ORCID

https://orcid.org/0000-0003-3025-125X

Date of Award

10-31-2023

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Molecular Genetics & Genomics)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

HIV infection remains a major global health burden with 1.7 million new acquisitions and 770,000 deaths in 2018 alone. While these numbers are declining due to antiretroviral therapy, only a handful of individuals have achieved complete remission from HIV. With the advent of antiretroviral therapy, HIV has turned a death sentence into a chronic, manageable condition. However, current antiretroviral therapies have only been studied on their ability to block infection and have not been able to clear the viral reservoir. Additionally, HIV cure strategies often rely upon sensing of highly variable epitopes such as the HIV envelope and have not achieved broad success. We previously identified a novel pattern recognition receptor, the caspase recruitment domain family member 8 (CARD8), that can sense HIV-1 protease activity in infected cells after dimerization by non-nucleoside reverse transcriptase inhibitors. CARD8 can be cleaved by HIV-1 protease and induces Caspase-1 inflammasome activation and pyroptosis of infected cells. The utilization of NNRTIs to induce CARD8 inflammasome activation offers a promising strategy for eliminating HIV-infected cells in a “Shock and Kill” approach due to its reliance upon viral protease activity which is less susceptible to mutation than its envelope counterpart. However, this process has reduced efficacy in vivo due to high binding affinity of NNRTI’s to serum proteins and antiretroviral drug resistance. Therefore, it is imperative to elucidate ways to sensitize the CARD8 inflammasome to NNRTI-induced activation. We show that this sensitization can be achieved through chemical inhibition of the CARD8 negative regulator DPP9. The DPP9 inhibitor Val-boroPro (VbP) kills HIV-1-infected cells without the presence of NNRTIs and acts synergistically with NNRTIs to promote clearance of HIV-1-infected cells in vitro and in humanized mice. More importantly, VbP can enhance clearance of residual HIV in CD4+ T cells isolated from people living with HIV (PLWH). We also show that VbP can ameliorate issues with NNRTI serum binding by reducing the threshold for CARD8 activation and can partially overcome NNRTI resistance. This offers a solution to enhance NNRTI efficacy in the elimination of HIV-1 reservoirs in PLWH. We also recently discovered that CARD8 inflammasome activation, immediately upon HIV-1 viral entry and release of HIV protease, may contribute substantially to CD4+ T cell loss. The loss of non-productively infected CD4+ T cells, known as bystander cell death, is the main contributor to HIV-1 disease progression and AIDS. Until the elucidation of CARD8’s role in this process, it was largely unknown how these cells undergo cell death without productive infection. Additionally, disease progression in non-human primate models of SIV infection display stark differences in their ability to recapitulate human disease. Some NHP species, such as those in the Cercocebus and Chlorocebus genera, do not develop CD4+ T cell loss and AIDS like symptoms in contrast to members of the Macaca genus. These non-pathogenic or “natural” hosts of SIV infection raise questions of CARD8’s role in lentiviral disease progression. We discovered loss-of-function mutations in the CARD8-coding genes from these “natural hosts”, which may explain the peculiarly non-pathogenic nature of these infections. These mutations were likely a byproduct of a novel reverse tandem gene duplication in the CARD8 gene of the Cercopithecoidae lineage which allows for increased mutational rates. As only the non-pathogenic hosts of SIV infection have naturally experienced SIV infection, it is possible that these loss-of-function mutations were selected to avoid CD4+ T Cell loss in endemic populations. We show that only the pathogenic hosts of infection have functional CARD8 proteins that can be cleaved and activated by SIV protease and that can induce rapid cell death. We propose this mechanism as a potential explanation for the difference in pathogenicity of non-human primate species.

Language

English (en)

Chair and Committee

Liang Shan

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