Assessing The Therapeutic Efficacy Of Disease-Specific T-Cell Biofactories

Current protein-based cancer therapies have several disadvantages, which include general toxicity, lack of response at the administered dose and differing responses from patients to patient. To circumvent these issues, we propose a platform in which engineered T-cells specifically recognize tumors and secrete therapeutic peptides directly at the disease site leading to targeted cancer cell killing. This approach will increase the specificity of the therapy and decrease toxicity to healthy cells. We transformed acute T-cell lymphoma cells into biofactories for site-specific synthesis of therapeutic proteins upon stimulation by antigen-presenting disease cells. The effector T-cell line was engineered by stably introducing a chimeric T-cell receptor to recognize Folate-Receptor alpha (FRα) or Mesothelin (MSLN) protein. Upon binding of the effector T-cells to the receptor, an intracellular cascade directed expression of non-human proteins is induced. Specific T-cell binding to human ovarian cancer cell lines and signaling was measured by in vitro co-culture using luciferase production as a surrogate for therapeutic peptide secretion. We demonstrated that T-cells can be genetically programed to synthesize and secrete proportionate amounts of engineered proteins upon engaging the tumor-associated antigens (FRα or MSLN) on a human ovarian cancer cell line, OVCAR3 (FRα+MSLN+). A FRα-MSLN- ovarian cancer cell line, A2780cis, was used as the non-targeted negative control. The difference in protein secretion following stimulation by the two cell lines, as measured by luciferase activity, was statistically significant within 1 hour. It reached ~35-fold within 1 to 3 days, and we observed stable expression for at least 10 days. The luminescent signal was proportionate to the number of OVCAR3 cells. To further validate the specificity of target engagement, we generated A2780cis-FRα positive and A2780cis-MSLN positive cell lines and demonstrated selective binding and activation of the corresponding effector cells in co-culture assays. No binding was detected to the A2780cis-vector control cells. In vivo results for T-cell biofactories targeting OVCAR3 tumors 24 hours post-stimulation validated the in vitro results. Currently, we are engineering T-cell biofactories to release cytotoxic peptides and are assessing their therapeutic efficacy against cancer cells in vitro using co-culture assays and supernatant transfer. Our results show that T-cells can be genetically reprogrammed to serve as biofactories for the synthesis of therapeutic proteins upon stimulation by antigen-presenting disease cells. Importantly, these studies demonstrate the feasibility of developing the next generation of adoptively transferred T-cell therapies to target tumors that express FRα (e.g., ovarian, breast, lung) and/or MSLN (e.g. ovarian, lung, pancreatic) on their cell surfaces for cancer therapy. Citation Format: Claire E. Repellin, Puja Patel, Lucia Beviglia, Harold Javitz, Lidia C. Sambucetti, Parijat Bhatnagar. Assessing the therapeutic efficacy of disease-specific T-cell biofactories [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2557.