# Quantifying bNAb neutralization of the HIV latent reservoir

> **NIH NIH R61** · ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI · 2024 · $425,302

## Abstract

Project Summary
 Broadly neutralizing antibodies (bNAb) against HIV Env represent a novel class of antivirals that have the
potential to neutralize viral entry, while simultaneously activating non-neutralizing immune effector functions.
However, a major obstacle for their clinical use is identifying the virologically suppressed HIV+ patients that are
best suited for a given bNAb or bNAb combination. Indeed, early-stage therapeutic studies with bNAb have not
found uniform concordance between viral outgrowth assay sequences (in vitro) and the Env resistance
mutations that arise in treated patients in vivo. Our laboratory has developed robust and quantitative flow
cytometry-based assays for neutralization of both cell-free and cell-to-cell infection. We have observed
decreases in the potency and efficacy of Abs using cell-associated versus cell-free HIV-1 neutralization
assays. The detection of these cell-associated phenotypes may be particularly important for cure-based
approaches where functional reactivity of bNAbs against infected cells may be important for in vivo efficacy. To
overcome the limitations of PCR-first based approaches that often identify non-functional Env, we propose the
development of a single cell Viral Outgrowth Neutralization Assay (scVONA) that will be performed in the
presence and absence of bNAb. This scVONA will simultaneously quantify viral latent reservoir sequences and
determine their sensitivity to bNAb. To accomplish this, scVONAs will leverage and enhance a highly sensitive
single cell reporter cell line to detect infection and identify cells to test for neutralization of cell-to-cell spread.
The ultimate readout are single cell flow cytometry measures that enable capture of resistant env genes by
sorting and sequencing insensitive versus sensitive clones. Long read Env sequencing will be employed for
bioinformatic analyses to determine linked sequences that correlate with resistance. Titration of bNAb will also
provide estimated IC50 and sequences may allow a measure of the fraction of sensitive clones. We believe
that scVONA will provide a rapid <2 week readout of neutralization sensitivity and simultaneously provide a
measure of functionally intact env genes for confirmatory analyses, significantly reducing the time, cost and
labor associated with identifying bNAb resistant sequences from HIV+ patient samples.

## Key facts

- **NIH application ID:** 10816554
- **Project number:** 5R61AI176585-02
- **Recipient organization:** ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
- **Principal Investigator:** BENJAMIN K CHEN
- **Activity code:** R61 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $425,302
- **Award type:** 5
- **Project period:** 2023-04-01 → 2026-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10816554

## Citation

> US National Institutes of Health, RePORTER application 10816554, Quantifying bNAb neutralization of the HIV latent reservoir (5R61AI176585-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10816554. Licensed CC0.

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