The overall goal of this Phase II SBIR grant proposal is to research and develop the GMP manufacturing process for next-generation tumor infiltrating lymphocyte (TIL) cancer therapeutic for solid tumors. In clinical trials, TIL-based immunotherapies have demonstrated tumor regression and increased survival rates against different cancer types. Despite these encouraging clinical results, durable antitumor responses are typically observed in a subset of patients with advanced cancers like melanoma. This stresses the need to develop more effective TIL-based strategies and to expand efficacy against different cancer types. TIL therapies are hindered in part by low TIL accumulation into tumors, low persistence, weak T cell receptor (TCR) affinity and/or avidity as well as the low expression of tumor antigens on the cell’s surface. Additionally, the presence of suppressive signals on T cells such as Lag3, Tim3, CD39, pose a major obstacle to generating effective and long-lived antitumor T cell responses. In our published and preliminary studies, activating MyD88 signaling in human or mouse T cells, increases antitumor activity and prolongs T cell survival. By fusing the high affinity CD8α (extracellular) to MyD88 (intracellular; CD8α:MyD88) we have developed a novel and universal platform that activates MyD88 signaling strictly upon the engagement of the TCR with the MHC-antigen on tumor cells. CD8α:MyD88 expression in TILs cells substantially increases responses to weak and suboptimal levels of tumor antigens including neoantigens, and in preliminary studies has demonstrated the extraordinary property of reducing the T cells entry into an ‘exhausted state’. Importantly, the use of the CD8α co-receptor represents a ‘universal’ approach to enhancing T cell responses, and can be used in patients regardless of the patient’s HLA type. Through this SBIR Phase II application, we propose the following aims. Aim 1 will research, develop, and manufacture GMP grade g-retroviral vectors. Here, we will generate high virus titer-producing CD8a:MyD88-EGFRt PG-13 cell lines, accompanying analytical assays, and manufacture the GMP CD8a:MyD88-EGFRt g-retroviral vectors. For this aim, we are partnering with a commercial contract development manufacturing organization (CDMO), Vigene Biosciences to generate GMP viral vectors. In Aim 2, we propose to develop and manufacture GMP CD8a:MYD88 engineered TILs (coR8:AMPTM TILs). In collaboration with the Gates Manufacturing Facility (GBF) at the University of Colorado Anschutz Medical Campus (UCAMC) we will conduct process & analytical assay, QC release assay, develop and manufacture of GMP grade coR8:AMPTM TILs. The successful completion of these aims will lead to the pre-IND and IND filings with the FDA for a Phase I clinical trial in patients with immune refractory solid tumors. This activity will be conducted in collaboration with the UCAMC’s Cell Therapy Operations Program (CTOP), Gates Biomanufacturing Facility, and our te...