Summary:

Cytotoxic T-lymphocytes will be a primary effecter in any successful HIV-1 vaccine. Knowledge of the protective CTL epitopes and their cross-subtype effectiveness is incomplete. The study proposes to use the setting of natural infection in individuals, multiply exposed to HIV subtypes A, C and D, to gain understanding of a protective CTL response. From a high-risk cohort in Tanzania, three groups will be identified: persistently seronegative, persistently infected with a single HIV subtype or recombinant, and superinfected with a second subtype. CTL ELISPOT and epitope mapping, viral load and diversity, HLA typing, and MHA (a new subtyping tool) will be employed. This study will be the first to define the CTL response capable of protecting from new HIV infections in persistent seronegatives and, in the same population and setting, to identify the flawed CTL responses that permit superinfection of already infected individuals. Personnel and infrastructure development will facilitate future vaccine trials.

Objectives:

The broad objective of this proposal is to investigate the hypothesis that the quality, breadth, and specificity of the CTL response against HIV-1 is a key determinant in protection, both from an initial HIV-1 infection, and from superinfection with a second HIV-1 subtype. The study will assess the quality of the CTL response, together with related parameters, such as viral load and diversity, HLA type, and, in the case of superinfection, the recognised CTL epitopes in the strains causing the initial infection and superinfection, respectively. A high-risk cohort in Tanzania exposed to HIV-1 subtypes A, C, and D will provide the clinical materials for evaluation. After three years of longitudinal follow-up, 30 individuals in each of the three categories will be studied: highly exposed uninfected, persistently singly infected and superinfected. Definition of protective CTL responses, and a direct evaluation of cross-subtype immunity, will guide the design and evaluation of candidate vaccines in the region. Infrastructure and personnel development in Tanzania during this project will also materially support preparation for candidate vaccine evaluation.

Description:

The proposed study will use samples that are collected during an ongoing cohort study of 600 bar workers at high risk for HIV infection that have been followed up on  quarterly since October 2000. Three different subgroups of the cohort will be studied. Highly exposed persistent seronegative indi.viduals (HEPS), HIV infected individuals that could resist HIV superinfection with a second strain (PSin), and individuals that became superinfected with a second HIV strain despite a immunological response against the initial HIV (SupIn). The individuals will be assigned to these subgroups based on the results of a newly developed assay (MHA) that screens in multiple genome regions for the presence of more than one HIV subtype. In each group, HLA type and CTL response to vaccinia-expressed HIV-1 gag, pol, env, and nef from subtypes A,C, and D will be measured using ELISPOT and, for proteins for which a repose is detected, epitope mapping.. Longitudinal evaluation of CTL response and epitope recognition will be conducted based on the initial cross-sectional determination. Viral load using a subtype-insensitive assay and the diversity of the viral quasispecies determined by a heteroduplex tracking assay will give a picture of the effectiveness of the CTL response in controlling viral load and diversity in each individual. The quarterly blood samples provide close scrutiny of the timing of infection in HEPS and of superinfection, which will in turn provide essential materials for determination of the CTL response at critical times. Sequencing of viral strains around the times of changing HIV-1 status, either from negative to positive, or from positive to superinfected, will help determine whether CTL escape variants play a role in the failure of protective immunity. Persistent single infections will provide a catalogue of CTL responses that may have been effective in the prevention of a second infection. Finally, the genetic analysis of superinfecting strains in relationship to the initial infection determined first by MHA and then by sequencing from PBMC DNA and plasma, will give concrete information about cross-subtype effectiveness of specific CTL responses. Much of the work will be conducted in laboratories in Tanzania and South Africa.

Milestones:

1) Identification of CTL responses capable of protecting from HIV-1 infection.
2) Identification of CTL responses capable of protecting from superinfection.
3) Identification of effective cross-subtype CTL responses.
4) Relationship of HLA type to effective CTL responses.
5) Investigation of viral load and/or viral diversity as surrogate markers of effective CTL responses.
6) Role of CTL escape in infection and superinfection.
7) Definition of protective epitopes for inclusion in candidate vaccines.
8) Development of laboratory capacity for HIV-1 vaccine evaluation in Tanzania.

Coordinator:

Partners:

2. Regional Medical Office of the Tanzanian Ministry of Health - Mbeya Research Board Mbeya, Tanzania
3. National Institute for Virology - Aids Research Unit of National Institute for Virology Johannesburg, South Africa
4. University of Cape Town - Division of Medical Virology - Faculty of Health Sciences Cape Town, South Africa
5. Kansanterveyslaitos, National Public Health Institute - Department of Infectious Disease Epidemiology, FI-Helsinki, Finland