The major obstacle to a cure for HIV infection is the persistence of replication-competent viral reservoirs during antiretroviral therapy. HIV-specific chimeric antigen receptor (CAR) T cells have been developed to target latently infected CD4+ T cells that express virus either spontaneously or after intentional latency reversal. Whether HIV-specific CAR-T cells can recognize and eliminate the follicular dendritic cell (FDC) reservoir of HIV-bound immune complexes (ICs) is unknown. We created HIV-specific CAR-T cells using human peripheral blood mononuclear cells (PBMCs) and a CAR construct that enables the expression of CD4 (domains 1 and 2) and the carbohydrate recognition domain of mannose binding lectin (MBL) to target native HIV Env (CD4-MBL CAR). We assessed CAR-T cell cytotoxicity using a carboxyfluorescein succinimidyl ester (CFSE) release assay and evaluated CAR-T cell activation through interferon gamma (IFN-γ) production and CD107a membrane accumulation by flow cytometry. CD4-MBL CAR-T cells displayed potent lytic and functional responses to Env-expressing cell lines and HIV-infected CD4+ T cells but were ineffective at targeting FDC bearing HIV-ICs. CD4-MBL CAR-T cells were unresponsive to cell-free HIV or concentrated, immobilized HIV-ICs in cell-free experiments. Blocking intercellular adhesion molecule-1 (ICAM-1) inhibited the cytolytic response of CD4-MBL CAR-T cells to Env-expressing cell lines and HIV-infected CD4+ T cells, suggesting that factors such as adhesion molecules are necessary for the stabilization of the CAR-Env interaction to elicit a cytotoxic response. Thus, CD4-MBL CAR-T cells are unable to eliminate the FDC-associated HIV reservoir, and alternative strategies to eradicate this reservoir must be sought. IMPORTANCE Efforts to cure HIV infection have focused primarily on the elimination of latently infected CD4+ T cells. Few studies have addressed the unique reservoir of infectious HIV that exists on follicular dendritic cells (FDCs), persists in vivo during antiretroviral therapy, and likely contributes to viral rebound upon cessation of antiretroviral therapy. We assessed the efficacy of a novel HIV-specific chimeric antigen receptor (CAR) T cell to target both HIV-infected CD4+ T cells and the FDC reservoir in vitro. Although CAR-T cells eliminated CD4+ T cells that express HIV, they did not respond to or eliminate FDC bound to HIV. These findings reveal a fundamental limitation to CAR-T cell therapy to eradicate HIV.

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Purpose of review-T cells within B cell follicles of secondary lymphoid tissues play key roles in HIV immunopathogenesis. This review highlights recent findings and identifies gaps in current knowledge. Recent findings-B cell follicles are major sites of virus replication and demonstrate significant impairments in generation of humoral immunity in HIV infection. Follicular T helper cells (Tfh), follicular T regulatory cells (Tfr) and follicular CD8 T cells (fCD8) play key roles in HIV immunopathogenesis. Tfh and more recently Tfr are highly permissive to HIV, and may serve as reservoirs of HIV in treated infection. Virus-specific CD8 T cells are less abundant in B cell follicles than extrafollicular regions, but their effector mechanisms remain an area of significant controversy. Impairments in Tfh likely contribute to impaired humoral immunity and potential mechanisms include B cell counter-regulatory mechanisms, Tfr suppression, and diminished repertoire breadth. A better understanding of the roles of Tfh, Tfr and fCD8 in HIV immunopathogenesis is critical to development of effective HIV vaccines and cure strategies. Summary-Tfh, Tfr, and fCD8 contribute to HIV persistence and impaired humoral immunity. A better understanding of their roles could facilitate vaccine development and HIV cure strategies.

A major barrier in the use of humanized mice as models of HIV-1 (HIV) infection is the inadequate generation of virus-specific antibody responses. Humanized DRAGA (hDRAGA) mice generate antigen-specific class switched antibodies to several pathogens, but whether they do so in HIV infection and the extent to which their secondary lymphoid tissues (sLT) support germinal center responses is unknown. hDRAGA mice were evaluated for their ability to support HIV replication, generate virus-specific antibody responses, develop splenocyte subsets, and organize sLT architecture. hDRAGA mice supported persistent HIV replication and developed modest levels of gp41-specific human IgM and IgG. Spleens from uninfected and HIV infected hDRAGA mice contained differentiated B and CD4+ T cell subsets including germinal center (GC) B cells and T follicular helper cells (TFH); relative expansions of TFH and CD8+ T cells, but not GC B cells, occurred in HIV-infected hDRAGA mice compared to uninfected animals. Immunofluorescent staining of spleen and mesenteric lymph node sections demonstrated atypical morphology. Most CD4+ and CD8+ T cells resided within CD20hi areas. CD20hi areas lacked canonical germinal centers, as defined by staining for IgD-Ki67+cells. No human follicular dendritic cells (FDC) were detected. Mouse FDC were distributed broadly throughout both CD20hi and CD20lo regions of sLT. HIV RNA particles were detected by in situ hybridization within CD20+ areas and some co-localized with mouse FDC. Viral RNA+ cells were more concentrated within CD20hi compared to CD20lo areas of sLT, but differences were diminished in spleen and eliminated in mesenteric lymph nodes when adjusted for CD4+ cell frequency. Thus, hDRAGA mice recapitulated multiple aspects of HIV pathogenesis including HIV replication, relative expansions in TFH and CD8+ T cells, and modest HIV-specific antibody production. Nevertheless, classical germinal center morphology in sLT was not observed, which may account for the inefficient expansion of GC B cells and generation of low titer human antibody responses to HIV-1 in this model.

Follicular dendritic cells (FDCs) are found in all secondary lymphoid tissues, where they function as a repository of antigens to maintain long‐term IgG and IgE responses. Antigens are trapped and retained on FDCs in the form of immune complexes; and while most immune complexes require large quantities to induce an immune response, FDC‐trapped antigens are remarkably immunogenic and only a few picogram can induce microgram concentrations of a specific antibody. In addition to providing antigens, FDCs provide a number of additional signals (e.g. BAFF, IL‐6) that further contribute to antibody production. In addition to their contributions to immunity in health, FDCs are involved in some pathological situations including HIV/AIDS (human immunodeficiency virus/acquired immune deficiency syndrome), sarcoma/lymphoma, prion‐mediated transmissible spongiform encephalopathies (e.g. Creutzfeldt–Jakob) and Castleman disease. A further understanding of FDCs and their functions in both health and disease may aid our ability to better regulate immunity and ameliorate some disease states. Key Concepts FDCs trap antigens as immune complexes that consist of antigen in the presence of either specific antibodies or complement proteins, or both. FDCs trap immune complexes using CD32 and/or CD21. FDC‐trapped antigens or iccosomes are highly immunogenic and minute amounts (picogram) can induce significant quantities (microgram) of a specific antibody. FDC‐trapped antigens remain on the surface of FDCs and are not internalised. These antigens are not degraded but maintain their native configuration and immunoreactivity for many months. In addition to trapping conventional antigens, FDCs also trap HIV (and potentially other viruses) and maintain the infectious nature for many months. FDC‐trapped HIV can transmit infection to adjacent target cells (e.g. CD4 + T lymphocytes). FDCs provide both antigen and other signals that are central to the induction and maintenance of specific antibody responses. FDCs can play roles in both health and disease (e.g. HIV/AIDS, prion diseases and follicular lymphomas).