Functional Cure for HIV: Lymphoid T cell Dynamics and PD-1 Immunotherapy 公开

Mylvaganam, Geetha Hanna (2016)

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Human immunodeficiency virus (HIV) has emerged as one of the most devastating global health burdens in history with approximately 36 million individuals infected worldwide. The introduction of anti-retroviral therapy (ART) has greatly enhanced viral control and the quality of life for individuals living with HIV, but ART remains a life long therapy due to latently infected CD4 T cells that are seeded early during primary infection. The latent HIV reservoir is directly responsible for viral resurgence post treatment interruption. Due to the limited success in generating a highly effective prophylactic vaccine for HIV and with 15 million and a growing number of HIV infected individuals on ART, researchers have re-directed their focus towards generating a more feasible approach to treating HIV, termed a "functional cure". The key to achieving a functional cure for HIV requires reduction/elimination of viral reservoirs and restoration of functional anti-viral CD8 T cells. The viral reservoirs are concentrated at lymphoid sites and thus there is a need for understanding the dynamics of virus-infected CD4 T cells and anti-viral CD8 T cells in the lymphoid tissue during chronic infection. Using a macaque model system of SIVmac251 pathogenesis, our studies revealed critical findings. Firstly, we observed that PD-1hi Tfh cells are aberrantly enriched in the lymph nodes (LN) and rectal mucosa of chronically SIV infected macaques and contribute to ongoing viral replication and production. Secondly, we identified a novel subset of germinal center infiltrating CXCR5+ SIV specific CD8 T cells that rapidly expand in vaccinated SIV controllers, can limit the expansion of virally infected Tfh, and are associated with enhanced viral control. Lastly, immunomodulation of the PD-1 pathway prior to and during the initiation phase of ART in a therapeutic SIV/ART macaque trial demonstrated significantly enhanced proliferation, cytotoxic potential, and polyfunctionality of anti-viral CD8 T cells resulting in markedly faster suppression of virus replication following the initiation of ART. Together, these findings further our understanding of some of the fundamental aspects of basic HIV/SIV biology and provide insight into novel therapeutic interventions that can be administered in combination with ART.

Table of Contents




Chapter 1: Introduction. 1

History and Global Burden of HIV/AIDS. 1

Transmission and Pathogenesis. 1

ART and Viral Reservoirs. 6

Immune Drivers of Viral Persistence. 9

Immune Response to HIV. 12

Innate immune response. 12

Adaptive immune response. 12

Humoral Immunity. 13

Cell mediated immunity. 14

T Follicular helper cells and HIV. 17

CD8 T cells and HIV. 20

Immune Exhaustion and Check-point Inhibition. 22

Summary. 27

Chapter 2: Diminished Viral Control during SIV Infection is Associated with Aberrant PD-1hi CD4 T cell Enrichment in the Lymphoid Follicles of the Rectal Mucosa. 30

Figure 2.1. Phenotypic characterization ofPD-1+ subsets in the blood, lymph node (LN), and rectum of healthy rhesus macaques (RM). 51

Figure 2.2. Characterization of PD-1hi CD4 T cells in the LN and rectum of chronically SIV infected vaccine-controller and non-controller RM. 52

Figure 2.3. CXCR5 expression and localization of PD-1hi CD4 T cells of chronically SIV infected vaccine-controller and non-controller RM. 53

Figure 2.4. Phenotypic characterization of PD-1hi CD4 T cells in the LN and rectum of chronically SIV infected vaccine-controller and non-controller RM. 54

Figure 2.5. Phenotypic characterization of CCR5 expression and infection status of PD-1hi CD4 T cells in the LN and rectum of chronically SIV infected vaccine-controller and non-controller RM. 56

Figure 2.6. Association between anti-viral CD8 T cells and PD-1hi CD4 T cells in the LN following SIV infection. 58

Figure 2.7. Accumulation of PD-1hi CD4 T cells in the LN of SIV infected non-controlling RM. 59

Figure 2.8. BCL-2 expression on Naïve and Memory CD4 T cells in the LN and Rectum of SIV infected macaques. 60

Figure 2.9. Accumulation of SIV infected Tfh within the LN and Rectal mucosa of SIV infected Non-controlling RM. 61

Chapter 3: Follicular Anti-viral CD8 T cells Contribute to Enhanced Control of Pathogenic SIV. 62

Figure 3.1. Rapid expansion of CXCR5+ SIV specific CD8 T cells is associated with enhanced control of chronic SIV infection. 79

Figure 3.2. CXCR5+ SIV specific CD8 T cells are more polyfunctional than CXCR5- SIV specific CD8 T cells in the blood. 80

Figure 3.3. CXCR5+ SIV-specific CD8 T cells are localized in the germinal centers (GC) of vaccinated controllers and limit Tfh expansion in vitro. 81

Figure 3.4. Global gene expression analysis revealed distinct gene expression profile for CXCR5+ SIV specific CD8 T cells. 82

Figure 3.5. Correlations between the percentage of CXCR5+ GagCM9+ CD8 T cells at week 2-3 and week 12-24 post SIV infection and set-point VL. 83

Figure 3.6. Percentage of CXCR5+ GagCM9 CD8 T cells in unvaccinated animals at week 12-24 post SIV infection inversely correlates with set-point viral load. 84

Figure 3.7. Representative FACS plots from a controller and non-controller RM at week 24 post SIV infection. 85

Figure 3.8. Representative immunofluorescence staining of CD8 T cells in LN sections from SIV+ non-controller and SIV+ vaccine controller RM at week 24 and week 2-3 post SIV infection. 86

Figure 3.9.Representative in situ tetramer staining images of MLN, spleen and rectal tissue of SIV infected controller RM. 87

Figure 3.10. Induction of perforin and granzyme co-expression on CXCR5+ and - CD8 T cells co-cultured with P11c antigen pulsed Tfh. 88

Figure 3.11. Representative histogram plots showing the expression of phenotypic markers on naïve and memory CXCR5+ and CXCR5- GagCM9 CD8 T cells, and Tfh cells. 89

Table 2. Cohort of SIV infected rhesus macaques used for sample collection . 90

Figure 3.12. Greater Frequency and Function of Germinal Center (GC) Infiltrating CXCR5+ CD8 T Cells During Controlled SIV Infection. 91

Chapter 4: In vivo PD-1 blockade as a Therapeutic. 92

Adjuvant to Antiretroviral TreatmentFigure 4.1. In vivo PD-1 ART trial design. 114

Figure 4.2. Rapid and discernable PD-1 blockade in vivo. 115

Figure 4.3. PD-1 blockade results in enhanced in vivo proliferationof CD4 and CD8 T cells. 116

Figure 4.4. PD-1 blockade results in enhanced polyfunctionality of SIV specific CD8 T cells. 117

Figure 4.5. PD-1 blockade results in enhanced functional quality of CD8 T cells. 118

Figure 4.6. In vivo PD-1 blockade synergizes with ART resulting in increased viral suppression. 119

Figure 4.7. In vivo PD-1 blockade enhances Th17 reconstitution in the rectal mucosa under suppressive ART. 120

Figure 4.8. Partial reconstitution of CD4 T cells in the blood and rectal mucosa of ART treated macaques. 121

Figure 4.9. Levels of cell associated Gag DNA within memory CD4 T cell subsets in PD-1 treated and control groups during suppressive ART. 122

Chapter 5: Discussion. 123

Early ART Initiation. 125

Targeting the Tfh Viral Reservoir. 127

HIV Sanctuary - Exposing the Germinal Center Reservoir. 131

Modulating Anti-viral Immunity under ART. 134

Biologicals. 134

Therapeutic Vaccines. 135

Cytokine based therapies and Latency Reversing Drugs (LRA's). 138

Conclusions. 141

Figure 5.1 Potential mechanisms of viral control by therapeutic vaccines. 142

References. 143

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