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Enhanced antibody-mediated neutralization of HIV-1 variants that are resistant to fusion inhibitors

View Article: PubMed Central - PubMed

ABSTRACT

Background: HIV-1 typically develops resistance to any single antiretroviral agent. Combined anti-retroviral therapy to reduce drug-resistance development is necessary to control HIV-1 infection. Here, to assess the utility of a combination of antibody and fusion inhibitor treatments, we investigated the potency of monoclonal antibodies at neutralizing HIV-1 variants that are resistant to fusion inhibitors.

Results: Mutations that confer resistance to four fusion inhibitors, enfuvirtide, C34, SC34, and SC34EK, were introduced into the envelope of HIV-1JR-FL, a CCR5-tropic tier 2 strain. Pseudoviruses with these mutations were prepared and used for the assessment of neutralization sensitivity to an array of antibodies. The resulting neutralization data indicate that the potencies of some antibodies, especially of those against the CD4 binding site, V3 loop, and membrane-proximal external region epitopes, were increased by the mutations in gp41 that conferred resistance to the fusion inhibitors. C34-, SC34-, and SC34EK-resistant mutants showed more sensitivity to monoclonal antibodies than enfuvirtide-resistant mutants. An analysis of C34-resistant mutations revealed that the I37K mutation in gp41 HR1 is a key mutation for C34 resistance, low infectivity, neutralization sensitivity, epitope exposure, and slow fusion kinetics. The N126K mutation in the gp41 HR2 domain contributed to C34 resistance and neutralization sensitivity to anti-CD4 binding site antibodies. In the absence of L204I, the effect of N126K was antagonistic to that of I37K. The results of a molecular dynamic simulation of the envelope trimer confirmation suggest that an I37K mutation induces the augmentation of structural fluctuations prominently in the interface between gp41 and gp120. Our observations indicate that the “conformational unmasking” of envelope glycoprotein by an I37K mutation is one of the mechanisms of neutralization sensitivity enhancement. Furthermore, the enhanced neutralization of C34-resistant mutants in vivo was shown by its high rate of neutralization by IgG from HIV patient samples.

Conclusions: Mutations in gp41 that confer fusion inhibitor resistance exert enhanced sensitivity to broad neutralizing antibodies (e.g., VRC01 and 10E8) and other conventional antibodies developed in HIV-1 infected patients. Therefore, next-generation fusion inhibitors and monoclonal antibodies could be a potential combination for future regimens of combined antiretroviral therapy.

Electronic supplementary material: The online version of this article (doi:10.1186/s12977-016-0304-7) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

Effect of C34 resistance-conferring mutations on the sensitivities to MAbs against V3 and MPER. Three mutations that together confer C34 resistance, I37K, N126K, and L204I, were examined for their effect on the neutralization sensitivity to the anti-V3 MAbs, KD247, 16G6, and 0.5γ, (a) and to the anti-MPER MAb 10E8 (b) using single (left) and double (right) mutants. In each graph, the x axis represents the log concentration of MAbs in µg/ml and the y axis represents the percent inhibition compared with the corresponding no inhibitor control. The results are shown as the means ± SEs of three independent experiments
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Fig3: Effect of C34 resistance-conferring mutations on the sensitivities to MAbs against V3 and MPER. Three mutations that together confer C34 resistance, I37K, N126K, and L204I, were examined for their effect on the neutralization sensitivity to the anti-V3 MAbs, KD247, 16G6, and 0.5γ, (a) and to the anti-MPER MAb 10E8 (b) using single (left) and double (right) mutants. In each graph, the x axis represents the log concentration of MAbs in µg/ml and the y axis represents the percent inhibition compared with the corresponding no inhibitor control. The results are shown as the means ± SEs of three independent experiments

Mentions: Although C34r was more sensitive to anti-V3 antibodies than WT (Fig. 1), single and double mutations of C34r did not affect its sensitivity to any of the tested anti-V3 antibodies except 16G6 (Fig. 3a). Mutants with I37K and L204I showed a fivefold increase in their sensitivity to 16G6; however, the sensitivities to KD247 and 0.5γ were similar between WT and mutants. This suggests that the combination of three mutations in C34r may act synchronously to impact the neutralization profile of anti-V3 antibodies (Fig. 3a).Fig. 3


Enhanced antibody-mediated neutralization of HIV-1 variants that are resistant to fusion inhibitors
Effect of C34 resistance-conferring mutations on the sensitivities to MAbs against V3 and MPER. Three mutations that together confer C34 resistance, I37K, N126K, and L204I, were examined for their effect on the neutralization sensitivity to the anti-V3 MAbs, KD247, 16G6, and 0.5γ, (a) and to the anti-MPER MAb 10E8 (b) using single (left) and double (right) mutants. In each graph, the x axis represents the log concentration of MAbs in µg/ml and the y axis represents the percent inhibition compared with the corresponding no inhibitor control. The results are shown as the means ± SEs of three independent experiments
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC5037607&req=5

Fig3: Effect of C34 resistance-conferring mutations on the sensitivities to MAbs against V3 and MPER. Three mutations that together confer C34 resistance, I37K, N126K, and L204I, were examined for their effect on the neutralization sensitivity to the anti-V3 MAbs, KD247, 16G6, and 0.5γ, (a) and to the anti-MPER MAb 10E8 (b) using single (left) and double (right) mutants. In each graph, the x axis represents the log concentration of MAbs in µg/ml and the y axis represents the percent inhibition compared with the corresponding no inhibitor control. The results are shown as the means ± SEs of three independent experiments
Mentions: Although C34r was more sensitive to anti-V3 antibodies than WT (Fig. 1), single and double mutations of C34r did not affect its sensitivity to any of the tested anti-V3 antibodies except 16G6 (Fig. 3a). Mutants with I37K and L204I showed a fivefold increase in their sensitivity to 16G6; however, the sensitivities to KD247 and 0.5γ were similar between WT and mutants. This suggests that the combination of three mutations in C34r may act synchronously to impact the neutralization profile of anti-V3 antibodies (Fig. 3a).Fig. 3

View Article: PubMed Central - PubMed

ABSTRACT

Background: HIV-1 typically develops resistance to any single antiretroviral agent. Combined anti-retroviral therapy to reduce drug-resistance development is necessary to control HIV-1 infection. Here, to assess the utility of a combination of antibody and fusion inhibitor treatments, we investigated the potency of monoclonal antibodies at neutralizing HIV-1 variants that are resistant to fusion inhibitors.

Results: Mutations that confer resistance to four fusion inhibitors, enfuvirtide, C34, SC34, and SC34EK, were introduced into the envelope of HIV-1JR-FL, a CCR5-tropic tier 2 strain. Pseudoviruses with these mutations were prepared and used for the assessment of neutralization sensitivity to an array of antibodies. The resulting neutralization data indicate that the potencies of some antibodies, especially of those against the CD4 binding site, V3 loop, and membrane-proximal external region epitopes, were increased by the mutations in gp41 that conferred resistance to the fusion inhibitors. C34-, SC34-, and SC34EK-resistant mutants showed more sensitivity to monoclonal antibodies than enfuvirtide-resistant mutants. An analysis of C34-resistant mutations revealed that the I37K mutation in gp41 HR1 is a key mutation for C34 resistance, low infectivity, neutralization sensitivity, epitope exposure, and slow fusion kinetics. The N126K mutation in the gp41 HR2 domain contributed to C34 resistance and neutralization sensitivity to anti-CD4 binding site antibodies. In the absence of L204I, the effect of N126K was antagonistic to that of I37K. The results of a molecular dynamic simulation of the envelope trimer confirmation suggest that an I37K mutation induces the augmentation of structural fluctuations prominently in the interface between gp41 and gp120. Our observations indicate that the “conformational unmasking” of envelope glycoprotein by an I37K mutation is one of the mechanisms of neutralization sensitivity enhancement. Furthermore, the enhanced neutralization of C34-resistant mutants in vivo was shown by its high rate of neutralization by IgG from HIV patient samples.

Conclusions: Mutations in gp41 that confer fusion inhibitor resistance exert enhanced sensitivity to broad neutralizing antibodies (e.g., VRC01 and 10E8) and other conventional antibodies developed in HIV-1 infected patients. Therefore, next-generation fusion inhibitors and monoclonal antibodies could be a potential combination for future regimens of combined antiretroviral therapy.

Electronic supplementary material: The online version of this article (doi:10.1186/s12977-016-0304-7) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus