prophylactic agents. 8,13,19,20 Here we investigated the capacity of CD4M33 to act in synergy with polyclonal sera that contain a high level of V3-directed neutralizing antibodies raised against the optimally constrained C4-V3 T303C-E322C peptide.…”