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.

T cell therapies require the removal and culture of T cells ex vivo to expand several thousandfold. However, these cells often lose the phenotype and cytotoxic functionality necessary to mediate an effective therapeutic response. The extracellular matrix has been used to preserve and augment cell phenotype; however, it has not been applied to cellular immunotherapies.Here we engineered a hyaluronic acid (HA)-based hydrogel to present the two stimulatory signals required for T-cell activation-termed an artificial T cell stimulating matrix (aTM).We found that biophysical properties of the aTM-stimulatory ligand density, stiffness, and extracellular matrix (ECM) proteins-potentiate T cell signaling and skew phenotype of both murine and human T cells. Importantly, it was only the combination of the ECM environment and mechanically sensitive TCR signaling from the aTM that produced nearly 4-times the number of rare, antigen-specific CD8+ T cells. Adoptive transfer of these tumor-specific cells significantly suppressed tumor growth and improved animal survival as compared with T cells stimulated by traditional methods. Beyond immediate immunotherapeutic applications, demonstrating that 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.

and Eisai. TAC has served as an advisor for Bristol Myers Squibb, Illumina, Eisai, and An2H. Under a licensing agreement between NexImmune and the Johns Hopkins University, JPS is entitled to shares of royalty received by the university on sales of artificial antigen-presenting cell products described in this article. He also owns NexImmune stock, which is subject to certain restrictions under university policy. JPS is a member of the company's Scientific Advisory Board. The terms of this arrangement are being managed by the Johns Hopkins University in accordance with its conflict-of-interest policies. JPS acknowledges grant funding from AstraZeneca.

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 (K-TRP2) or to a model foreign antigen (K-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. .