Discovery of novel 1,4-disubstituted 1,2,3-triazole phenylalanine derivatives as HIV-1 capsid inhibitors

Abstract

The HIV-1 capsid (CA) protein plays crucial roles in both early and late stages of the viral life cycle, which has intrigued researchers to target it to develop anti-HIV drugs. Accordingly, in this research, we report the design, synthesis and biological evaluation of a series of novel phenylalanine derivatives as HIV-1 CA protein inhibitors using the Cu(I)-catalyzed azide and alkyne 1,3-dipolar cycloaddition (CuAAC) reaction. Among this series of inhibitors, compound II-10c displayed a remarkable anti-HIV activity (EC₅₀ = 2.13 μM, CC₅₀ > 35.49 μM). Furthermore, surface plasmon resonance (SPR) binding assays showed that compounds II-10c and PF-74 (lead compound) have similar affinities to HIV-1 CA monomer. Further investigation showed that the weak permeability and water solubility of representative compounds were probably the important factors that restricted their cell-based activity. Preliminary structure-activity relationships (SARs) were inferred based on the activities of these compounds, and their known structure. The most promising new compound was studied with molecular dynamics simulation (MD) to determine the preferred interactions with the drug target. Finally, the activities of members of this series of inhibitors were deeply inspected to find the potential reasons for their anti-HIV-1 activity from various perspectives. This highlights the important factors required to design compounds with improved potency.

Keywords: CuAAC; HIV-1; HIV-1 capsid protein; Molecular dynamics simulation; Phenylalanine derivatives; SAR; Surface plasmon resonance.

Fig. 1. The design of novel phenylalanine derivatives as HIV-1 CA inhibitors. (a) Structure of PF-74; (b) the binding mode of PF-74 in the NTD-CTD interface of CA protein hexamer. Yellow dashed lines indicate H-bond interactions. (c) Several reported phenylalanine derivatives as HIV-1 CA protein inhibitors. (d) Target compounds designed in this work.

Scheme 1. Reagents and conditions used to synthesize I-8(a–u). (i) 4-methoxy-N-methylaniline, PyBop, DIEA, CH₂Cl₂, r.t., 8–10 h; (ii): CF₃COOH; CH₂Cl₂, r.t., 5–6 h; (iii) propiolic acid, HATU, DIEA, CH₂Cl₂, r.t., 10 h; (vii): azide substituents; CuSO₄·5H₂O, VcNa, THF/H₂O (v : v = 1 : 1), 30–60 °C, 4–6 h.

Scheme 2. Reagents and conditions used to synthesize II-10(a–o). (i) 3-ethynylbenzoic acid, HATU, DIEA, CH₂Cl₂, r.t., 10 h; (ii): azide substituents; CuSO₄·5H₂O, VcNa, THF/H₂O (v : v = 1 : 1), 30–60 °C, 4–6 h.

Scheme 3. Reagents and conditions used to synthesize III-15(a–o). (i) 4-methoxy-N-methylaniline, PyBop, DIEA, CH₂Cl₂, r.t., 8–10 h; (ii): CF₃COOH; CH₂Cl₂, r.t., 5–6 h; (iii) 3-ethynylbenzoic acid, HATU, DIEA, CH₂Cl₂, r.t., 10 h; (vii): azide substituents; CuSO₄·5H₂O, VcNa, THF/H₂O (v : v = 1 : 1), 30–60 °C, 4–6 h.

Fig. 2. SPR isotherms of compound II-10c (a) binding to two variants of the CA protein (monomer and disulfide-stabilized hexamer) respectively with PF-74 as the reference (b). Isotherms are an average of 3 replicates with error bars indicating SEM.

Fig. 3. (a) RMSD (heavy atoms) of amino acids of CA HIV-1 monomer in reference to the first frame of the MD simulation. (b) RMSD (heavy atoms) of the bound II-10c in reference to the docked conformer.

Fig. 4. Binding interactions of II-10c in the first (a) and second (b) clusters.