Project Grant F31CA317816

PRECLINICAL DEVELOPMENT OF AN ANTI- PTPRZ1 CAR T CELL FOR PRECISION IMMUNOTHERAPY IN GLIOBLASTOMA - GLIOBLASTOMA (GB) REMAINS UNIVERSALLY FATAL DESPITE AGGRESSIVE TREATMENT, WITH A MEDIAN SURVIVAL OF 15-20 MONTHS.1 CART CELL THERAPY HAS TRANSFORMED THE TREATMENT OF HEMATOLOGIC MALIGNANCIES BUT HAS FAILED IN GB PRIMARILY DUE TO ANTIGEN HETEROGENEITY. CURRENT CAR T CELL TARGETS USED IN GB, SUCH AS EGFRVLLL, ARE EXPRESSED ON ONLY 20-30% OF TUMOR CELLS, ENABLING IMMUNE ESCAPE. WE HAVE IDENTIFIED PROTEIN TYROSINE PHOSPHATASE RECEPTOR 21 (PTPRZ1) AS A PROMISING CART TARGET THROUGH SINGLERCELL RNA SEQUENCING, DEMONSTRATING HOMOGENEOUS EXPRESSION ACROSS GB CELLS WITH MINIMAL NORMAL BRAIN EXPRESSION. CRITICALLY, WE HAVE ADDRESSED A CENTRAL LIMITATION OF PRIOR PTPRZ1-TARGETING EFFORTS BY GENERATING BOTH SINGLE-CHAIN VARIABLE FRAGMENTS (SCFVS) AND NANOBODIES THAT SELECTIVELY AND ROBUSTLY RECOGNIZE ENDOGENOUS LEVELS OF PTPRZ1 ACROSS PATIENT-DERIVED GB SAMPLES, GLIOMA STEM CELLS, AND ESTABLISHED GB CELL LINES. THIS F31 PROPOSAL WILL OPTIMIZE AND VALIDATE PTPRZ1-TARGETED CART THERAPY THROUGH TWO SPECIFIC AIMS. AIM 1 WILL CHARACTERIZE BINDING THROUGH IHC AND SPR AND EVALUATE OFF-TARGET TOXICITY USING EPILEPSY PATIENTDERIVED BRAIN SLICE CULTURE TISSUE AND SYNGENEIC MOUSE MODELS. AIM 2 WILL SYSTEMATICALLY OPTIMIZE CAR ARCHITECTURE BY SCREENING SCFV AND NANOBODY BINDING DOMAINS PAIRED WITH NINE HINGE VARIANTS, FIVE TRANSMEMBRANE DOMAINS, AND NINE COSTIMULATORY CONFIGURATIONS, THEN EVALUATE TOP CANDIDATES IN VIVO USING PATIENT-DERIVED XENOGRAFL GB MODELS. SUCCESSFUL COMPLETION OF THIS RESEARCH WILL LEAD TO IDENTIFICATION OF AN OPTIMIZED PTPRZ1-TARGETED CAR CONSTRUCT WITH SUPERIOR TUMOR CONTROL, SUSTAINED T CELL FUNCTION, AND A RIGOROUSLY DEFINED SAFETY PROFILE APPROPRIATE FOR TRANSLATIONAL ADVANCEMENT. THIS WORK WILL HELP ESTABLISH A CLINICALLY TRANSLATABLE IMMUNOTHERAPY FOR GB. FURTHERMORE, THIS F31 FELLOWSHIP WILL PROVIDE ESSENTIAL TRAINING IN CAR T ENGINEERING, TUMOR IMMUNOLOGY, AND TRANSLATIONAL RESEARCH, POSITIONING ME FOR AN INDEPENDENT CAREER DEVELOPING NOVEL CANCER IMMUNOTHERAPIES. THE LONG-TERM GOAL OF THIS PROJECT IS TO EXPLOIT THE UNIFORM OVEREXPRESSION OF PTPRZ1 IN GB TO OVERCOME ANTIGEN HETEROGENEITY AND ADVANCE A RATIONALLY DESIGNED CAR T CELL THERAPY TOWARD CLINICAL TRANSLATION FOR PATIENTS WITH GB.