CROI 2025 Abstract eBook

Abstract eBook

Poster Abstracts

507

Temsavir Treatment Improves Recognition of HIV-1 Infected Cells by Broadly Neutralizing Antibodies Robert Ferris, James Schawalder, Cristin Galardi, Richard Dunham, Heather Madsen, Hangfei Qi ViiV Healthcare, Durham, NC, USA Background: HIV-1 envelope glycoprotein (Env) is expressed on the surface of infected cells, making it an attractive target for the clearance of these cells through antibody-mediated immune activities, such as antibody-dependent cellular cytotoxicity (ADCC). It was previously demonstrated that treatment with temsavir (TMR), a small molecule, can enhance the binding of the broadly neutralizing antibody (bnAb) N6, leading to improved clearance of HIV-1 infected cells. TMR is the active form of fostemsavir (FTR), a drug approved for use in heavily treatment-experienced adults with multidrug-resistant HIV-1 infection. TMR binds to and stabilizes Env in a 'closed' conformation, which facilitates better recognition of Env by N6 and potentially other bnAbs. Methods: To investigate whether TMR can modulate Env binding on infected cells by bnAbs other than N6, we conducted flow cytometry studies using HIV-1 infected primary CD4+ T cells. CD4+ T cells that were isolated from healthy PBMCs were infected with HIV-1 and treated with TMR for 24h prior to measurement of antibody binding in flow cytometry. Results: We found that TMR treatment enhances binding of most bnAbs, including antibodies targeting CD4bs, V1V2, V3 glycans and gp120/41 interface with the exception of 10E8, to HIV-1 infected cells, leading to increased clearance of infected cells through ADCC. The enhanced binding of bnAb by TMR treatment is most prominently observed on infected cells that maintain CD4 expression, the cells that are otherwise difficult to target by bnAbs alone. When tested with a panel of clinical isolate viruses, TMR was found to enhance bnAb binding at concentrations similar to its antiviral potency. This suggests that the mechanism of action (MOA) is on-target and that this synergistic effect could potentially be achieved in clinical settings with approved doses of FTR. In kinetics experiments, we found that effective enhancement of antibody binding was observed with a treatment duration of 6h and longer, but a treatment duration of 30min was insufficient to produce this benefit. Conclusions: Our results demonstrate that TMR treatment enhances antibody binding to the infected cells that maintain CD4 expression with a majority of bnAbs, at concentrations similar to the antiviral potency of TMR. These suggest that combining bnAbs with TMR can broaden the range of HIV-1 infected cells susceptible to antibody-mediated clearance, potentially increasing the likelihood of reservoir reduction in clinical settings. Maximizing Benefits to Participants in Analytic Treatment Interruption With Antibody Infusions Morgane Rolland 1 , Hongjun Bai 1 , Mélanie Merbah 1 , Gabriel Smith 2 , Elizabeth Comeau 1 , Matthew Lind 1 , Carlo Sacdalan 3 , Nittaya Phanuphak 4 , Kiat Ruxrungtham 5 , Sandhya Vasan 2 , Merlin Robb 1 , Donn Colby 1 , Lydie Trautmann 2 , Yifan Li 1 , for the RV254/RV630 Study Group 1 US Military HIV Research Program, Bethesda, MD, USA, 2 Henry M Jackson Foundation, Bethesda, MD, USA, 3 SEARCH, Bangkok, Thailand, 4 Institute of HIV Research and Innovation, Bangkok, Thailand, 5 Thai Red Cross AIDS Research Centre, Bangkok, Thailand Background: Analytic treatment interruption (ATI) studies utilizing broadly neutralizing antibody (bnAb) infusions are associated with delayed viral rebound if viruses found in the participants are sensitive to the bnAbs. Yet, viral sensitivity is typically evaluated only after the ATI has been conducted. For a future ATI that will enroll 35 participants from the RV254 Thai cohort, we screened HIV-1 sequences for sensitivity to the bnAbs that will be used in the ATI: PGDM1400LS and VRC07-523LS. Methods: Plasma samples were sequenced via single genome amplification or the Pacific Biosciences single molecule real-time platform using unique molecular identifiers. Envelope (env) sequences were cloned and pseudoviruses with these Env were tested for neutralization sensitivity to bnAbs using IC80<1ug/mL as a cutoff. Results: Among 649 participants enrolled during acute HIV-1 infection who initiated antiretroviral treatment (ART) within days of diagnosis, 168 (98% male) met study eligibility criteria and 39,907 env sequences were obtained from 160 participants. Most participants had CRF01_AE env (85%). Sequence analysis showed minimal diversity (average mean pairwise diversity=0.004) with a single HIV-1 lineage in most participants (61%). Half of the infections with multiple founder lineages showed rare lineages that represented <3% of the viral population. For each participant, in silico predictions of bnAb

sensitivity were used to select representative env sequences for neutralization assays. Neutralization data from 116 participants showed 54 (46.5%) sensitive to PGDM1400LS, 55 (46.7%) sensitive to VRC07-523LS, and 25 (21.5%) sensitive to both. We also modeled the Instantaneous Inhibitory Potential (IIP) values; at week 20 post ATI, 75 participants had clones with IIP (PGDM1400 + VRC07)>3.5, allowing prioritization of participants for study enrolment. Conclusions: We developed a strategy combining sequence analysis, in silico bnAb sensitivity predictions and neutralization assays to prioritize enrollment of participants with the most sensitive viruses to the bnAbs that will be used in an ATI. Our approach benefitted from the fact that participants initiated ART in acute infection and therefore had a typically homogeneous viral reservoir recapitulated by the limited number of distinct env sequences we sampled. While the benefits of this approach will only be ascertained after the ATI is conducted, these data provide the largest description of sensitivity to two potent bnAbs across CRF01_AE viruses. Myeloid Targeted and Enhanced Endosomal Escapable LNPs Improve Latent HIV-1 Elimination Efficacy Soumya S. Dey, Sudipta Panja, Bharat N. Chaudhary, Mohammad Uzair Ali, Santhi Gorantla, Howard Gendelman University of Nebraska Medical Center, Omaha, NE, USA Background: A cure for HIV remains elusive as the excision of latent proviral DNA by adenovirus-based therapeutic systems has failed to date due to vector immunogenicity and limited targeting to viral reservoirs. In contrast, lipid nanoparticles (LNPs) are promising for targeting viral reservoirs, offering greater cargo-carrying capacity and reduced immune responses. However, limitations in endosomal escape remain a critical impediment. Therefore, the current system, must show sufficient cytoplasmic release of its cargo to impact clinical outcomes. Herein, disulfide-containing ionizable lipid-based LNPs were developed to leverage the high levels of reactive oxygen species (ROS) in myeloid phagolysosomes. The ROS-sensitive LNPs promote cleavage of disulfide bonds, leading to enhanced endosomal escape and improved cytosolic delivery of CRISPR guide RNAs (gRNAs). Methods: A four-step synthetic procedure was used to synthesize a library of 20 ionizable lipids with cleavable disulfide linkages. Transfection efficacy was assessed by a FLuc reporter mRNA in human monocyte-derived macrophages (MDMs). FDA-approved ionizable lipids, MC3 and ALC-0315, were compactors against the lead candidate (DS-LNP). IVIS was used to evaluate biodistribution. The toxicity profile of the new ionizable lipid was tested via CTB assay. The lead candidate delivered CRISPR-Cas9 gRNAs targeting five HIV-1 exons (tat1-2/ rev1-2/gp41), and CRISPR-Cas9-mediated excision was validated by PCR and gel electrophoresis in HIV-infected MDMs. Results: Our lead LNP formulation (DS-LNP), consisting of a novel disulfide based ionizable lipid, DOPC, Cholesterol, and DMG-PEG2K, was nontoxic and demonstrated a 20-fold higher mRNA transfection efficiency in MDMs compared to MC3 and ALC-0315, which are limited by poor endosomal escape in myeloid cells. The DS-LNP biodistribution was specifically observed in the lymphoid and pulmonary viral reservoirs of humanized mice, marking a significant advancement over conventional formulations that inadequately target these reservoirs. CCR5-targeted DS-LNP improved MDMs targeting and endosomal escape, achieving 80% excision of HIV-1 proviral DNA, a clear advancement over conventional formulations. Conclusions: Disulfide-based LNPs offer a significant advantage in overcoming endosomal escape barriers in MDMs, facilitating effective CRISPR-Cas9 delivery to HIV reservoirs. This approach potentially targets the lymphoreticular and pulmonary system and can improve viral DNA elimination. A Novel and Promising Strategy of HIV Eradication via LTR-LTR Recombination Feng Li 1 , Guanhan Li 1 , Luke Kelley 1 , Robert Gorelick 2 , Frank Maldarelli 1 1 National Institutes of Health, Bethesda, MD, USA, 2 Frederick National Laboratory for Cancer Research, Frederick, MD, USA Background: Long-lived HIV-infected cells persist and undergo clonal expansion, preventing viral eradication during antiretroviral therapy. The majority of infected cells contain defective proviruses with mutations, deletions, or insertions. Defective proviruses are also generated by LTR-LTR recombination, excising the entire coding sequence, leaving only a solo LTR The figure, table, or graphic for this abstract has been removed.

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