SUMMARY/ABSTRACT: The use of autologous gene modified cells that are resistant to HIV infection to reduce viral reservoir size and delay or prevent HIV recrudescence is on the forefront of HIV curative science. However, many current gene modification approaches focus solely on reducing CCR5 expression on hematopoietic stem cells or mature CD4 T cells. These approaches may have limited impact in people with HIV (PWH) that have virus able to use other coreceptors for entry, and only provide a single layer of protection against infection in vivo. As a result, the AMC #097 study, “A Phase I Study of Stem Cell Gene Therapy for HIV Mediated by Lentivector Transduced, Pre-Selected CD34+ Cells: A Trial of the AIDS Malignancy Consortium,” was implemented to combine multiple anti-HIV genes into a single lentiviral vector that is designed to block HIV-1 infection at different stages of the HIV-1 life cycle providing strong pre-integration inhibition of HIV-1 infection using a lentiviral vector that including a CCR5 shRNA, a chimeric human-macaqueTRIM5α restriction factor, and a HIV TAR decoy. This strategy has been shown to prevent infection in vitro and in vivo, and 12 PWH requiring autologous SCT for lymphoma have already received gene modified stem cells through AMC097. One participant stopped ART outside study protocol following SCT and experienced post-treatment control. Whereas the primary endpoints of the study are to evaluate the safety of such an approach, support is urgently needed to perform in- depth HIV reservoir analyses and to implement assays to determine if gene modified cells become infected following transplantation with or without analytical treatment interruption. We will: (1) test the hypothesis that autologous SCT with gene modified stem cells simultaneously targeting different stages of the HIV-1 replication cycle will lead to blood and gut tissue expansion and maintenance of a transduced CD4+ T and other immune cells resistant to HIV-1 infection; (2) test the hypothesis that SCT with gene modified stem cells will reduce the size of the HIV-1 reservoir and residual HIV-1 transcriptional activity, and lead to post-treatment HIV control following withdrawal of ART, and (3) determine if gene modified cells become infected prior to and following cessation of ART a novel duplexed single-cell-droplet (scdPCR) assay with the ability to simultaneously detect cellular HIV-1 DNA or RNA and the integrated lentiviral vector or chimeric TRIM5α transcriptional activity.