This proposal sets the stage for clinical trials of gene therapy for hemophilia A, while simultaneously initiating studies on a novel vector system. We initially will determine the most effective and safest gene correction strategy for factor VIII (FVIII) deficiency in our mouse model. We have previously shown that functional VIII can be delivered from the suprabasal epidermis to the circulation in a transgenic mouse and a skin explant. We will improve FVII delivery by use of 1) a FVII resistant to inactivation and 2) a species-specific FVIII. We then use the optimal FVIII construct to deliver the gene in a retroviral vector to human keratinocytes. These cells which contain a high fraction of stem cells will be grafted to FVIII-deficient immunodeficient mice to show correction of FVIII-deficiency from transfected keratinocytes. In addition, we follow- up observations suggesting that intradermal injection of an FVIII transgene can result in long-term circulating FVIII. In a second section of the proposal, we seek to improve upon modest long-term correction achieved by mouse FVIII cDNA delivered in an adenoviral vector by tail vein injection. We test a number of variables, including improved gene construct and short-term immunosuppression. We also use a """"""""gutted"""""""" adenoviral vector as a delivery vehicle to improve results. These studies should provide sufficient information concerning optimal modes of therapy to justify attempts to correct a dog model of hemophilia A in the later years of the grant. In a fourth Specific Aim, we initiate studies on a novel vector, the endogenous L1 retro-transposon and determine the efficacy of this promising vector in the delivery of the factor IX gene in cultured cells.
| French, Robert A; Samelson-Jones, Benjamin J; Niemeyer, Glenn P et al. (2018) Complete correction of hemophilia B phenotype by FIX-Padua skeletal muscle gene therapy in an inhibitor-prone dog model. Blood Adv 2:505-508 |
| George, Lindsey A; Sullivan, Spencer K; Giermasz, Adam et al. (2017) Hemophilia B Gene Therapy with a High-Specific-Activity Factor IX Variant. N Engl J Med 377:2215-2227 |
| Gollomp, Kandace; Lambert, Michele P; Poncz, Mortimer (2017) Current status of blood 'pharming': megakaryoctye transfusions as a source of platelets. Curr Opin Hematol 24:565-571 |
| High, Katherine A; Anguela, Xavier M (2016) Adeno-associated viral vectors for the treatment of hemophilia. Hum Mol Genet 25:R36-41 |
| Siner, Joshua I; Samelson-Jones, Benjamin J; Crudele, Julie M et al. (2016) Circumventing furin enhances factor VIII biological activity and ameliorates bleeding phenotypes in hemophilia models. JCI Insight 1:e89371 |
| Zhang, Nanyan; Zhi, Huiying; Curtis, Brian R et al. (2016) CRISPR/Cas9-mediated conversion of human platelet alloantigen allotypes. Blood 127:675-80 |
| Sim, Xiuli; Poncz, Mortimer; Gadue, Paul et al. (2016) Understanding platelet generation from megakaryocytes: implications for in vitro-derived platelets. Blood 127:1227-33 |
| Marcos-Contreras, Oscar A; Smith, Shannon M; Bellinger, Dwight A et al. (2016) Sustained correction of FVII deficiency in dogs using AAV-mediated expression of zymogen FVII. Blood 127:565-71 |
| Arruda, V R; Samelson-Jones, B J (2016) Gene therapy for immune tolerance induction in hemophilia with inhibitors. J Thromb Haemost 14:1121-34 |
| Lyde, R; Sabatino, D; Sullivan, S K et al. (2015) Platelet-delivered therapeutics. J Thromb Haemost 13 Suppl 1:S143-50 |
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