Michelle C. Sabo, Ph.D. - US grants

DESCRIPTION (provided by applicant): Hepatitis C virus is a chronic, viral disease that infects the liver. Infection is transmitted by injection drug use or contaminated blood products, and approximately 80% of infected individuals become chronically infected. Chronic HCV infection is associated with an increased risk of liver cirrhosis and hepatocellular carcinoma. Indeed, HCV is the leading cause of liver transplantation in the United States. Currently, there is no vaccine for HCV and the treatment regimen, pegylated Interferon-?2b and ribavirin, is only effective in ~50% of patients. Due to a lack of small animal model and, until recently, a cell culture adapted virus, little is known about the immune response against HCV infection. HCV Envelope-2 (E2) is expressed on the surface of the virion and is thought to be important in mediating viral attachment and entry. Neutralizing antibodies against the E2 protein are found in infected patients, although the mechanisms by which these antibodies act and the epitopes to which they bind remains poorly understood. An improved understanding of how anti-E2 antibodies neutralize infection and identification of the important neutralizing epitopes would provide a framework for the future development of vaccines and novel therapeutics against HCV. The objective of this proposal is to study the mechanism of antibody neutralization directed against HCV E2 and to identify the antibody binding regions within the protein that are important in controlling infection. To achieve this objective, we propose two specific aims. In the first aim we propose to test the neutralization capabilities of a large panel of anti-E2 antibodies in vitro. We will test the ability of HCV E2 antibodies to interfere with different phases of the viral life cycle, to block infection of viral particles of different densities, and to interact with the complement system to enhance neutralization of infection. This aim will enable us to better understand how antibodies function to limit HCV infection. In our second aim, we propose to investigate the molecular mechanism of antibody-mediated neutralization of HCV E2. In this aim we plan to map the critical antibody binding residues within E2 and to test antibodies for their ability to inhibit HCV binding to CD81. This will provide a more thorough understanding of the critical residues within E2 that are necessary for viral entry. Together, these aims will allow us to generate a better understanding of how HCV neutralizing antibodies inhibit infection and whether specific viral epitopes can be used as targets for vaccine development or the generation of novel therapeutics. PUBLIC HEALTH RELEVANCE: Hepatitis C Virus is a blood born pathogen that results in chronic disease in ~80% of infected individuals and is associated with an increased risk of cirrhosis and hepatocellular carcinoma. Currently there is no vaccine for HCV and the available treatment is only effective in ~50% of patients. We intend to study the mechanism by which antibodies provide protection from HCV infection to better inform vaccine and therapeutic development.

ABSTRACT Summary: This proposal supports a five-year training program for Dr. Sabo to gain the skills she needs to become an independent physician-scientist and an expert in reproductive immunology. Dr. Sabo has a background in basic science and immunology, and plans to shift her focus to clinically-oriented, translational research. She has developed a set of aims that align with her research interests, and six core learning objectives to help her make this transition. Research plan: Bacterial vaginosis (BV) and sub-optimal vaginal bacterial taxa have been associated with HIV acquisition. The precise mechanism by which this occurs is unknown, but may be related to increased cervical inflammation. Dendritic cells (DCs) are critical for regulating the inflammatory state of mucosal tissues, and likely play a role in genital acquisition of HIV. However, DCs remain minimally characterized in the cervix. The purpose of this proposal is to examine if sub-optimal vaginal bacteria and increased vaginal bacterial species diversity are associated with increased total cervical DCs. We have proposed three aims to answer this question. For AIMS 1 and 2, we will perform a cross sectional study of female sex workers (FSWs) enrolled in a longitudinal, open cohort study in Mombasa, Kenya. Vaginal swabs and cervical biopsy samples will be collected at a single visit. To determine if high-risk bacterial taxa are associated with increased DC number (AIM 1), we will evaluate the association between concentrations of vaginal bacteria (measured by quantitative PCR [qPCR]) and total numbers of cervical DCs (measured by flow cytometry from tissue digests of cervical biopsies). To examine the relationship between vaginal bacterial species diversity and cervical DCs (AIM 2), we will perform broad range PCR with high throughput sequencing of vaginal swabs to calculate the Shannon Diversity Index (SDI) for each patient, and the SDI will be compared to the number of DCs isolated by flow cytometry. In AIM 3, we will test the hypothesis that treatment of BV reduces the number of cervical DCs. As an exploratory outcome, DC activation will be assessed by measurement of cell surface co-stimulatory markers and intracellular pro-inflammatory cytokines for all aims. Together, these aims have the potential to provide evidence for a mechanistic link between sub-optimal vaginal microbiota, cervical inflammation, and HIV susceptibility in women. Training plan: The six core learning objectives for this K23 award period include training in epidemiology, clinical studies, global health, high dimensional data analysis, mucosal immunology, and the vaginal microbiome. These goals were selected to ensure that Dr. Sabo acquires the skills necessary to lead her own clinical studies and launch an independent research career performing translational studies to elucidate the role of the reproductive immune system in health and disease. Training to complete the six key learning objectives will be achieved through a combination of mentorship, coursework, implementation of the proposed research plan, local seminars, and national and international meetings. .