Project Grant F31HL184854

OPTIMIZATION OF ENHANCED IN SITU PROTEIN PRODUCTION FOR FVIII REPLACEMENT USING MODULAR, RATIONALLY DESIGNED TRANSLATABLE RIBONUCLEIC ACIDS EXPRESSING SELF-REPLICATIVE MACHINERY - PROJECT SUMMARY/ABSTRACT THIS FELLOWSHIP WILL SUPPORT THE INTERDISCIPLINARY RESEARCH AND TRAINING OF MR. WYATT BECICKA UNDER THE SPONSORSHIP OF BOSTON UNIVERSITY'S DR. MARK GRINSTAFF. THE PROPOSED RESEARCH ADDRESSES THE CURRENT LIMITATIONS OF FACTOR VIII (FVIII) REPLACEMENT THERAPY FOR HEMOPHILIA A. DUE TO A SHORT HALF-LIFE AND LIMITED THERAPEUTIC WINDOW OF FVIII, THE CURRENT STANDARD OF CARE FOR HEMOPHILIA A PATIENTS ENTAILS MULTIWEEKLY PROPHYLACTIC FVIII INFUSIONS. THIS COST-INTENSIVE DOSING REGIMEN IS REQUIRED TO PROVIDE VITAL YET SUBOPTIMAL COAGULANT ACTIVITY. MEANWHILE, ACUTELY SUPRATHERAPEUTIC FVIII LEVELS POST-INFUSION PROMOTE THE DEVELOPMENT OF NEUTRALIZING ANTIBODIES IN OVER 30% OF SEVERE HEMOPHILIA A PATIENTS, FOR WHICH THE CURRENT TREATMENT IS PROLONGED IMMUNE TOLERANCE INDUCTION WITH LIMITED SUCCESS RATES. THIS ESTABLISHES A NEED FOR AN ACCESSIBLE MODALITY FOR FVIII PROTEIN REPLACEMENT WITH IMPROVED PHARMACOKINETICS. SELF-AMPLIFYING RNA (SARNA) UTILIZES REPLICATIVE MACHINERY TO EXTEND THE DURATION AND MAGNITUDE OF CARGO PROTEIN PRODUCTION IN VIVO; HOWEVER, CURRENT CLINICAL SARNAS HAVE BEEN DEVELOPED WITH A LIMITED SUBSET OF REPLICATIVE PROGRAMS THAT MAXIMIZE RNA REPLICATION AND PROTEIN PRODUCTION WITH MINIMAL CONSIDERATION FOR IMMUNOGENICITY AND SUSTAINABLE REDOSING. I HYPOTHESIZE THAT THE REPLICATION KINETICS AND CELLULAR DISTRIBUTION OF SARNA CAN BE PREDICTABLY CONTROLLED TO ELICIT DISTINCT IN VIVO PROTEIN EXPRESSION PROFILES AND IMMUNE RESPONSES IN A MODULAR FASHION THAT IS INDEPENDENT OF CARGO PROTEIN IDENTITY. IN TURN, I PROPOSE ESTABLISHING A SET OF SARNAS WITH NOVEL IN VIVO EXPRESSION PROFILES IN ORDER TO BOTH INVESTIGATE AND OPTIMIZE THE RELATIONSHIP BETWEEN FVIII DOSE, HOST IMMUNITY, AND LONG-TERM EFFICACY OF FVIII REPLACEMENT THERAPY. THE SPECIFIC AIMS OF THIS FELLOWSHIP PROPOSAL ARE AS FOLLOWS. AIM 1 LEVERAGES A BARCODED SARNA SYSTEM TO ESTABLISH A GOLD STANDARD LIPID NANOPARTICLE (LNP) DELIVERY SYSTEM FOR INTRAMUSCULAR (IM) SARNA PROTEIN REPLACEMENT. AIM 2 INVESTIGATES THE IMPACT OF DIFFERENT SARNA REPLICATION PROGRAMS ON THE SAFETY AND EFFICACY OF PROTEIN REPLACEMENT. AIM 3 ESTABLISHES OPTIMIZED, FUNCTIONAL FVIII REPLACEMENT IN A MOUSE MODEL OF HEMOPHILIA A. THE FELLOW HAS RECRUITED LEADERS IN RNA SYNTHESIS, IN VIVO RNA DELIVERY, CLINICAL PHARMACOLOGY, TRANSLATIONAL IMMUNOLOGY, AND MODELS OF VASCULAR DISEASE TO HIS THESIS COMMITTEE. HIS SPONSOR, DR. MARK GRINSTAFF, WILL CONTINUE TO MEET REGULARLY WITH THE FELLOW AS PART OF HIS THREE-YEAR TRAINING PLAN, WHICH INCLUDES: 1) TRAINING IN RNA AND LNP SYNTHESIS, IN VIVO TRANSGENE DELIVERY, PRECLINICAL STUDIES OF PRIMARY TISSUES AND CELLS, AND WORK WITH ADVANCED MOUSE MODELS OF DISEASE, 2) TRAINING IN RESPONSIBLE CONDUCT OF RESEARCH; 3) ATTENDING SEMINARS, WORKSHOPS, AND NETWORKING EVENTS IN THE BIOMEDICAL ENGINEERING DEPARTMENT; 4) PREPARING FOR AND PRESENTING AT CONFERENCES TO SHARE RESULTS AND BUILD PROFESSIONAL NETWORKS; AND, 5) PROFESSIONAL DEVELOPMENT VIA HIS ADVISORS AND HIS RUBRIC- BASED INDIVIDUAL DEVELOPMENT PLAN TO ATTAIN HIS ULTIMATE GOAL OF BECOMING AN INDEPENDENT RESEARCHER AND PROFESSOR.