Prior to 1996, the prognosis of HIV disease was near-certain death; however, biomedical advancements in the past 20 years established HIV as a chronic manageable disease with a nearly normal life span. Recent advancements suggest the potential for a cure. One outcome of current medical treatments is that 50% of all HIV positive individuals are older (-?¥ 50years), and a substantial number of those individuals are long-term (-?¥ 20 years) survivors. Existing research Qualitative research has provided little insight about what older long-term HIV survivors believe about their disease circumstances and aging with the disease. A qualitative method in the phenomenological tradition was used to explore older long-term HIV survivors' notions about aging with HIV and an HIV cure. The self-regulation model of illness representations and the preventive and corrective, proactivity (PCP) model of aging with HIV disease for older adults guided the study. Using strategically placed flyers in HIV services environments, 12 older long-term HIV survivors volunteered to describe their beliefs about aging with HIV and an HIV cure. Participants' statements were entered into discrete cells in an electronic spreadsheet (Excel) and were coded, sorted, and categorized. The categories were sorted for commonality, and emergent themes and subthemes were identified. Older long-term HIV survivors believed they had few issues aging with HIV, expected to live a long time, and believed that finding a cure would have little effect on their lives. These research findings may be beneficial to healthcare providers and researchers who provide quality of life interventions and information to older adults living long-term with HIV who are concerned about aging, longevity, and a cure.

The Human Immunodeficiency virus-1 can stay undetected and unaffected by host immune surveillance and antiretroviral therapy. This phenomenon is called proviral latency and the cells harbouring such viruses are part of the latently infected cell reservoir. In this situation, the viral genome integrates into the host's genome upon infection, whereby infected cells exhibit either very low levels or no viral transcription, and hence no viral proteins or egress viruses are produced that can be detected by the immune system. However, viral transcription can be re-activated to produce infectious viruses under certain circumstances. Host-encoded retroviral restriction factors like APOBEC3 (A3) proteins are part of our intrinsic immune defences against retroviral infection, introducing mutations in viral replication intermediates. We hypothesize that low levels of G-to-A transition mutations in the HIV-1 LTR region, introduced by APOBEC3G/F, could lead to a latency-like phenotype. These latent viruses pose major hurdles for HIV-1 cure therapies. Our lab previously created a library of clones possessing mutations in the LTR introduced by A3G/F. Later, mutated LTRs were cloned into 3 types of plasmid backbones: 1) a pEGFP expression vector to study the transcriptional activity of the mutated promoter, 2) into non-replicative pNL4 ∆env ∆vif viral expression vector, and 3) into a replicative pNL4-CXCR4 viral vector to study infection and induction by latency reversal agent (LRA) treatment to better understand the mechanism of latency and transcriptional induction. Viruses produced from these plasmids carrying mutated promoters are referred to as latency-prone viruses or LPVs in this thesis. Characterizing the transcription, infection, and induction to PMA/I of the LPVs would essentially help in evaluating the role of A3 mutations in viral latency and further help in the development of new therapeutics.

Human Immunodeficiency Virus (HIV) infection initiates major metabolic and cell- survival complications. Anti-retroviral therapy (ART) is the current approach to suppress active HIV replication to a level of undetected viral load, but it is not a curative approach. Newer and sophisticated gene editing technologies could indeed be a potent antiviral therapy to achieve a clinical sterilization/cure of HIV infection. Chronic HIV patients, even under a successful ART regimen, exhibit a low-grade inflammation, immune senescence, premature aging, telomeric DNA attrition, T cell apoptosis, and cellular homeostasis. In this dissertation, we investigated CD4 T cell homeostasis, degree of T cell apoptosis, an associated telomeric DNA damage, DNA damage repair signaling, and the apoptotic pathways in CD4 T cells during HIV infection with or without ART treatment. Our data support a DNA damage accumulation, and impaired DNA damage repair in chromosome ends via recruitment of 53BP1 protein to the damaged foci. We found that a key player of DNA damage and repair enzyme, ATM, and its associated checkpoint proteins (CHK1, CKH2) are affected by HIV infection. HIV infection also altered another multifunctional master regulator protein AKT that is crucial in maintaining cellular homeostasis. Curing HIV is the ultimate redemption against HIV-associated complications. To explore the possibility of a functional cure, we investigated the use of a transient and a non-viral CRISPR/Cas9-based gene editing technology targeting the latently incorporated HIV provirus. After performing a nucleofection/electroporation using an in vitro formulated ribonucleoprotein (RNP) constituting a synthetic guide RNA (gRNA) and Cas9 nuclease protein, we demonstrated a significant (maximum 97%) reduction of HIV-mRNA and p24-capsid protein expression, upon stimulation (using PMA) and latency reactivation of latently HIV-infected CD4 T cells and latent-monocytes. Notably, the RNP treatment did not induce any cytotoxic effects, without affecting the abilility of cell proliferation. A sequence specific cleavage of HIV-provirus in two crucial gene locations (targeting vpr/tat genes) showed the most significant suppression of HIV reactivation or latency reversal. We have used DNA sequencing, and T7EI assay to confirm the target-site-specific cleavage of the HIV-proviral genome. Our data confirm the activation of non- homologous end joining (NHEJ) pathway to repair the double-stranded DNA break created by the CRISPR/Cas9 treatment. Taken together, this study provides a new gene therapeutic approach using synthetic gRNA/Cas9 targeting HIV genome, which warrant further in vivo animal and human studies.