Project Grant F32AI202861

STABLE CELL STATE ENGINEERING FOR REGULATORY T CELL THERAPIES - ABSTRACT AUTOIMMUNE DISEASES SUCH AS MULTIPLE SCLEROSIS, TYPE 1 DIABETES, INFLAMMATORY BOWEL DISEASE, AND RHEUMATOID ARTHRITIS AFFECT TENS OF MILLIONS OF PEOPLE AND ARE LARGELY MANAGED WITH SYSTEMIC IMMUNOSUPPRESSION THAT COMPROMISES HOST DEFENSE AND RARELY INDUCES CURE. ANTIGEN-SPECIFIC TOLERANCE-RETRAINING THE IMMUNE SYSTEM TO IGNORE SELF WHILE PRESERVING PROTECTIVE IMMUNITY-IS A MAJOR UNMET NEED DIRECTLY ALIGNED WITH THE NIH MISSION TO REDUCE THE BURDEN OF CHRONIC, IMMUNE-MEDIATED DISEASE. THIS FELLOWSHIP AIMS TO ENGINEER STABLE CELL THERAPIES FOR RESTORING ANTIGEN-SPECIFIC TOLERANCE. IT WILL FOCUS PRIMARILY ON IL-10-EXPRESSING TYPE 1 REGULATORY T (TR1) CELLS BECAUSE THEY EMERGE NATURALLY IN INFLAMED PERIPHERAL TISSUES TO RESTRAINS PATHOGENIC IMMUNE RESPONSES. THEREFORE, TO ACHIEVE THE OVERARCHING GOAL OF ENGINEERING A STABLE TR1 CELL STATE, THIS PROPOSAL WILL FIRST IDENTIFY GENETIC PERTURBATIONS THAT DRIVE TR1 DIFFERENTIATION (AIM 1) AND THEN ENGINEER SOLUTIONS THAT STABILIZE IT UNDER INFLAMMATORY PRESSURE (AIM 2). IN AIM 1, THE APPLICANT WILL SCREEN THOUSANDS OF MULTI-GENE KNOCK-INS USING STATE-OF-THE-ART EXPERIMENTAL APPROACHES AND THEN REFINE CONSTRUCT DESIGNS USING ADVANCED ALGORITHMS FOR PERTURBATION MODELLING. BY ITERATIVELY SCREENING CONSTRUCTS AND REFINING DESIGNS, AIM 1 WILL CONVERGE ON SOLUTIONS FOR TR1 STATE ENGINEERING WHILE COMPARING THE EFFECTIVENESS OF TWO DIFFERENT STRATEGIES: "RATIONALLY DESIGNED" CONSTRUCTS (ENCODING PUTATIVE TR1-ASSOCIATED REGULATORS) VERSUS UNBIASED "DIVERSITY- ORIENTED" CONSTRUCTS (SYNTHETIC TRANSCRIPTION FACTORS WITH MIXED EFFECTOR AND DNA BINDING DOMAINS). IN AIM 2, THE APPLICANT WILL ENGINEER AND SCREEN CYTOKINE SWITCH RECEPTORS THAT CONVERT COMMON INFLAMMATORY CUES INTO TOLEROGENIC SIGNALS. AIM 2 WILL THEN TEST THE ABILITY OF THESE SYNTHETIC RECEPTORS TO PRESERVE TR1 IDENTITY AND SUPPRESSIVE FUNCTION UNDER TH1-, TH17-, AND TH2-LIKE CYTOKINE MILIEUS IN VITRO AND IN HUMANIZED MOUSE MODELS OF COLITIS. TOGETHER, THESE STUDIES WILL IDENTIFY A TRANSLATION-READY CELL THERAPY CANDIDATE FOR AUTOIMMUNITY WHILE ESTABLISHING DESIGN PRINCIPLES AND A BROADLY APPLICABLE FRAMEWORK FOR "STABLE CELL STATE ENGINEERING" IN ANY CONTEXT. THE INTEGRATED RESEARCH AND TRAINING PLAN WILL PROVIDE RIGOROUS MENTORING IN SYNTHETIC BIOLOGY, DYNAMICAL MODELING, CLINICAL AND TRANSLATIONAL IMMUNOLOGY, AND STRUCTURED CAREER DEVELOPMENT IN GRANTSMANSHIP, COMMUNICATION, AND LEADERSHIP. SUCCESSFUL COMPLETION OF THE RESEARCH AIMS AND TRAINING GOALS WILL DIRECTLY SUPPORT THE APPLICANT'S LONG-TERM CAREER GOAL OF LEADING AN INDEPENDENT, NIH-FUNDED RESEARCH LABORATORY FOCUSED ON CELL STATE ENGINEERING AND TRANSLATABLE THERAPIES FOR INDUCING ANTIGEN-SPECIFIC TOLERANCE.