Anti-retroviral therapy (ART) is effective in reducing plasma HIV load to undetectable levels and restoring partial immunity. However, viral populations rapidly rebound once therapy is removed. Furthermore, HIV-associated co-morbidities, particularly certain cancers, remain a significant co-morbidity even with effective ART. We hypothesized that HIV-infected macrophages in tumors may both protect HIV from ART and contribute to metastasis.
The AIDS and Cancer Specimen Resource (ACSR) provided 26 post mortem tissues from three HIV+/ART+ patients with no detectable viral load. Two died from metastatic lymphoma (Pt02, Pt04), and one from lung cancer (Pt05). Tissues were assessed for HIV using digital drop PCR. Single genome amplification was used to generate env and nef sequences. We inferred maximum-likelihood phylogenies and performed multiple statistical tests to investigate viral evolutionary patterns and compartmentalization. We used an HIV signal detection technique (RNAscope) combined with histological staining to visualize the cellular location of HIV RNA in tissues from Pt02.
ddPCR was positive for all tissues. HIV env-nef DNA sequences were generated from 8/11, 3/6 and 9/9 tissues from each of the three patients, respectively. HIV RNA was also isolated from three tissues in Pt02 (lymph node tumor, cerebellum and aorta) and two tissues from Pt05 (lymph node and spleen). Maximum-likelihood phylogenies and statistical testing showed little evidence of viral compartmentalization among tissues for RNA and DNA. Considering that the patients had been on ART, some clades showed surprisingly high diversity and evidence of on-going evolution, while others were consistent with a pattern of clonal expansion. RNAscope for Pt02 showed that RNA expression of HIV gag-pol was clustered and co-localized with CD163+/CD68+ macrophages in tumor lymph node and the cerebellum, with lymphoma involvement due to meningeal metastasis.. Infiltrating macrophages surrounded HIV infected cells (Figure).
Our results suggest that lymphatic and cancer tissues may offer a privileged environment for persistent HIV replication within macrophages during ART and may promote tumor growth and metastasis. Two distinct patterns of tissue virus evolution suggest that different modes of replication/spread underlie persistence: ART-resistant HIV infected macrophages with persistent evolution/spread and clonal expansion of HIV infected cell populations.