Genetic divergence between reservoir and pre-therapy plasma HIV-1 Env variants does not confer consistent compartment-specific differences in broadly neutralizing antibody sensitivity

Broadly neutralizing antibodies (bNAbs) are a promising strategy to target the HIV-1 reservoir, the main barrier to cure. HIV-1 variants may differ in Env sequence and neutralization sensitivity across anatomical compartments. We investigated genetic differences and differences in neutralization sensitivity between Env variants persisting in peripheral blood mononuclear cell (PBMC) and lymph node (LN) reservoirs, and pre-therapy plasma variants. The study included 11 individuals on antiretroviral therapy in Durban, South Africa. Env sequences derived from pre-therapy plasma and from PBMCs and LN reservoirs were assessed for reservoir-specific genetic differences. Recombinant viruses encoding Env variants were generated for six participants and tested for neutralization sensitivity using a panel of nine clinically relevant bNAbs. We observed genetic divergence between compartments, with a median of 6 and 19 amino acid differences between pre-therapy plasma and LN and PBMC reservoirs, respectively. Of these, 63% (LN) and 73% (PBMC) were confirmed by next-generation sequencing to be unique to the reservoir, with few occurring at sites previously associated with altered bNAb sensitivity. Phenotypic analysis revealed no consistent compartment-specific patterns in neutralization sensitivity. CD4-binding site (CD4bs) and gp120-gp41 interface bNAbs showed broad coverage across compartments, while sensitivity to V2 and V3 antibodies was more variable. Although reservoir-derived Env variants show genetic divergence from pre-therapy viruses, this variation is largely not at sites reported to alter bNAb sensitivity and does not translate into consistent compartment-specific differences in bNAb sensitivity. The broad activity of CD4bs and interface-targeting bNAbs across compartments supports their suitability in antibody-based HIV cure strategies. IMPORTANCE: HIV persists in reservoirs during antiretroviral therapy, preventing a cure. Broadly neutralizing antibodies (bNAbs) are a promising strategy to target these reservoirs, but it is unclear whether viruses in different tissues respond differently to these antibodies. Here, we show that although HIV variants in blood and lymph node reservoirs differ genetically from viruses present before treatment; they do not show consistent changes in antibody sensitivity. Importantly, antibodies targeting the CD4-binding site and the gp120-gp41 interface remained broadly effective across all compartments. These findings suggest that key bNAbs can target HIV reservoirs throughout different body compartments, supporting their use in cure strategies and highlighting the value of combining antibodies to overcome viral diversity.