Limited Immunogenicity of an HLA-A*03:01-restricted Epitope of Erv-k-env in Non-hiv-1 Settings: Implications for Adoptive Cell Therapy in Cancer.

Repetitive elements (REs) are often expressed at higher levels in tumor cells than normal cells, implicating these genomic regions as an untapped pool of tumor-associated antigens. In ovarian cancer (OC), protein from the RE ERV-K is frequently expressed by tumor cells. Here we determined whether the targeting of a previously identified immunogenic epitope in the envelope gene (env) of ERV-K resulted in target antigen specificity in non-HIV-1 settings. We found that transducing healthy donor T cells with an ERV-K-Env-specific T cell receptor construct resulted in antigen specificity only when co-cultured with HLA-A*03:01 B lymphoblastoid cells. Furthermore, these transduced T cells were not specific for HLA-A*03:01 + OC cells nor for the cognate peptide in HLA-matched systems from multiple healthy donors. These data suggest that the ERV-K-Env epitope recognized by this T cell receptor is of low immunogenicity and has limited potential as a T cell target for OC.

Engineered tumor-specific T cells using immunostimulatory photothermal nanoparticles.

BACKGROUND: Adoptive T cell therapy (ATCT) has been successful in treating hematological malignancies and is currently under investigation for solid-tumor therapy. In contrast to existing chimeric antigen receptor (CAR) T cell and/or antigen-specific T cell approaches, which require known targets, and responsive to the need for targeting a broad repertoire of antigens in solid tumors, we describe the first use of immunostimulatory photothermal nanoparticles to generate tumor-specific T cells. METHODS: Specifically, we subject whole tumor cells to Prussian blue nanoparticle-based photothermal therapy (PBNP-PTT) before culturing with dendritic cells (DCs), and subsequent stimulation of T cells. This strategy differs from previous approaches using tumor cell lysates because we use nanoparticles to mediate thermal and immunogenic cell death in tumor cells, rendering them enhanced antigen sources. RESULTS: In proof-of-concept studies using two glioblastoma (GBM) tumor cell lines, we first demonstrated that when PBNP-PTT was administered at a "thermal dose" targeted to induce the immunogenicity of U87 GBM cells, we effectively expanded U87-specific T cells. Further, we found that DCs cultured ex vivo with PBNP-PTT-treated U87 cells enabled 9- to 30-fold expansion of CD4+ and CD8+ T cells. Upon co-culture with target U87 cells, these T cells secreted interferon-É£ in a tumor-specific and dose-dependent manner (up to 647-fold over controls). Furthermore, T cells manufactured using PBNP-PTT ex vivo expansion elicited specific cytolytic activity against target U87 cells (donor-dependent 32-93% killing at an effector to target cell (E:T) ratio of 20:1) while sparing normal human astrocytes and peripheral blood mononuclear cells from the same donors. In contrast, T cells generated using U87 cell lysates expanded only 6- to 24-fold and killed 2- to 3-fold less U87 target cells at matched E:T ratios compared with T cell products expanded using the PBNP-PTT approach. These results were reproducible even when a different GBM cell line (SNB19) was used, wherein the PBNP-PTT-mediated approach resulted in a 7- to 39-fold expansion of T cells, which elicited 25-66% killing of the SNB19 cells at an E:T ratio of 20:1, depending on the donor. CONCLUSIONS: These findings provide proof-of-concept data supporting the use of PBNP-PTT to stimulate and expand tumor-specific T cells ex vivo for potential use as an adoptive T cell therapy approach for the treatment of patients with solid tumors.

PLGA nanodepots co-encapsulating prostratin and anti-CD25 enhance primary natural killer cell antiviral and antitumor function.

Natural killer (NK) cells are attractive effector cells of the innate immune system against human immunodeficiency virus (HIV) and cancer. However, NK cell therapies are limited by the fact that target cells evade NK cells, for example, in latent reservoirs (in HIV) or through upregulation of inhibitory signals (in cancer). To address this limitation, we describe a biodegradable nanoparticle-based "priming" approach to enhance the cytotoxic efficacy of peripheral blood mononuclear cell-derived NK cells. We present poly(lactic-co-glycolic acid) (PLGA) nanodepots (NDs) that co-encapsulate prostratin, a latency-reversing agent, and anti-CD25 (aCD25), a cell surface binding antibody, to enhance primary NK cell function against HIV and cancer. We utilize a nanoemulsion synthesis scheme to encapsulate both prostratin and aCD25 within the PLGA NDs (termed Pro-aCD25-NDs). Physicochemical characterization studies of the NDs demonstrated that our synthesis scheme resulted in stable and monodisperse Pro-aCD25-NDs. The NDs successfully released both active prostratin and anti-CD25, and with controllable release kinetics. When Pro-aCD25-NDs were administered in an in vitro model of latent HIV and acute T cell leukemia using J-Lat 10.6 cells, the NDs were observed to prime J-Lat cells resulting in significantly increased NK cell-mediated cytotoxicity compared to free prostratin plus anti-CD25, and other controls. These findings demonstrate the feasibility of using our Pro-aCD25-NDs to prime target cells for enhancing the cytotoxicity of NK cells as antiviral or antitumor agents.

Engineering the TGFß Receptor to Enhance the Therapeutic Potential of Natural Killer Cells as an Immunotherapy for Neuroblastoma.

PURPOSE: The ability of natural killer (NK) cells to lyse allogeneic targets, without the need for explicit matching or priming, makes them an attractive platform for cell-based immunotherapy. Umbilical cord blood is a practical source for generating banks of such third-party NK cells for "off-the-shelf" cell therapy applications. NK cells are highly cytolytic, and their potent antitumor effects can be rapidly triggered by a lack of HLA expression on interacting target cells, as is the case for a majority of solid tumors, including neuroblastoma. Neuroblastoma is a leading cause of pediatric cancer-related deaths and an ideal candidate for NK-cell therapy. However, the antitumor efficacy of NK cells is limited by immunosuppressive cytokines in the tumor microenvironment, such as TGFß, which impair NK cell function and survival. EXPERIMENTAL DESIGN: To overcome this, we genetically modified NK cells to express variant TGFß receptors, which couple a mutant TGFß dominant-negative receptor to NK-specific activating domains. We hypothesized that with these engineered receptors, inhibitory TGFß signals are effectively converted to activating signals. RESULTS: Modified NK cells exhibited higher cytotoxic activity against neuroblastoma in a TGFß-rich environment in vitro and superior progression-free survival in vivo, as compared with their unmodified controls. CONCLUSIONS: Our results support the development of "off-the-shelf" gene-modified NK cells, that overcome TGFß-mediated immune evasion, in patients with neuroblastoma and other TGFß-secreting malignancies.