CROI 2018 Abstract eBook

Abstract eBook

Oral Abstracts

needed on i) expanding use of innovative strategies for HIV prevention and treatment access for the young, males and vulnerable populations, ii) expansion of flexible and adaptive strategies to respond rapidly to diverse epidemics, and iii) sustained funding to enable expanded coverage of HIV treatment and prevention services. 111 IDENTIFICATION OF HIV-1 ENV MUTATIONS THAT CONFER BROAD RESISTANCE TO ARVs IN VITRO Rachel Van Duyne , Phuong Pham, Eric O. Freed National Cancer Institute, Frederick, MD, USA Background: We have identified compensatory mutations within the HIV-1 envelope (Env) glycoprotein that rescue defects in replication conferred by mutations in Gag. Despite severe deficiencies in cell-free infectivity, these Env mutants replicate with wild-type (WT) kinetics and are capable of efficient cell-to-cell transmission. The goal of this study was to characterize the ability of the Env mutant viruses to confer a broad replication advantage and determine whether they can overcome or escape inhibition by antiretrovirals (ARVs). Methods: We passaged Env mutants in T-cell lines to establish a spreading infection and measured replication kinetics in the presence or absence of ARVs. We also selected for viral isolates exhibiting at least partial resistance to ARVs as indicated by efficient replication in the presence of the inhibitors. Results: We demonstrate that the Env compensatory mutants replicate with WT or faster-than-WT kinetics despite severe defects in cell-free, single-cycle infectivity. We also observed that the Env mutants can rescue a replication- defective integrase mutant, suggesting that they might also be able to confer resistance to ARVs. Indeed, we found that the Env mutants exhibit markedly reduced sensitivity to Ritonavir (RTV), Nelfinavir (NFV), Dolutegravir (DTG), Tenofovir (TFV), and Rilpivirine (TMC) at drug concentrations that block or delay WT virus replication. Remarkably, long-term passage of WT virus in the presence of several of these inhibitors resulted in the selection of partially resistant ARV-escape mutants containing substitutions in Env. This escape occurred in the absence of resistance mutations in enzyme targets of these drugs. The positions of the selected Env mutations are highly conserved (>90%) across all clade B viruses, highlighting the specificity of selection of mutations at these positions. Conclusion: These results demonstrate that mutations in Env that promote efficient cell-cell transfer, at the expense of cell-free particle infectivity, can broadly contribute to drug resistance in vitro. Cell-to-cell HIV-1 transmission occurs more efficiently and rapidly than cell-free infection, supporting the relevance of this mode of viral dissemination in vivo. These results also raise the possibility that the acquisition of Env mutations represents an unrecognized, transient stepping stone towards the development of high-level HIV-1 drug resistance. 112 STRAIN-DEPENDENT ACTIVATION AND INHIBITION OF HIV-1 ENTRY BY A PF-68742 DIASTEREOMER Connie A. Zhao 1 , Amy Princiotto 1 , Mark Farrell 2 , Amos B. Smith 2 , Navid Madani 1 , Joseph G. Sodroski 1 1 Dana–Farber Cancer Institute, Boston, MA, USA, 2 University of Pennsylvania, Philadelphia, PA, USA Background: HIV-1 entry into cells is mediated by the envelope (Env) trimer of gp120-gp41 heterodimers. Sequential binding to target cell receptors, CD4 and CCR5 or CXCR4, triggers the metastable Env to undergo entry-related conformational changes. PF-68742 was recently identified as a small molecule that inhibits infection of a subset of HIV-1 strains by interfering with an Env function other than receptor binding, with resistance determinants mapping to the gp41 disulfide loop and fusion peptide. We investigated the antiviral mechanism of PF-68742. Methods: Recombinant luciferase-expressing HIV-1 pseudotyped by wild-type or mutant HIV-1 Envs was incubated with increasing concentrations of PF-68742 alone or in the presence of other entry inhibitors or antibodies. The virus- inhibitor mixture was added to CD4+ CCR5+, CD4+ CXCR4+, or CD4- CCR5+ target cells, and luciferase activity was measured 48 to 72 hr later. Results: Of the four PF-68742 diastereomers, only one, MF275, inhibited the infection of CD4+ CCR5+ cells by some HIV-1 strains. Unexpectedly, MF275 activated the infection of CD4- CCR5+ cells by several HIV-1 strains resistant to the compound’s inhibitory effects in CD4+ CCR5+ target cells. In both cases, the strain susceptibility profiles were unique from those of other entry inhibitors. Sensitivity to other entry inhibitors indicated that MF275-activated virus entry requires CCR5 binding as well as gp41 heptad repeat (HR1) formation and

exposure. In contrast to CD4 complementation by CD4-mimetic compounds, activation of CD4-independent infection by MF275 did not depend upon availability of the gp120 Phe 43 cavity; moreover, the MF275-activated state was long-lived relative to that induced by CD4-mimetic compounds. While MF275 and a CD4-mimetic both enhanced susceptibility of some HIV-1 strains to the 17b and 19b antibodies against a CD4-induced epitope and the gp120 V3 loop, respectively, only MF275 enhanced susceptibility to the broadly neutralizing antibody 4e10 against the gp41 membrane-proximal external region. Conclusion: MF275 apparently binds a site on the HIV-1 Env unique from the CD4 binding site to activate a conformational cascade that leads to virus entry. This pathway is parallel to but distinct from that triggered by CD4 and CD4- mimetic compounds. Thus, HIV-1 Env samples conformations within an energy landscape in which there are multiple pathways to entry (or inactivation). An understanding of the mechanisms of activity of MF275 should assist efforts to optimize its utility. 113LB TRISPECIFIC ANTIBODIES FOR PREVENTION AND TREATMENT OF HIV-1 INFECTION Amarendra Pegu 1 , Zhi-yong Yang 2 , Ling Xu 2 , Ercole Rao 2 , Nicole Doria-Rose 1 , Jochen Beninga 2 , Krisha McKee 1 , Dana M. Lord 2 , Ronnie R. Wei 2 , Gejing Deng 2 , Stephen Schmidt 1 , Wang Keyun 1 , Mario Roederer 1 , John R. Mascola 1 , Gary Nabel 2 1 Vaccine Research Center, NIAID, Bethesda, MD, USA, 2 Sanofi, Cambridge, MA, USA Background: Broadly neutralizing anti-HIV-1 antibodies (bnAbs) are promising candidates for use in both prevention and treatment of HIV-1. These bnAbs have been shown to be efficacious in animal models and ongoing clinical studies suggest they mediate anti-viral activity in HIV-1 infected humans. Due to the diversity of circulating HIV-1 strains, the use of a single bnAb will likely lead to development or selection of resistant strains. Therefore, a combination of two or more bnAbs will be required for effective anti-HIV-1 therapy or to cover diverse strains for immunoprophylaxis. Methods: Here, we engineered trispecific antibodies (Abs) that allow a single molecule to interact with three independent HIV-1 envelope determinants: 1) the CD4 binding site, 2) the membrane proximal external region (MPER) and 3) the V1V2 glycan site. These trispecific Abs were then assessed in neutralization assays against a representative global panel of HIV-1 viruses. The most potent and broad trispecific Abs were then evaluated for pharmacokinetics and protective efficacy in non-human primates (NHPs). Results: Trispecific Abs exhibited higher potency and breadth than any previously described single bnAb. They showed pharmacokinetics like human bnAbs in naive rhesus macaques. Compared to single bnAbs that allowed breakthrough infection frommixed SHIV challenge of rhesus macaques, the trispecific Ab conferred complete immunity against a mixture of SHIVs. Conclusion: Trispecific Abs thus constitute a platform to engage multiple therapeutic targets through a single protein, and could be applicable for diverse diseases, including infections, cancer and autoimmunity. Background: The human genome encodes seven APOBEC3 (A3) enzymes, at least four of which (A3D, A3F, A3G, and A3H) can induce G-to-A mutations in HIV-1 genomes. These enzymes leave two distinct hypermutation signatures: GG-to-AG and GA-to-AA. The former signature is dominant in viral sequences hypermutated in vitro by A3G, whereas the latter is prevalent in sequences hypermutated in vitro by A3D, A3F, or A3H. Current, in vitro based models posit that all HIV-restrictive A3 enzymes are ubiquitously active and cooperate to hypermutate HIV. However we have discovered that an in vivo hypermutated virus typically bears a dominant GG-to-AG signature or a dominant GA-to-AA signature, which would be expected from independent encounters with A3G or A3D/F/H, but not from both enzyme classes simultaneously. This hypermutation bias towards GG-to-AG or GA-to-AA suggests the existence of a mechanism that prevents A3 proteins from simultaneously targeting HIV-1. Methods: We performed four independent analyses: 1) We analyzed all reported in vivo hypermutated HIV-1 sequences (1164 sequences from 988 patients) using two independent methods (non-alignment-based and alignment-based); 2) We analyzed all of the 564 SNPs of the A3 locus in 2504 individuals from 26 populations (1000 Genomes Project). 3) We quantified, using RNAseq data, all of the reported A3 transcripts in 461 donors from the 114 MOLECULAR DETERMINANTS OF HIV HYPERMUTATION Diako Ebrahimi , Christopher Richards, Michael A. Carpenter, Jennifer McCann, Adam Cheng, Terumasa Ikeda, Daniel Salamango, Reuben S. Harris University of Minnesota, Minneapolis, MN, USA

Oral Abstracts

39

CROI 2018