SUMMARY CORE B: GMP CELL and RNA MANUFACTURING Core B is the Clinical Cell and Vaccine Production Facility (CVPF). The CVPF is a Foundation for the Accreditation of Cellular Therapy (FACT)-accredited, Good Manufacturing Practices (GMP) compliant cell and RNA manufacturing facility. This facility supports “bench to bedside” translation of innovative investigational cell and gene biologics for a wide range of cancers, infectious diseases, and other therapeutic indications at the University of Pennsylvania (Penn) and its collaborating institutions. The CVPF supports manufacturing for 18 investigational new drug applications (INDs), and has produced over 2400 cellular vaccines that have been safely administered to more than 900 patients. The CVPF is designated as the Core B GMP cell and RNA manufacturing resource for the projects outlined in this proposal, and supports a) process development and validation experiments for successful GMP clinical translation of cell and RNA manufacturing and b) clinical trial manufacturing and final product testing of gene modified cells for human administration. The CVPF will provide specific support for the proposed projects through the development of protocols to manufacture (1) tandem CD19 CAR T cell and CD22 CAR T cell products, (2) TCR negative, HLA-I negative CD19 CAR T cells, (3) CD33 negative hematopoietic stem cells and CD33 CAR T cells, and (4) tandem CD19 CAR T cell and BCMA CAR T cell products. To achieve effective deletion of the above mentioned targets (TCR, HLA-I, CD33) we will manufacture GMP-grade guide RNAs and Cas9 mRNA. We will electroporate therapeutic cells with these RNA constructs to create CRISPR/Cas9 mediate, target negative cells. Quality control assays and release criteria will be developed for each therapeutic cell type to ensure consistent production of safe and high quality modified cells.