Commensal bacteria stimulate antitumor responses via T cell cross-reactivity.

Recent studies show gut microbiota modulate antitumor immune responses; one proposed mechanism is cross-reactivity between antigens expressed in commensal bacteria and neoepitopes. We found that T cells targeting an epitope called SVYRYYGL (SVY), expressed in the commensal bacterium Bifidobacterium breve (B. breve), cross-react with a model neoantigen, SIYRYYGL (SIY). Mice lacking B. breve had decreased SVY-reactive T cells compared with B. breve-colonized mice, and the T cell response was transferable by SVY immunization or by cohousing mice without Bifidobacterium with ones colonized with Bifidobacterium. Tumors expressing the model SIY neoantigen also grew faster in mice lacking B. breve compared with Bifidobacterium-colonized animals. B. breve colonization also shaped the SVY-reactive TCR repertoire. Finally, SVY-specific T cells recognized SIY-expressing melanomas in vivo and led to decreased tumor growth and extended survival. Our work demonstrates that commensal bacteria can stimulate antitumor immune responses via cross-reactivity and how bacterial antigens affect the T cell landscape.

Engineering an Artificial T-Cell Stimulating Matrix for Immunotherapy.

T cell therapies require the removal and culture of T cells ex vivo to expand several thousand-fold. However, these cells often lose the phenotype and cytotoxic functionality for mediating effective therapeutic responses. The extracellular matrix (ECM) has been used to preserve and augment cell phenotype; however, it has not been applied to cellular immunotherapies. Here, a hyaluronic acid (HA)-based hydrogel is engineered to present the two stimulatory signals required for T-cell activation-termed an artificial T-cell stimulating matrix (aTM). It is found that biophysical properties of the aTM-stimulatory ligand density, stiffness, and ECM proteins-potentiate T cell signaling and skew phenotype of both murine and human T cells. Importantly, the combination of the ECM environment and mechanically sensitive TCR signaling from the aTM results in a rapid and robust expansion of rare, antigen-specific CD8+ T cells. Adoptive transfer of these tumor-specific cells significantly suppresses tumor growth and improves animal survival compared with T cells stimulated by traditional methods. Beyond immediate immunotherapeutic applications, demonstrating the environment influences the cellular therapeutic product delineates the importance of the ECM and provides a case study of how to engineer ECM-mimetic materials for therapeutic immune stimulation in the future.

ImmunoMap: A Bioinformatics Tool for T-cell Repertoire Analysis.

Despite a dramatic increase in T-cell receptor (TCR) sequencing, few approaches biologically parse the data in a fashion that both helps yield new information about immune responses and may guide immunotherapeutic interventions. To address this issue, we developed a method, ImmunoMap, that utilizes a sequence analysis approach inspired by phylogenetics to examine TCR repertoire relatedness. ImmunoMap analysis of the CD8 T-cell response to self-antigen (Kb-TRP2) or to a model foreign antigen (Kb-SIY) in naïve and tumor-bearing B6 mice showed differences in the T-cell repertoire of self- versus foreign antigen-specific responses, potentially reflecting immune pressure by the tumor, and also detected lymphoid organ-specific differences in TCR repertoires. When ImmunoMap was used to analyze clinical trial data of tumor-infiltrating lymphocytes from patients being treated with anti-PD-1, ImmunoMap, but not standard TCR sequence analyses, revealed a clinically predicative signature in pre- and posttherapy samples. Cancer Immunol Res; 6(2); 151-62. ©2017 AACR.

Dual Targeting Nanoparticle Stimulates the Immune System To Inhibit Tumor Growth.

We describe the development of a nanoparticle platform that overcomes the immunosuppressive tumor microenvironment. These nanoparticles are coated with two different antibodies that simultaneously block the inhibitory checkpoint PD-L1 signal and stimulate T cells via the 4-1BB co-stimulatory pathway. These "immunoswitch" particles significantly delay tumor growth and extend survival in multiple in vivo models of murine melanoma and colon cancer in comparison to the use of soluble antibodies or nanoparticles separately conjugated with the inhibitory and stimulating antibodies. Immunoswitch particles enhance effector-target cell conjugation and bypass the requirement for a priori knowledge of tumor antigens. The use of the immunoswitch nanoparticles resulted in an increased density, specificity, and in vivo functionality of tumor-infiltrating CD8+ T cells. Changes in the T cell receptor repertoire against a single tumor antigen indicate immunoswitch particles expand an effective set of T cell clones. Our data show the potential of a signal-switching approach to cancer immunotherapy that simultaneously targets two stages of the cancer immunity cycle resulting in robust antitumor activity.

CD47 Enhances In Vivo Functionality of Artificial Antigen-Presenting Cells.

PURPOSE: Artificial antigen-presenting cells, aAPC, have successfully been used to stimulate antigen-specific T-cell responses in vitro as well as in vivo. Although aAPC compare favorably with autologous dendritic cells in vitro, their effect in vivo might be diminished through rapid clearance by macrophages. Therefore, to prevent uptake and minimize clearance of aAPC by macrophages, thereby increasing in vivo functionality, we investigated the efficiency of "don't eat me" three-signal aAPC compared with classical two-signal aAPC. EXPERIMENTAL DESIGN: To generate "don't eat me" aAPC, CD47 was additionally immobilized onto classical aAPC (aAPC(CD47+)). aAPC and aAPC(CD47+) were analyzed in in vitro human primary T-cell and macrophage cocultures. In vivo efficiency was compared in a NOD/SCID T-cell proliferation and a B16-SIY melanoma model. RESULTS: This study demonstrates that aAPC(CD47+) in coculture with human macrophages show a CD47 concentration-dependent inhibition of phagocytosis, whereas their ability to generate and expand antigen-specific T cells was not affected. Furthermore, aAPC(CD47+)-generated T cells displayed equivalent killing abilities and polyfunctionality when compared with aAPC-generated T cells. In addition, in vivo studies demonstrated an enhanced stimulatory capacity and tumor inhibition of aAPC(CD47+) over normal aAPC in conjunction with diverging biodistribution in different organs. CONCLUSIONS: Our data for the first time show that aAPC functionalized with CD47 maintain their stimulatory capacity in vitro and demonstrate enhanced in vivo efficiency. Thus, these next-generation aAPC(CD47+) have a unique potential to enhance the application of the aAPC technology for future immunotherapy approaches.

Sprouty-2 regulates HIV-specific T cell polyfunctionality.

The ability of individual T cells to perform multiple effector functions is crucial for protective immunity against viruses and cancer. This polyfunctionality is frequently lost during chronic infections; however, the molecular mechanisms driving T cell polyfunctionality are poorly understood. We found that human T cells stimulated by a high concentration of antigen lacked polyfunctionality and expressed a transcription profile similar to that of exhausted T cells. One specific pathway implicated by the transcription profile in control of T cell polyfunctionality was the MAPK/ERK pathway. This pathway was altered in response to different antigen concentrations, and polyfunctionality correlated with upregulation of phosphorylated ERK. T cells that were stimulated with a high concentration of antigen upregulated sprouty-2 (SPRY2), a negative regulator of the MAPK/ERK pathway. The clinical relevance of SPRY2 was confirmed by examining SPRY2 expression in HIV-specific T cells, where high levels of SPRY2 were seen in HIV-specific T cells and inhibition of SPRY2 expression enhanced the HIV-specific polyfunctional response independently of the PD-1 pathway. Our findings indicate that increased SPRY2 expression during chronic viral infection reduces T cell polyfunctionality and identify SPRY2 as a potential target for immunotherapy.