Enhanced Exposure of Human Immunodeficiency Virus Type 1 Primary Isolate Neutralization Epitopes through Binding of CD4 Mimetic Compounds

Abstract

N-(4-Chlorophenyl)-N-(2,2,6,6-tetramethyl-piperidin-4-yl)-oxalamide (NBD-556) is a low-molecular-weight compound that reportedly blocks the interaction between human immunodeficiency virus type 1 (HIV-1) gp120 and its receptor CD4. We investigated whether the enhancement of binding of anti-gp120 monoclonal antibodies (MAbs) toward envelope (Env) protein with NBD-556 are similar to those of soluble CD4 (sCD4) by comparing the binding profiles of the individual MAbs to Env-expressing cell surfaces. In flow cytometric analyses, the binding profiles of anti-CD4-induced epitope (CD4i) MAbs toward NBD-556-pretreated Envexpressing cell surfaces were similar to the binding profiles toward sCD4-pretreated cell surfaces. To investigate the binding position of NBD-556 on gp120, we induced HIV-1 variants that were resistant to NBD-556 and sCD4 in vitro. At passage 21 in the presence of 50 M NBD-556, two amino acid substitutions (S375N in C3 and A433T in C4) were identified. On the other hand, in the selection with sCD4, seven mutations (E211G, P212L, V255E, N280K, S375N, G380R, and G431E) appeared during the passages. The profiles of the mutations after the selections with NBD-556 and sCD4 were very similar in their three-dimensional positions. Moreover, combinations of NBD-556 with anti-gp120 MAbs showed highly synergistic interactions against HIV-1. We further found that after enhancing the neutralizing activity by adding NBD-556, the contemporaneous virus became highly sensitive to antibodies in the patient's plasma. These findings suggest that small compounds such as NBDs may enhance the neutralizing activities of CD4i and anti-V3 antibodies in vivo.Human immunodeficiency virus type 1 (HIV-1) replicates continuously in the face of a strong antibody (Ab) response, although Abs effectively control many viral infections (3). Neutralizing Abs (NAbs) are directed against the HIV-1 envelope (Env) protein, which is a heterodimer comprising an extensively glycosylated CD4-binding subunit (gp120) and an associated transmembrane protein (gp41). Env proteins are present on the virion surface as "spikes" composed of trimers of three gp120-gp41 complexes (20,21,29). These spikes resist neutralization through epitope occlusion within the oligomer, extensive glycosylation, extension of variable loops from the surface of the complex, and steric and conformational blocking of receptor binding sites (16,18,20).Ab access to conserved regions is further limited because viral entry is a stepwise process involving conformational changes that lead to only transient exposure of conserved domains such as the coreceptor binding site (4, 5). However, some early strains of HIV-1 appear to be highly susceptible to neutralization by Abs (1, 10). For instance, subtype A HIV-1 envelopes from the early stage of infection exhibit a broad range of neutralization sensitivities to both autologous and heterologous plasma (1), suggesting that at least a subset of the envelopes have some preserved and/or exposed neutralization epitopes. It is well known that ...

Impact of the Maraviroc-Resistant Mutation M434I in the C4 Region of HIV-1 gp120 on Sensitivity to Antibody-Mediated Neutralization

“…Patient plasma IgGs and sera: Autologous plasma samples were collected and purified using Protein ASepharose (Affi-gel Protein A; Bio-Rad, Hercules, CA, USA) (22,30,33). The purified plasma IgGs were designated KP-5-IgG-1, -2, -5, and -7.…”

“…Antibodies and inhibitors: Anti-V3 NMAbs: KD-247 (27,28) and 447-52D (29), anti-CD4bs: 0.5d (3D6) (30,31) and b12 (32), and anti-CD4i: 4E9C (30) were used for neutralization susceptibility assays. KD-247 was kindly provided by the Chemo-Sero-Therapeutic Research Institute (Kumamoto, Japan).…”

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

“…8,9,30 Therefore, further improvement of the small organic CD4-mimetics is required to make them suitable for practical applications as preexposure prophylaxis. CD4-mimetic compounds have the potential to be used as an additional option to treat patients infected with HIV-1.…”

“…6,7 Importantly, from the perspective of the present study, peptides and small organic molecules can mimic sCD4 and expose the V3 loop for neutralization. 8,9 We recently designed a synthetic constrained V3 peptide immunogen that elicited an antibody response capable of neutralizing a spectrum of clade-B tier 1 strains differing in the sequences of the V3 b-strands. 10 This neutralizing antibody response was five times stronger than that elicited by a gp120 construct with deleted V1/V2 and 30-fold stronger in comparison with the corresponding linear V3 peptide.…”

“…9 CI values were determined for only two laboratory adapted viruses. Overall, the CI values of 0.174 and 0.043 for 50% and 75% inhibition, respectively, determined for NBD-556 are somewhat better than those determined for CD4M33 in the present work.…”