Melinda Mata

Antitumor effects of chimeric receptor engineered human T cells directed to tumor stroma.

Cancer-associated fibroblasts (CAFs), the principle component of the tumor-associated stroma, form a highly protumorigenic and immunosuppressive microenvironment that mediates therapeutic resistance. Co-targeting CAFs in addition to cancer cells may therefore augment the antitumor response. Fibroblast activation protein-α (FAP), a type 2 dipeptidyl peptidase, is expressed on CAFs in a majority of solid tumors making it an attractive immunotherapeutic target. To target FAP-positive CAFs in the tumor-associated stroma, we genetically modified T cells to express a FAP-specific chimeric antigen receptor (CAR). The resulting FAP-specific T cells recognized and killed FAP-positive target cells as determined by proinflammatory cytokine release and target cell lysis. In an established A549 lung cancer model, adoptive transfer of FAP-specific T cells significantly reduced FAP-positive stromal cells, with a concomitant decrease in tumor growth. Combining these FAP-specific T cells with T cells that targeted the EphA2 antigen on the A549 cancer cells themselves significantly enhanced overall antitumor activity and conferred a survival advantage compared to either alone. Our study underscores the value of co-targeting both CAFs and cancer cells to increase the benefits of T-cell immunotherapy for solid tumors.

Toward immunotherapy with redirected T cells in a large animal model: ex vivo activation, expansion, and genetic modification of canine T cells.

Adoptive transfer of T cells expressing chimeric antigen receptors (CARs) has shown promising antitumor activity in early phase clinical studies, especially for hematological malignancies. However, most preclinical models do not reliably mimic human disease. We reasoned that developing an adoptive T-cell therapy approach for spontaneous osteosarcoma (OS) occurring in dogs would more closely reproduce the condition in human cancer. To generate CAR-expressing canine T cells, we developed expansion and transduction protocols that allow for the generation of sufficient numbers of CAR-expressing canine T cells for future clinical studies in dogs within 2 weeks of ex vivo culture. To evaluate the functionality of CAR-expressing canine T cells, we targeted HER2+ OS. We demonstrate that canine OS is positive for HER2, and that canine T cells expressing a HER2-specific CAR with human-derived transmembrane and CD28.&zgr; signaling domains recognize and kill HER2+ canine OS cell lines in an antigen-dependent manner. To reduce the potential immunogenicity of the CAR, we evaluated a CAR with canine-derived transmembrane and signaling domains, and found no functional difference between human and canine CARs. Hence, we have successfully developed a strategy to generate CAR-expressing canine T cells for future preclinical studies in dogs. Testing T-cell therapies in an immunocompetent, outbred animal model may improve our ability to predict their safety and efficacy before conducting studies in humans.

Inducible Activation of MyD88 and CD40 in CAR T Cells Results in Controllable and Potent Antitumor Activity in Preclinical Solid Tumor Models

Adoptive immunotherapy with T cells expressing chimeric antigen receptors (CAR) has had limited success for solid tumors in early-phase clinical studies. We reasoned that introducing into CAR T cells an inducible costimulatory (iCO) molecule consisting of a chemical inducer of dimerization (CID)-binding domain and the MyD88 and CD40 signaling domains would improve and control CAR T-cell activation. In the presence of CID, T cells expressing HER2-CARζ and a MyD88/CD40-based iCO molecule (HER2ζ.iCO T cells) had superior T-cell proliferation, cytokine production, and ability to sequentially kill targets in vitro relative to HER2ζ.iCO T cells without CID and T cells expressing HER2-CAR.CD28ζ. HER2ζ.iCO T cells with CID also significantly improved survival in vivo in two xenograft models. Repeat injections of CID were able to further increase the antitumor activity of HER2ζ.iCO T cells in vivo Thus, expressing MyD88/CD40-based iCO molecules in CAR T cells has the potential to improve the efficacy of CAR T-cell therapy approaches for solid tumors.Significance: Inducible activation of MyD88 and CD40 in CAR T cells with a small-molecule drug not only enhances their effector function, resulting in potent antitumor activity in preclinical solid tumors, but also enables their remote control post infusion. Cancer Discov; 7(11); 1306-19. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 1201.

Adoptive cell therapy for sarcoma

Current therapy for sarcomas, though effective in treating local disease, is often ineffective for patients with recurrent or metastatic disease. To improve outcomes, novel approaches are needed and cell therapy has the potential to meet this need since it does not rely on the cytotoxic mechanisms of conventional therapies. The recent successes of T-cell therapies for hematological malignancies have led to renewed interest in exploring cell therapies for solid tumors such as sarcomas. In this review, we will discuss current cell therapies for sarcoma with special emphasis on genetic approaches to improve the effector function of adoptively transferred cells.

MyD88/CD40-based inducible co-stimulation to improve CAR T cell therapy

Meeting abstracts Adoptive immunotherapy with genetically modified T cells holds promise in improving outcomes for cancer patients. While a broad array of genetic modification strategies are being explored, few allow for the specific manipulation of adoptively transferred T cells in vivo . One

Abstract PR16: Genetically engineered T cells redirected against cancer-associated fibroblasts to potentiate immunotherapeutic effects.

Adoptive T-cell therapy has had considerable success in effectuating anti-tumor responses, however complete eradication of bulky solid tumors is rarely observed. This limited efficacy can in part be attributed to the tumor associated stroma; an immunosuppressive microenvironment that greatly decreases the efficacy of the T-cell product. Specifically, cancer associated fibroblasts (CAFs); the central component of the tumor stroma, secrete inhibitory factors and nutrient depleting enzymes that are detrimental to effector T cell function. In addition, CAFs secrete components of the extra-cellular matrix that contributes to desmoplasia and tumor growth. Recent reports also suggest a more pivotal role of the CAFs in regulating the self-renewing cancer stem cell (CSC) niche. CAFs express fibroblast activation protein alpha (FAP); a membrane bound serine protease, in a number of a solid tumors making it an attractive immunotherapeutic target. We hypothesized that targeting CAFs with FAP-specific T cells will destroy the ‘tumor promoting haven’, resulting in significant antitumor effects. To test this hypothesis, we successfully generated FAP-specific T cells using a second-generation chimeric antigen receptor (CAR) specific for FAP. A prototypical CAR combines the antigen specificity of an antibody with the signaling function of a T cell. The resulting genetically engineered FAP-specific T cells recognized and killed human as well as murine FAP-positive target cells ex vivo. To analyze the effects of targeting the tumor associated stroma in vivo we utilized 1) a locoregional model using the FAP negative lymphoblastoid cell line (LCL) tumor and 2) a systemic model using the FAP negative A549 lung adenocarcinoma. In the local model, mice receiving a subcutaneous admixture of FAP-specific T cells and LCL showed a significant decrease in tumor engraftment and growth. Both untreated and control T cell treated mice showed progressive tumor growth with a concomitant induction of murine FAP rich supportive stroma. In our systemic model, the FAP negative non-small cell lung carcinoma (NSCLC) A549 cell line induced the expression of a murine FAP positive reactive stroma in the lungs when injected intravenously through the tail vein. Treatment with FAP-specific T cells showed a significant decrease in murine FAP expression in the lungs with a concomitant decrease in tumor growth and improved survival. Thus targeting FAP in the tumor stroma alone, resulted in a significant anti-tumor effect both in our local and systemic models. Given the reciprocal relationship between tumor cells and CAFs, we hypothesized that co-targeting the tumor cells and CAFs would result in enhanced anti-tumor response than targeting either alone. Erythropoietin-producing hepatocellular carcinoma-A2 (EphA2) CAR positive T cells were used to target the A549 tumor cells. EphA2-specific T cells when administered together with FAP-T cells, resulted in a significant decrease in tumor growth and increased survival compared to mice that received either EphA2 or FAP-T cells alone. Our research therefore underscores the importance of targeting the tumor associated stroma in addition to tumor targeting for a more efficacious therapeutic response. This abstract is also presented as Poster A91. Citation Format: Sunitha Kakarla, Kevin Chow, Melinda Mata, Xiao-Tong Song, Meng-Fen Wu, Hao Liu, Lisa Wang, David Rowley, Klaus Pfizenmaier, Stephen Gottschalk. Genetically engineered T cells redirected against cancer-associated fibroblasts to potentiate immunotherapeutic effects. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr PR16.