Novel Immunogens and Mucosal Vaccination For Improving Protection Against HIV-1 Open Access

Jones, Andrew (Spring 2018)

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Despite continuous efforts since its initial discovery, an effective prophylactic vaccine against Human Immunodeficiency Virus (HIV) -1 has yet to be developed.  Insights from vaccine efficacy trials in both animal models and humans, as well as basic research to understand the complexities of HIV-1, have greatly informed current efforts in HIV-1 vaccine design.  Current efforts are focused on the design and characterization of novel immunogens that promote protective antibody responses.  Many immunogens are designed to better recapitulate the trimeric structure of the native HIV-1 Envelope, composed of heterotrimers of gp120 and gp41, which mediates viral entry.

            This dissertation will characterize the immunogenicity and efficacy of a novel trimeric HIV-1 gp120 immunogen, named cycP-gp120, in rabbits and rhesus macaques.  We demonstrate cycP-gp120 to be robust vaccine immunogen, generating high titers of cross-reactive antibodies targeting the variable loops 1 and 2 (V1V2) of gp120, a key correlate of protection in the RV144 vaccine efficacy trial in humans.  We show that cycP-gp120 in conjunction with priming with the poxvirus vector modified vaccinia Ankara (MVA-HIV) promotes both V1V2-directed antibodies and antibody-dependent cellular cytotoxicity (ADCC) in rabbits.

            As HIV-1 is primarily transmitted across mucosal surfaces via sexual contact, generating mucosal immunity to HIV-1 is thought to be a crucial component to an effective HIV-1 vaccine.  Here we describe a novel method of oral immunization in which we immunized the sublingual and buccal tissue of rhesus macaques with a needle-free injector.  Needle-free immunization resulted in a large expansion of both systemic and mucosally localized vaccine-specific antibodies, demonstrating this route as an easy and effective method of mucosal vaccination.

            To test the protective efficacy of MVA-HIV/cycP-gp120 immunization in rhesus macaques, as well as the needle-free oral vaccination route, we challenged immunized animals with a weekly low dose of pathogenic SHIV-SF162P3.  We observed significant protection in rhesus macaques immunized with via needle-free oral immunization or intradermally/subcutaneously.  Protective efficacy correlated with anti-gp120 binding IgG, anti-V2 peptide antibodies, and antibody-dependent cell-mediated viral inhibition (ADCVI).  These studies thus describe cycP-gp120 as an attractive vaccine immunogen for further studies in non-human primates and humans, as well as the needle-free oral immunization route as mucosal vaccination technique.    

Table of Contents

Chapter 1:  Introduction ---------------------------------------------------------  1

Virology and Pathogenesis of Human Immunodeficiency Virus-1 (HIV-1) -------------   2

Protective Immunity to HIV-1:  Lessons from Vaccine Trials ----------------------------- 10 Current HIV-1 Vaccine Strategies:  Development of Novel Immunogens                                                            and Prime-Boost Regimens --------------------------------------------------------------------- 22

Mucosal Vaccine Development for HIV-1 --------------------------------------------------- 29

Summary of Introduction ----------------------------------------------------------------------- 36

Figures and Legends -------------------------------------------------------------------

Fig 1. Structure of HIV-1 --------------------------------------------------------------- 37

Fig 2. Structure of HIV-1 Envelope --------------------------------------------------- 38

Fig 3. Neutralizing and non-neutralizing antibody effector functions ------------ 39

Fig 4. Timeline of HIV-1 Vaccine Efficacy Trials ---------------------------------- 40

Fig 5. The V1V2 region of HIV-1 gp120 --------------------------------------------- 41

Fig 6. The V2-hotspot region of HIV-1 gp120 --------------------------------------- 42

Fig 7. Generation of trimeric gp120 (cycP-gp120) ---------------------------------- 43

Fig 8. V1V2 loops in cycP-gp120 -----------------------------------------------------  44

Fig 9. MVA-HIV expresses trimeric HIV-1 Env ------------------------------------  45

Chapter II:  A trimeric HIV-1 envelope gp120 induces potent and broad anti-V1V2 loop antibodies against HIV-1 in rabbits and rhesus macaques -------------  46

            Abstract -------------------------------------------------------------------------------------------  47

            Importance ----------------------------------------------------------------------------------------  48

Introduction ---------------------------------------------------------------------------------------  49

            Results ---------------------------------------------------------------------------------------------  52

            Discussion -----------------------------------------------------------------------------------------  61 

            Methods--------------------------------------------------------------------------------------------- 66

            Acknowledgements ------------------------------------------------------------------------------- 71

            Figures and Legends -------------------------------------------------------------------------

Fig 1. CycP-gp120 induces high titers of high avidity anti-gp120 and                                                      tier-1A and -1B neutralizing antibodies in rabbits -----------------------------------  72

Fig 2. Binding Antibody Multiplex Assay (BAMA) analysis of rabbit immune                                       sera reactivity to a global panel of HIV-1 gp120, gp140,                                                                         and gp70-V1V2 scaffolds ---------------------------------------------------------------- 74

Fig 3. V2-directed antibodies generated by cycP-gp120 ----------------------------  75

Fig 4. Boosting MVA-primed rabbits with cycP-gp120 promotes neutralizing                             antibodies, increased gp120/gp140, and gp70-V1V2 directed antibodies --------- 76

Fig 5. ADCC activity in rabbits immunized with cycP-gp120 or                                                          MVA-HIV prime cycP-gp120 boost ---------------------------------------------------- 77

Fig 6. Boosting rhesus macaques primed with a DNA/MVA vaccine                                                      with cycP-gp120 --------------------------------------------------------------------------- 78

Table 1. Neutralizing antibody titers against tier-1A, -1B, and -2 HIV isolates -  79

Chapter III: Systemic or Oral MVA/Protein HIV-1 immunization with a needle-free injector induces protective antibody responses in rhesus macaques -------  80

Abstract --------------------------------------------------------------------------------------------  81

Introduction ---------------------------------------------------------------------------------------- 83

            Results ---------------------------------------------------------------------------------------------  88

            Discussion ------------------------------------------------------------------------------------------ 99

            Methods-------------------------------------------------------------------------------------------- 105

   Figures and Legends ---------------------------------------------------------------------------

Fig 1. Dendritic Cells in the Sublingual and Buccal Tissue------------------------- 110

Fig 2. Detection of aldehyde dehydrogenase (aldh) activity in                                                          dendritic cells in oral tissue and lymph nodes --------------------------------------- 111

Fig 3. Sublingual and buccal immunization with a needle-free injector ----------112

Fig 4. Vaccine Scheme ------------------------------------------------------------------ 113

Fig 5. Anti-gp120 antibodies in the sera ---------------------------------------------- 114

Fig 6. Anti-gp120 antibodies in the mucosal secretions ---------------------------- 115

Fig 7. Neutralizing Antibodies --------------------------------------------------------- 116

Fig 8. Anti-Env breadth and cross-reactivity ----------------------------------------- 117

Fig 9. Anti-V1V2 antibody responses ------------------------------------------------- 118

Fig 10. Anti-V2 hotspot antibodies ---------------------------------------------------- 119

Fig 11. Intra-rectal challenge with pathogenic SHIV-SF162P3 -------------------  120

Fig 12. Immune-correlates of protection ---------------------------------------------- 121

Fig 13. Binding to B.6240 gp120 correlates with protection ----------------------- 122

Fig 14. Antibody dependent cell-mediated viral inhibition                                                              correlates with protection --------------------------------------------------------------- 123

Fig 15. Anti-dmlt antibodies generated by immunization with                                                               cycP-gp120 and dmLT -------------------------------------------------------------------124

Fig 16. Serum and rectal IgA responses do not correlate with protection -------- 125

Fig. 17.  Peak CD4+ T-cell responses and phenotype of rectal                                                              CD4+ T-cells at the pre-challenge time-point ---------------------------------------- 126

Chapter IV: Discussion -----------------------------------------------------------------------   127

            The RV144 trial and V1V2-directed antibody responses ----------------------------------  128

            Neutralizing vs non-neutralizing antibody responses --------------------------------------- 132

            Mucosal vaccination for HIV-1 via sublingual and buccal immunization --------------- 139

References ------------------------------------------------------------------------------- 142

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