In Vivo Editing for Hemophilia Gene Therapy

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Project Summary The exceptional promise of gene therapy for hemophilia has been shown in recent studies that result in long term correction of factor IX deficiency, via hepatic transduction with an adeno-associated vector (AAV). In spite of this watershed event in the history of the gene therapy field, there presently exist barriers to the fullest implementation of hemophilia gene therapy. The limited packaging capacity of AAV for larger gene constructs practically confounds the use of this vector for factor VIII correction and vector-related toxicities have been noteworthy for the context of in vivo transduction of the liver. We propose to develop a novel vector approach that addresses these limitations. Of note, our recent strategies to retarget adenoviral vectors (Ad) have allowed mitigation of vector-associated liver toxicities and in vivo transduction of the pulmonary endothelium that has demonstrated that this site can serve as a fully effective platform to reconstitute deficient serum proteins. Moreover, new generation Ads with multiple deletions can be modified to incorporate an expanded range of gene constructs and larger payloads relevant to gene therapy for the full spectrum of hemophilia disorders. We propose to utilize the CRISPR/Cas 9 system which is fully commensurate with Ad technology, in conjunction with our pulmonary endothelial-targeted Ad, to achieve stable incorporation of corrective hemophilia genes within the pulmonary endothelium. Our novel strategy thereby offers the potential for stable genetic correction in a manner that circumvents potential vector-associated toxicities for both factor IX and factor VIII hemophilia. Successful completion of the aims within this proof of feasibility Phase 1 STTR will set the stage for the application of corrective gene therapy for hemophilia to the widest context of patients and provide the rationale for translational development of our highly novel approach into lead optimization and selection in Phase II. Our groups have unparalleled track records for successful bench-to-bed translation of novel gene therapy strategies. Our strategy to accomplish stable expression of deficient hemophilia factors deriving from a pulmonary vascular source clearly represents a highly original approach. In addition to circumventing the issue of vector-mediated hepatotoxicities, the expanded packaging capacity of adenovirus vectors also feasibilizes gene therapy for factor VIII, as well as factor IX deficiency disorders. To hasten translation of our approach, we will utilize adenoviral vectors derived from non-human primate adenoviruses and will specifically use gorilla Ad vectors (GAd) that address a pivotal human use barrier by circumventing preformed adenoviral immunity and are show excellent performance characteristics in therapeutic applications. The profile of efficacy studies herein will constitute the rationale for translational development of this novel approach as well as a leap to establishing the pulmonary endothelium as a source to provide secreted factors as an important strategy for the range of inherited genetic disorders based upon deficient serum factors as well as other potential therapeutic applications.