CROI 2019 Abstract eBook

Abstract eBook

Poster Abstracts

310 A NOVEL PHAGE-DISPLAY APPROACH MAPS LINEAR EPITOPES OF GP41- SPECIFIC MABS Meghan E. Garrett , Katherine L. Williams, Theodore Gobillot, Ryan Basom, Megan M. Stumpf, Katharine Dusenbury, Julie M. Overbaugh Fred Hutchinson Cancer Research Center, Seattle, WA, USA Background: Antibodies targeting the HIV Envelope (Env) protein can limit viral infection and provide immunity. However, the field has yet to generate a vaccine capable of producing protective antibodies in humans. With epitope- based vaccine design we can engineer immunogens to elicit antibodies against specific regions of Env, but we are limited in part by the low throughput nature of current epitope mapping methods. Here we developed and tested a high- throughput, comprehensive approach to map the epitopes of recently identified HIV-specific monoclonal antibodies (mAbs) that mediate ADCC. Methods: We applied a phage display method that used deep sequencing, Phage Immunoprecipitation-Sequencing (PhIP-seq), to identify the linear epitopes of four newly identified gp41-specific monoclonal antibodies (mAbs): QA255.006, QA255.016, QA255.067, and QA255.072 as well as 240-D, a gp41- specific antibody previously mapped using peptide arrays. We first generated a synthesized oligonucleotide library encoding for 39 amino acid long peptides that tile along the entire length of several Env and full-length HIV sequences from different clades. This library was cloned into a T7 bacteriophage display vector. To perform a PhIP-seq experiment, mAbs were coated on beads and incubated with the phage library, samples were sequenced in parallel, and then peptides specifically enriched by the mAb were computationally identified. Competition ELISAs were performed to compare epitope mapping results using a more traditional approach. Results: We mapped the linear epitope of QA255.067 and QA255.072 to Env amino acids 592-606 and 596-609 (HXB2 numbering), a region corresponding to the immunodominant C-C loop region of gp41. Competition ELISAs confirmed these results. We also more finely mapped the epitope of 240-D to amino acids 596-605, which is consistent with findings from structural studies. We were unable to see specific enrichment of any peptides in PhIP-seq with QA255.006 and QA25.016, but competition ELISA results indicated these mAbs target a discontinuous epitope on gp41. Conclusion: PhIP-seq mapped overlapping but distinct epitopes of two newly identified gp41 mAbs and 240-D. This method may be useful for mapping HIV antibodies that target linear epitopes, and particularly, antibodies that recognize the gp41 protein, which is an attractive vaccine target because it is relatively conserved. 311 RAPID DEVELOPMENT OF AN INFANT-DERIVED HIV-1 BROADLY NEUTRALIZING ANTIBODY Laura Doepker 1 , Cassandra Simonich 2 , Duncan Ralph 1 , James A. Williams 2 , Amrit Dhar 2 , Christopher T. Small 1 , Brian Oliver 3 , Vladimir Vigdorovich 3 , Vidya Mangala Prasad 2 , Ruth Nduati 4 , D. N. Sather 3 , Kelly K. Lee 2 , Frederick Matsen 1 , Julie M. Overbaugh 1 1 Fred Hutchinson Cancer Research Center, Seattle, WA, USA, 2 University of Washington, Seattle, WA, USA, 3 Center for Infectious Disease Research, Seattle, WA, USA, 4 University of Nairobi, Nairobi, Kenya Background: HIV-infected infants develop broadly neutralizing plasma responses with more rapid kinetics and lower somatic hypermutation than adults, suggesting the ontogeny of infant responses could inform a better path to achievable vaccine targets. We previously isolated BF520.1, the first and only infant-derived broadly HIV-neutralizing antibody (bnAb). A thorough investigation of how BF520.1 developed will highlight possible pathways of rapid bnAb development that may be useful in vaccine design. Furthermore, resolving the structural basis of BF520.1’s interaction with HIV envelope will inform the design of effective vaccine immunogens. Methods: We sequenced antibody genes from a blood sample collected midway between HIV infection and the isolation of BF520.1. We developed robust computational methods to reconstruct the developmental lineage of BF520.1 that include using personalized germline gene sets to infer the antibody sequences of BF520.1’s naïve ancestor, identifying midpoint sequences that were clonally-related to BF520.1, and phylogenetically determining likely mutational pathways that generated the mature bnAb. We compared our Bayesian lineage reconstruction approach to lineage inference by maximum likelihood, a common approach in the field. Lastly, we used single particle cryo- electron microscopy to explore the structural interaction of BF520.1 with the HIV envelope trimer BG505.SOSIP.664.

(PI3K) inhibitor, PTEN, for identifying antigen-specific MBC from HIV-infected individuals compared to healthy controls. Gene co-expression analysis showed that in addition to overexpression of PTEN, there was increased co-expression of type I interferon-associated genes with PTEN on single cell level in HIV compared to controls. Conclusion: Overall, this signature reinforces the concept of an imbalance in the interferon pathway leading toward an impairment of the ability of B cells to mature where PTEN seems to play a central role. Further, this study provides a framework for analysis of antigen-specific cells using single cell gene expression analysis and provides insight into persistent defects in B cell-mediated immunity in the context of treated HIV infection and introduce potential targets of intervention to improve vaccine responses. 309 AUTOLOGOUS NEUTRALIZING ANTIBODIES DRIVE VIRUS EVOLUTION DURING REBOUND AFTER ATI D. B. Salantes 1 , Felicity Mampe 1 , Anya Bauer 1 , Emily Lindemuth 1 , Emmanouil Papasavvas 2 , Luis Montaner 2 , Katharine J. Bar 1 1 University of Pennsylvania, Philadelphia, PA, USA, 2 Wistar Institute, Philadelphia, PA, USA Background: Characterization of the viral dynamics and host immune pressures present during HIV-1 rebound after analytical treatment interruption (ATI) provides insight into the environment in which therapeutic and curative strategies must act. Methods: We studied plasma samples from a previously conducted clinical trial (NCT00051818) in which chronically-ART initiated participants underwent a single or multiple sequential ATIs. Single genome sequencing of env genes was performed on plasma vRNA from first detectable rebound through up to 1 year of ATI. Select envs were cloned and tested as pseudoviruses for sensitivity to autologous plasma neutralizing antibody (nAb) responses in the TZM.bl assay. Results: Phylogenetic analysis of env sequences from first detectable plasma rebound in 11 participants undergoing a single ATI revealed multiple genetically distinct lineages replicating in each participant (median=5, range 2 to >10). Over time, total env diversity and the number of genetically distinct lineages expanded, with evidence of virus evolution, recombination, and reactivation of new populations. In 6 of 7 participants with adequate sampling, however, all or many of the initial rebounding lineages were not sampled in subsequent timepoints. Virus lineages that were cleared were significantly more sensitive to autologous plasma nAbs than those that persisted (median reciprocal IC50 titers of 906 vs. 153, p=0.0286, by Wilcoxon). IgG from plasma at the time of ART interruption had modest activity against cleared viruses; IgG fromweek 4 and 8 of ATI increased its potency against rebound viruses by 100 to 10,000-fold (p=0.0079, by Wilcoxon). In 2 participants from the multiple ATI arm of the study, initial rebound was comprised of multiple distinct lineages (6-7 lineages). Over 3 subsequent ATIs, a substantial number of initial lineages were no longer sampled, with autologous nAbs showing a non-significant trend towards greater potency against cleared viruses. Conclusion: In ART suppressed individuals undergoing ATI, we found that multiple virus populations arise from latency and diversify rapidly over ATI, with selective sweeps of initial rebound virus populations observed in most participants. Autologous nAbs are modest initially, but quickly expand to drive virus selection over subsequent weeks. Results suggest that autologous nAbs are an important component of the immune dynamics of rebound and should be considered in immunotherapeutic approaches to virus suppression and cure.

Poster Abstracts

CROI 2019 113