CROI 2016 Abstract eBook

Abstract Listing

Poster Abstracts

functional inhibitory profile of these Abs was defined using the well-standardized TZM-bl neutralization assay, the conventional neutralization assay on PBMCs, or the Fc-mediated inhibitory assay on macrophages. Results: We found that the anti-W614A-3S purified Abs display efficient and broad neutralizing activity. They inhibit transmitted founder Tier 2 viruses, neutralize primary isolates on primary cells and display Fc-mediated inhibitory functions at ng to µg/ml concentrations. The detection of anti-W614A-3S Abs was specifically correlated both with lower viral DNA (p<0.0001), viral load (p<0.0001), and other clinical parameters (CD4 + T cells, HLA protective alleles,…) suggesting that anti-W614A-3S neutralizing Abs participate in the control of HIV replication in ALT patients. Conclusions: These results demonstrate that ALT patients develop efficient neutralizing Abs that can be purified using W614A-3S mutant protein capture assays. These Abs are distinct to that recently isolated from ELITE neutralizer patients. Abs directed against W614A-3S may therefore be considered as a new family of broadly neutralizing Abs, which need to be further characterized, considering their potential role on viral load and viral DNA. 305 WITHDRAWN

306 Neutralization Differences Between CCR5 and CXCR4 Tropic Viruses in HIV-1B Infection

Ludy Registre ; Manish Sagar Boston Univ, Boston, MA, USA

Background: The mechanism for emergence of CXCR4 tropic HIV-1 variants remains poorly understood. We hypothesized that neutralizing antibodies (nAb) generated within an infected host potentially influence emergence of CXCR4-utilizing variants. We examined neutralization susceptibility among co-existing and inter-subject CCR5- and CXCR4-using viruses. Methods: Full-length envelopes (envs) were isolated from 21 individuals from ACTG study A5095 using single genome amplification. Envs were incorporated into an NL4-3 backbone to generate replication competent recombinant viruses. Viruses were examined for co-receptor usage and neutralization sensitivity using TZM-bl cells. Neutralization was assessed by measure of area under the curve (AUC), which assesses average neutralization within the range of two concentrations. Chimeric envelopes with swapped V3 loops were generated to determine the viral determinant influencing observed neutralization susceptibility differences. Comparisons were done using the Wilcoxon rank sum test. Results: A median of 13 envs (range 3-27) was isolated from 21 individuals. Exclusively CCR5-using envs (R5) were isolated from 12 individuals, envs able to utilize both CCR5 and CXCR4 (R5X4) and or only CXCR4 (X4) were isolated from 7 subjects, and a combination of R5, X4, and R5X4 envs were amplified from 2 individuals. Sequence analysis showed that all envs were subtype B. None of the R5 or R5X4 envs (n = 52), but all the X4 variants (n = 19) had a 2 – 3 amino acid insertion prior to the V3 loop GPGR crown. In the 2 subjects with co-circulating X4 and R5 variants, X4 envs were less neutralization sensitive to autologous contemporaneous plasma compared to the R5 envs (subject 1: R5 median AUC-0.46 vs. X4 AUC- 0.19; subject 2: R5 median AUC- 0.19 vs. X4 AUC- 0.0). In inter-subject comparisons, X4 as compared to R5 variants were more neutralization resistant to heterologous pooled plasma (p= 0.03). Conclusions: Similar to previously described subtype C HIV-1, subtype B env X4 variants possess a unique V3 loop genotypic signature. In some individuals, X4 variants potentially emerge as neutralization escape variants against the host generated nAbs. Chimeric envelope results will elucidate the role of the unique V3 amino acid insertion in conferring neutralization resistance. 307 Humoral Immune Pressure Selects for HIV-1 CXCR4-Using Variants Nina Lin 1 ; Oscar Gonzalez 1 ; Carlos Becerril 2 ; Behzad Etemad 2 ; Hong Lu 3 ; XuelingWu 3 ; Shahin Lockman 2 ; M. Essex 4 ; Daniel Kuritzkes 5 ; Manish Sagar 1 1 Boston Univ, Boston, MA, USA; 2 Brigham and Women’s Hosp, Harvard Med Sch, Boston, MA, USA; 3 Aaron Diamond AIDS Rsr Cntr, New York, NY, USA; 4 Harvard Sch of PH, Boston, MA, USA; 5 Harvard Med Sch, Boston, MA, USA Background: The emergence of a dual-mixed (DM) population (mixture of CCR5-using (R5), exclusively CXCR4-utilizing (X4), or R5X4 viruses) as opposed to presence of only R5 variants is associated with accelerated CD4+ T cells decline. Previous studies have not clarified if CXCR4-utilizing variants accelerate disease progression or if faster CD4+ T cell decline facilitates the emergence of DM virus populations. Methods: Seven individuals enrolled in the Mashi study with documented DM populations and not on treatment were selected for this study. Envelopes (envs) were isolated using bulk PCR cloning. Envs were used to generate pseudovirions, replication competent viruses, or codon optimized gp120s. Infectious viruses were examined for neutralization sensitivity using TZM-bl assay, coreceptor usage using U87 cells, and evolution using in-vitro passage in the presence or absence of autologous plasma. Envs using different coreceptors were compared using the Wilcoxon rank-sum test. Results: A median of 11 subtype C env clones (range 9 – 18) were examined from each individual. The X4 envs as compared to co-circulating CCR5-utilizing variants were less sensitive to neutralization by both contemporaneous autologous plasma (p = 0.03) and plasma pools from individuals that harbor only R5 HIV-1 (p = 0.12). The X4 and co-existing R5 envs, however, had similar sensitivity to two different broadly neutralizing antibodies (bNab), VRC01 and 10E8 (p > 0.05). Among 2 individuals with variants susceptible to env V3 directed bNab, PGT128, X4 as compared to co-circulating R5 variants were more neutralization resistant in 1 case (p = 0.003). Chimeric envs with X4 as compared to R5 V3 domains were less sensitive to pooled R5 plasma and PGT128, although the differences were relatively small. Chimeric gp120s with X4 as compared to R5 V3 segments also showed decreased binding to autologous plasma and V3 directed monoclonal antibody, 447-52D. In-vitro passage in activated CD4+ T cell cultures of a neutralization sensitive CCR5-using virus led to the emergence of a CXCR4-utilizing virus stock in 1 of 3 cases in the presence but not the absence of autologous plasma. Conclusions: In some individuals, X4 as compared to co-circulating R5 variants are relatively neutralization resistant to host generated antibodies that potentially target the env V3 loop. In these individuals, CXCR4-utilizing viruses likely emerge as a consequence of humoral immune pressure.

Poster Abstracts

118

CROI 2016