Mechanisms of Human Immunodeficiency Virus (HIV) Persistence and Strategies to Elicit Cytotoxic T Lymphocyte-Mediated Clearance of HIV-Infected Cells
Creators: name: Painter, Mark
Publisher: name: My University
Year: 2020
Subjects: subject: Human immunodeficiency virus (HIV)
subject: Latency
subject: HIV cure
subject: Cytotoxic T lymphocytes
subject: Hematopoietic stem and progenitor cells
subject: Nef
subject: Microbiology and Immunology
subject: Science

description: Human immunodeficiency virus (HIV), the cause of a decades-long pandemic responsible for over 30 million deaths and 38 million ongoing infections, establishes a chronic infection for which there is no cure. While the development of combination antiretroviral therapy (ART) has radically transformed the course of the pandemic, individuals living with HIV must maintain therapy for the remainder of their lives. This is the result of cells harboring latent HIV proviruses, which are stably integrated in the host cell DNA but remain transcriptionally silent. These latent proviruses are not targeted by ART and evade clearance by the host immune response, but can begin to express viral genes and re-establish an ongoing infection in the event of ART interruption. The leading theoretical framework to achieve an HIV cure is the “shock and kill” approach, in which latent proviruses are therapeutically reactivated to express viral genes and subsequently killed by the cytopathic effects of the virus or the host immune response. The latent reservoir of replication-competent HIV is found in a multitude of quiescent cell types residing in diverse tissues, including resting memory CD4+ T cells and hematopoietic stem and progenitor cells (HSPCs). Understanding the mechanisms regulating HIV latency and reactivation in quiescent cells will enable the development of targeted latency reversing agents (LRAs). Chapter 2 describes two modifications to in vitro cultures that independently maintain primary HSPCs in a quiescent state. These quiescent HSPCs are susceptible to HIV infection, but preferentially harbor latent proviruses that have a significantly reduced likelihood of spontaneous reactivation. Latent proviruses in quiescent cells are resistant to therapeutic reactivation by histone deacetylase inhibition or P-TEFb activation, but are responsive to NFkB activation. Collectively, this work provides a path forward to identify mechanisms contributing to latency and latency reversal in quiescent primary cells. In the event that a potent shock or sequential shocks successfully induce HIV gene expression in every cell harboring a replication-competent provirus, these cells need to be killed before the latent reservoir can be reseeded. Cytotoxic T lymphocytes (CTLs) are the main effectors of the adaptive immune system responsible for eliminating HIV-infected cells by recognizing HIV peptides presented by MHC-I on the cell surface. HIV evades these responses through the activity of the accessory protein Nef, which downregulates MHC-I by redirecting it to the lysosome instead of the plasma membrane. The work described in Chapter 3 led to the identification of concanamycin A (CMA) as a potent inhibitor of HIV Nef. CMA counteracted Nef at sub-nanomolar concentrations that did not interfere with lysosomal acidification or degradation and were non-toxic in primary cell cultures. CMA specifically reversed Nef-mediated downregulation of MHC-I, but not CD4, and cells treated with CMA showed reduced formation of the AP-1:Nef:MHC-I complex required for MHC-I downregulation. CMA restored expression of diverse allotypes of MHC-I in Nef-expressing cells and inhibited Nef alleles from divergent clades of HIV and SIV, including from primary patient isolates. Importantly, restoration of MHC-I in HIV-infected cells was accompanied by enhanced CTL-mediated clearance of infected cells comparable to genetic deletion of Nef. Thus, CMA is a promising lead compound for therapeutic inhibition of Nef to enhance immune-mediated clearance of HIV-infected cells.