Transposon-Mediated Gene Therapy for Fanconi Anemia

DESCRIPTION (provided by applicant): Fanconi anemia (FA) is a rare, autosomal recessive inherited disease caused by the absence of any of eight different proteins that regulate DNA repair. Affected individuals suffer from bone marrow failure early in life. FA can be effectively treated by allogeneic hematopoietic stem cell transplant. However, most patients lack a matched related donor, and there is a marked increase in morbidity and mortality following transplant using stem cells from an unrelated source. Discovery Genomics, Inc. (DGI) is developing its Sleeping Beauty (SB) transposon system as a non- viral vector for gene therapy of FA. The two-component SB system consists of a transposon (inverted repeats flanking a therapeutic gene of interest) and a transposase that catalyzes excision of the transposon at the ends of the IR's and then integration into host cell chromosomal sequence. In Phase I studies, DGI demonstrated the successful loading of transposon DNA and RNA into cultured hematopoietic cells, and then correction of human lymphoblastoid cells deficient in Fanconi anemia complementation group C (FANC-C) by SB-mediated transposition. In this Phase II application, the overall goal is to provide key preclinical data that will be necessary to support the anticipated efficacy of SB-mediated gene therapy for FA. In this regard, there are two key questions that will need to be addressed; (i) What is the effectiveness of SB-mediated FANC gene insertion in the treatment of an animal model of FA? (ii) For the human trial, what is the clinically applicable methodology that will be used for introduction of FANC-encoding transposon DNA into human hematopoietic stem cells (HSC)? There are two Specific Aims: Aim 1. Evaluate Sleeping Beauty transposon-mediated integration and long-term expression of the FANC-C gene in hematopoietic stem cells of FANC-C deficient mice as a model for SB-mediated gene therapy for Fanconi anemia. Aim 2. Evaluate Sleeping Beauty transposon-mediated integration and long-term expression in human CD34+ hematopoietic stem cells. For both of these Aims, SB transposon DNA and transposase-encoding sequence will be introduced into HSC using Cyto Pulse electroporation technology. Results from these experiments will provide the technical basis for achieving transposon-mediated integration and long-term expression in hematopoietic stem cells, the cell population that will be targeted for FA gene therapy, and will also provide the commercial basis for growth of DGI in the development of FA gene therapy using the SB transposon system. This project describes the development of a non-viral gene transfer method for treatment of Fanconi anemia, a rare disorder of the blood, by Discovery Genomics, Inc. (DGI), a small biotech startup company. Successful treatment of Fanconi anemia using DGI's transposon system is intended to provide proof of principle for the potential application of this technology to the treatment of other disorders of the blood as well.