Background: Follicular helper CD4 T cells (TFH) are located within the germinal centers (GC) of lymph nodes (LN) and represent a major contributor to the latent reservoir. Bispecific antibodies that target HIV Env and CD3 are being developed to eliminate the latent reservoir by activating HIV from CD4 T cells and inducing killing of those cells by CD8 T cells. We characterized the localization, frequency, and function of CD8 T cells in GCs to determine if they were present and capable of killing HIV-expressing cells in the context of HIV Env/CD3-targeting bispecific antibodies.
Methods: The phenotype, localization and function of CD8 T cells in tonsils and LNs from non-infected and HIV-infected viremic individuals was investigated. Polychromatic flow cytometry was used for phenotypic analysis and confocal imaging for spacial localization. Histo-Cytometry was performed for the quantitative analysis of GC cell populations. Function (IFN, TNF, MIP-1, and Perforin, GzB production) was assessed by intracellular staining after stimulation with anti-CD3. (5 hour). In vitro cytolytic activity of sorted CD8 T cell populations was tested in a killing assay using an anti-HIV Env/anti-CD3 bispecific-antibody. Cytokines and soluble cell death mediators were analyzed by Luminex.
Results: Phenotypic analysis of tonsillar cells revealed a memory population of CD8 T cells expressing a CCR7lowCXCR5high profile compatible with follicular localization. Histo-Cytometry analysis confirmed the presence of a small population of CD8 T cells within the GC. These GC CD8 T cells were expanded in HIV-infected LNs compared to non-infected tonsils. Follicular CD8 T cells (defined by CXCR5 expression and loss of CCR7) exerted a superior killing capacity judged by in vitro mobilization of GzB/Perforin. Production of MIP-1ï¡ and GzB predominated over IFN and TNF production in GC CD8 T cells. Of all tissue CD8 T cell populations tested, GC localized CD8 T cells had the greatest ability to mediate killing of HIV-infected target cells after cross-linking with an anti-HIV Env/anti-CD3 bispecific antibody.
Conclusions: HIV infection is characterized by accumulation of CD8 T cells within LN follicles. These CD8 T cells are functionally capable of mediating bispecific antibody-mediated killing of HIV-infected CD4 T cells. These data add credence to the use of bispecific antibody therapy to purge the LN reservoir