Date

4-8-2010

UMMS Affiliation

Graduate School of Biomedical Sciences, Immunology and Virology Program

Document Type

Dissertation, Doctoral

Subjects

AIDS Vaccines; HIV Antibodies; HIV-1; Vaccines, DNA; env Gene Products, Human Immunodeficiency Virus; Dissertations, UMMS

Disciplines

Immunology and Infectious Disease | Life Sciences | Medicine and Health Sciences | Virology

Abstract

The best known correlate of protection provided by vaccines is the presence of pathogen specific antibodies after immunization. However, against the Human Immunodeficiency Virus-1 (HIV-1) the mere presence of antibodies specific for the viral Envelope (Env) protein is not sufficient to provide protection. This necessitates in depth study of the humoral responses elicited during infection and by vaccination. While a significant amount of effort has been invested in studying the evolution of antibody responses to viral infection, only limited progress in understanding antibody responses elicited through vaccination has been made. In the studies described here, I attempt to rectify this deficiency by investigating how the quality of a humoral response is altered with the use of different immunization regimens, in particular a DNA prime-protein boost regimen, or with the use of different model HIV-1 Env gp120 immunogens. In a New Zealand White (NZW) rabbit model, we demonstrate that the broader neutralizing activity elicited with the DNA prime-protein boost regimen may be the result of the elicitation of a higher avidity antibody response and a unique profile of antibody specificities. Specifically, use of a DNA prime-protein boost regimen elicits antibodies targeted to the CD4 binding domain of the HIV-1 Env, a specificity that was not frequently observed when only protein based immunizations were administered.

We extended this analysis to sera from healthy human volunteers who participated in early phase HIV vaccine trials utilizing either a protein alone immunization regimen, a canarypox prime-protein boost immunization regimen, or a DNA prime-protein boost immunization regimen. Evaluation of sera from these trials demonstrated that the use of a DNA prime-protein boost regimen results in an antibody response with greater neutralization breadth characterized by an increased frequency and titer of antibodies targeted toward the CD4 binding site (CD4bs). In addition to this, the antibody response elicited by the DNA prime-protein boost regimen also exhibited the capability to mediate antibody dependent cell-mediated cytotoxicity (ADCC) activity as well as activation of the complement system.

Additionally, in an attempt to better understand the capabilities of antibodies elicited by a DNA prime-protein boost regimen, we generated gp120 specific monoclonal antibodies (mAbs) from a single DNA primed-protein boosted NZW rabbit. Analysis of mAbs produced from this animal revealed that use of this immunization regimen elicits an antibody repertoire with diverse epitope specificity and cross reactivity. Furthermore, these select mAbs are capable of neutralizing heterologous HIV isolates. Further application of mAb generation in rabbits may provide a valuable tool to study immunogenicity of different vaccines and immunization regimens.

Concurrently, while demonstrating that a DNA prime-protein boost regimen elicits a higher quality antibody response than that observed with other leading techniques, we also demonstrated that immunogen selection can play a vital role in the quality of the resulting antibody response. By immunizing with two closely related but phenotypically distinct model gp120 immunogens, known as B33 and LN40, we demonstrated that disparate gp120s have different intrinsic abilities to raise a heterologous neutralizing antibody response. Additionally, we showed that residues found within and flanking the b12 and CD4 binding sites play critical roles in modulating neutralizing activity of sera from animals immunized with LN40 gp120, indicating that the broader neutralizing activity seen with this immunogen may be due to differential elicitation of antibodies to this domain.