Project Grant R21TR005162

DURABILITY OF A GENE EDITING THERAPY THAT RESTORES DYSTROPHIN IN A HUMANIZED MOUSE MODEL OF DUCHENNE MUSCULAR DYSTROPHY - PROJECT SUMMARY GENE EDITING OFFERS SIGNIFICANT ADVANTAGES FOR TREATING RARE DISEASES SINCE IT EDITS THE PATIENT'S OWN DNA. THUS, THE NORMAL REGULATORY ENVIRONMENT OF THE GENE IS MAINTAINED, IT CAN BE USED TO CORRECT ANY GENE REGARDLESS OF GENE SIZE, AND IT GENERATES EDITS THAT ARE EXPECTED TO BE MORE DURABLE THAN GENE REPLACEMENT APPROACHES WHERE LOSS OF THE EPISOMAL TRANSGENE MAY OCCUR. AS NEXT GENERATION IN VIVO GENE EDITING THERAPIES ADVANCE TOWARDS CLINICAL DEVELOPMENT, PRECLINICAL STUDIES ARE NEEDED TO ASSESS LONG-TERM DURABILITY. THE GENE EDITING THERAPY DESCRIBED IN THIS APPLICATION IS DESIGNED TO REMOVE DMD EXONS 45-55 TO RESTORE THE READING FRAME FOR 50% OF DUCHENNE MUSCULAR DYSTROPHY PATIENTS WHILE RETAINING 87% OF THE PROTEIN CODING SEQUENCE. THIS IS ~3X MORE OF THE CODING SEQUENCE THAN GENE REPLACEMENT THERAPIES WHICH DELIVER A "MICRO-DYSTROPHIN" AND INTRODUCES FEWER NON-NATIVE JUNCTIONS. IT ALSO REPLICATES A PATIENT GENOTYPE ASSOCIATED WITH NO SYMPTOMS OR JUST A VERY MILD DISEASE COURSE. THOROUGH PRECLINICAL EFFICACY OF THIS GENE EDITING THERAPY HAS BEEN DEMONSTRATED IN AN ESTABLISHED HUMANIZED MOUSE MODE OF DUCHENNE AT 2 MONTHS POST-TREATMENT. IN THIS PROPOSAL, EFFICACY WILL BE COMPARED IN LONG-TERM 6-MONTH STUDIES VERSUS SHORT-TERM 2-MONTH STUDIES. MOLECULAR AND FUNCTIONAL EFFICACY OUTCOMES WILL ASSESS DMD EDITING, VECTOR COPY NUMBER PER CELL, RESTORATION OF BOTH THE DMD GENE PRODUCT (DYSTROPHIN) AND THE DYSTROPHIN-ASSOCIATED GLYCOPROTEIN COMPLEX, MUSCLE HISTOLOGY, AND MUSCLE FUNCTION TESTING USING TWO DIFFERENT NON- INVASIVE ASSESSMENTS. MULTIPLE DOSING STRATEGIES WILL BE COMPARED, INCLUDING A TWO-FOLD HIGHER SINGLE DOSE AND A REPEAT DOSE AT DAYS 0 AND 56. THIS WORK WILL CLARIFY THE LONG-TERM DURABILITY OF GENE EDITING EFFICACY AND HOW IT IS INFLUENCED BY DOSING REGIMENS. LEARNINGS FROM THIS PROPOSAL WILL HAVE WIDE APPLICABILITY TO OTHER GENE EDITING THERAPIES WHICH TARGET POST-MITOTIC TISSUES LIKE MUSCLE. THESE DATA WILL BE INCLUDED IN FUTURE IND FILINGS AND WILL HELP ADVANCE DEVELOPMENT OF AN INNOVATIVE GENE EDITING THERAPY FOR DUCHENNE MUSCULAR DYSTROPHY.