Hematopoietic Stem Cell-Derived Chimeric Antigen Receptor Expressing Cells Traffic to HIV Reservoir Sites in SHIV-Infected Non-Human Primates

Thesis (Master's)--University of Washington, 2019The successful treatment of 2 HIV+ individuals with allogeneic stem cell transplantations has garnered much interest in its utility as a therapy for HIV. However, there are significant limitations that prevent allogeneic stem cell transplantation from being a viable therapy for HIV infection, including toxicity due to Graft-versus-host disease, and the limited availability of donors who are homozygous for the CCR5Δ32 mutation. One viable option to circumvent these limitations and provide a functional cure, through immune-mediated elimination of the virus, is with chimeric antigen receptor (CAR) immunotherapy. By introducing the CAR engineered to target HIV infected cells into hematopoietic stem cells, we have the potential to generate a self-renewing population of CAR T-cells, thus circumventing some of the limitations with longevity that are seen with CAR T-cells. Here, we report trafficking of hematopoietic stem and progenitor (HSPC)-derived CAR+ cells to HIV tissue reservoir sites, in a pigtail macaque model of HIV infection. CAR+ cells were identified in the lymphoid germinal centers, the parenchyma of the central nervous system, and the gastrointestinal tract by immunohistochemistry, almost 2 years after initial engraftment with lentiviral modified HSPCs. Multilineage engraftment of CAR+ cells were identified in lymphoid germinal centers and the gastrointestinal tract, consisting of T-cells, B-cells, and myeloid lineage cells. CAR+ B-cells in the germinal centers were also actively replicating, characterized by robust Ki-67 expression. No difference was observed in trafficking, engraftment, and cell cycle activity between CAR animals and control animals that carry a “tailless” CAR which lacks the cytoplasmic signal transduction domain. These results show the HSPC-derived CAR+ cells will traffic to, and persist in, HIV tissue reservoir sites, with multilineage engraftment of CAR+ cells. The method could be used as a platform for immunotherapeutic delivery treatment of HIV infection, as well as a variety of other medical conditions

Phenotypic and functional characterization of pharmacologically expanded Vγ9Vδ2 T cells in pigtail macaques

Summary: While gaining interest as treatment for cancer and infectious disease, the clinical efficacy of Vγ9Vδ2 T cell-based immunotherapeutics has to date been limited. An improved understanding of γδ T cell heterogeneity across lymphoid and non-lymphoid tissues, before and after pharmacological expansion, is required. Here, we describe the phenotype and tissue distribution of Vγ9Vδ2 T cells at steady state and following in vivo pharmacological expansion in pigtail macaques. Intravenous phosphoantigen administration with subcutaneous rhIL-2 drove robust expansion of Vγ9Vδ2 T cells in blood and pulmonary mucosa, while expansion was confined to the pulmonary mucosa following intratracheal antigen administration. Peripheral blood Vγ9Vδ2 T cell expansion was polyclonal, and associated with a significant loss of CCR6 expression due to IL-2-mediated receptor downregulation. Overall, we show the tissue distribution and phenotype of in vivo pharmacologically expanded Vγ9Vδ2 T cells can be altered based on the antigen administration route, with implications for tissue trafficking and the clinical efficacy of Vγ9Vδ2 T cell immunotherapeutics

Stem cell-derived CAR T cells traffic to HIV reservoirs in macaques.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) with CCR5- donor cells is the only treatment known to cure HIV-1 in patients with underlying malignancy. This is likely due to a donor cell-mediated graft-versus-host effect targeting HIV reservoirs. Allo-HSCT would not be an acceptable therapy for most people living with HIV due to the transplant-related side effects. Chimeric antigen receptor (CAR) immunotherapies specifically traffic to malignant lymphoid tissues (lymphomas) and, in some settings, are able to replace allo-HSCT. Here, we quantified the engraftment of HSC-derived, virus-directed CAR T cells within HIV reservoirs in a macaque model of HIV infection, using potentially novel IHC assays. HSC-derived CAR cells trafficked to and displayed multilineage engraftment within tissue-associated viral reservoirs, persisting for nearly 2 years in lymphoid germinal centers, the brain, and the gastrointestinal tract. Our findings demonstrate that HSC-derived CAR+ cells reside long-term and proliferate in numerous tissues relevant for HIV infection and cancer

Stem cell–derived CAR T cells traffic to HIV reservoirs in macaques

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) with CCR5– donor cells is the only treatment known to cure HIV-1 in patients with underlying malignancy. This is likely due to a donor cell–mediated graft-versus-host effect targeting HIV reservoirs. Allo-HSCT would not be an acceptable therapy for most people living with HIV due to the transplant-related side effects. Chimeric antigen receptor (CAR) immunotherapies specifically traffic to malignant lymphoid tissues (lymphomas) and, in some settings, are able to replace allo-HSCT. Here, we quantified the engraftment of HSC-derived, virus-directed CAR T cells within HIV reservoirs in a macaque model of HIV infection, using potentially novel IHC assays. HSC-derived CAR cells trafficked to and displayed multilineage engraftment within tissue-associated viral reservoirs, persisting for nearly 2 years in lymphoid germinal centers, the brain, and the gastrointestinal tract. Our findings demonstrate that HSC-derived CAR+ cells reside long-term and proliferate in numerous tissues relevant for HIV infection and cancer