Background
DUALITY was a randomized trial that showed comparable reductions in HIV-1 DNA and RNA in peripheral CD4⁺ T cells from ART-naïve people with HIV (PWH) initiating DTG+3TC (2DR) or DTG+FTC/TAF (3DR) as first-line ART. Here, we applied multimodal imaging to characterize spatial dynamics of HIV-1 and antiretroviral (ARV) distribution in lymph nodes (LN) during the first year of ART.
Methods
39 individuals underwent ultrasound-guided inguinal LN excision and paired blood sampling at baseline or weeks 2, 4, 12, or 48 after ART initiation (n=4/arm/timepoint). Serial LN cryosections were analyzed using DNA/RNAscope in situ hybridization (ISH) for HIV-1 DNA and RNA (vDNA, vRNA), immunohistochemistry (IHC) for CD20⁺ B cells and collagen deposition, and mass spectrometry imaging (MSI) for ARV distribution. Spatial overlap of HIV markers and ARVs was assessed by nearest-neighbor analysis of co-registered images (MATLAB). Due to sampling constraints, changes in vDNA and vRNA in LN were analyzed descriptively as ratios of medians week48/week0, with bootstrap-derived 95% CI.
Results
vDNA and vRNA levels in LN infected-cells declined rapidly upon ART initiation. At week 48, vDNA decreased by 84.2% (58.4–96.4) in 2DR and 94.5% (30.8–99.3) in 3DR. Similarly, vRNA levels declined by 95.1% (46.8–99.8) in 2DR and 99.9% (99.1–99.9) in 3DR. Cell-associated vDNA in LN correlated with total and intact HIV-1 DNA levels in peripheral CD4⁺ T cells (r = 0.55, p = 0.0005; r = 0.67, p = 0.0039, respectively). Among 3DR MSI ARV targets, tenofovir (TFV) was not detectable, although LC-MS/MS confirmed TFV-dp and FTC-tp concentrations exceeding EC50 (36.7 and 29.6 fmol/106 cells, respectively); so spatial analysis focused on 2DR. Spatial measurement of DTG+3TC indicated exposure of ≥1 ARV across median 66.1% (range, 19.7 – 81.8%) of LN tissue – mainly DTG, with 33.8% (4.0 – 64.0%) of tissue being covered by both 3TC and DTG. Notably, 100% (50 – 100%) of vDNA+ cells and 100% (67 – 100%) of vRNA+ cells neighbored ≥1 ARV, with 50% (0 – 100%) of vDNA+ cells and 55% (10 – 100%) of vRNA+ cells being directly colocalized with ≥1 ARV. High collagen, a marker of LN fibrosis, was associated with reduced ARV coverage and lower DTG concentrations.
Conclusions
Consistent with findings in peripheral CD4⁺ T cells, DTG+3TC induced a rapid vDNA and vRNA decline in LN, comparable to DTG+FTC/TAF. Spatial mapping showed extensive LN penetration of DTG+3TC, supporting effective drug delivery to tissue reservoirs.