? ? The unifying theme of the Weill Cornell PEGT is the challenge of transferring the technology of ex. vivo and in vivo gene transfer to treat human disease. In yr 01-05 of the Weill Cornell PEGT, we focused on developing gene transfer technology, understanding vector biology, and fine tuning the controls that will permit safe and effective gene transfer to humans. We believe the time is ripe to translate our results to the clinic. Thus, for the PEGT renewal, 3 of the 4 projects include clinical studies, while the 4th project continues at the pre-clinical level to be translated to treating human disease in the future. To accomplish these goals, this proposal maintains the same leadership as in our current PEGT projects, including 3 PIs at Weill Cornell (S. Raffi, S. Worgall, R. Crystal) and 1 at Memorial-Sloan Kettering (M. Sadelain). At the center of our gene therapy program is the Belfer Gene Therapy Core Facility, a 12,000 ft2, state-of-the-art wet laboratory, experimental animal and GMP production facility. Together, the PIs, Core Directors and extensive gene transfer infrastructure, represent an integrated program to translate gene therapy to the clinic. The 4 proposed projects (* = clinical) are: Project 1* - Genetic treatment of ?-thalassemia by lentivirus- mediated transfer of a regulated human ?-globin gene (M. Sadelain); Project 2 - Reconstitution of hemangiogenesis by targeted gene delivery of stem cell active chemokines (S. Rafii); Project 3*- Immunization against Pseudomonas aeruginosa by capsid modified Ad vectors expressing Pseudomonas OprF (S. Worgall); and Project 4* - A novel strategy for gene therapy for ?1AT deficiency: Intrapleural delivery of AAV5 vectors expressing al AT (R. Crystal). The Local Cores supporting the projects include: Vector; Analysis; Experimental Animal; Clinical Operations and Regulatory Review; Training and Education; and Administration. The Weill Cornell PEGT also includes the National Clinical Grade Vector Production Core, making plasmid, adenovirus, adeno-associated virus and lentivirus vectors available to NHLBI investigators. (End of Abstract) ? ? INDIVIDUAL PROJECTS AND CORE UNITS: ? ? Project 1: Genetic Treatment of ?-thalassemia by Lentivirus-mediated Transfer of a Regulate ? Human ?-globin Gene. (Sadelain, M.) ? ? DESCRIPTION (provided by applicant): ? ? ?-Thalassemia is one of the commonest inherited diseases in humans, characterized by a severe hemolytic; anemia and ineffective erythropoiesis. While transfusion and chelation therapy do not represent a radical treatment, the use of bone marrow replacement is limited by complications of allogeneic transplantation and the need for aggressive conditioning regimens. The ultimate goal of this proposal is to establish a treatment for 6-thalassemia based on the genetic modification of autologous hematopoietic stem cells. The approach we propose is based on lentiviral-mediated transfer of the normal ?-globin gene. This proposal rests in large part on our earlier demonstration that lentiviral vectors enable stable and efficient transmission of the human ?-globin gene and large fragments of the human ?-globin locus control region (May et al, Nature, 2000), resulting in elevated, therapeutically relevant expression of the human ?-globin gene in long-term murine bone marrow chimeras. We have now extended these findings and demonstrated sustained amelioration of anemia, correction of extramedullary hematopoiesis and prevention of iron accumulation in long-term chimeras with ?-thalassemia intermedia, as well as rescue from lethal ??-thalassemia in a novel adult disease model we created. Combined with new data on long-term expression of the vector encoded human ?-globin gene, busulfan conditioning, and hematopoietic cell transduction, we now propose to investigate specific issues pertaining to the efficacy and safety of globin gene transfer before undertaking a clinical trial to assess the safety and the therapeutic potential of this strategy.
The first aim i s to optimize specific parameters that will impact on the efficacy of Antiviral mediated globin gene therapy, focusing on transcriptional regulation and vector titer.
The second aim addresses safety aspects of lentiviral-mediated globin gene therapy, focusing on oncogene trans-activation, the conditioning regimen, and vector immunogenicity.
This aim i ncludes studies on globin gene transfer in nonmyeloablated rhesus macaques.
The third aim i s to test the safety of lentiviral-mediated globin gene transfer in patients with B-thalassemia and define a minimally toxic, non-myeloablative conditioning regimen that will permit transduced hematopoietic progenitors to durably engraft. To this end, we will first develop and validate a CD34+ cell transduction protocol using serum-free conditions in a closed system and next undertake a pilot study to investigate the safety and efficacy of lentiviral mediated globin gene transfer in busulfan-conditioned patients with 6-thalassemia major. (End of Abstract) ? ?
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