β-Thalassemia and Sickle Cell Disease are the most common monogenic diseases worldwide, with >317,000 newborn/year, caused by defects of β-globin expression or structure. They lead to transfusion-dependent anemia, multi-organ damage and early death. Allogeneic Hematopoietic Cell Transplantation (HCT) is the only cure, available for few patients. Ex vivo gene therapy (GT), i.e. autologous transplantation of Hematopoietic Stem and progenitor Cells (HSCs) modified by lentiviral (LV) gene-transfer to express a therapeutic hemoglobin (Hb), is an extremely promising approach for those not eligible to HCT. The most promising GT approaches aim at the production of anti-sickling (AS) Hb, or at the reactivation of Fetal Hb (HbF). Only partial clinical efficacy has been achieved so far, mainly due to suboptimal Hb production and inner LV restraints. Here we propose to develop an original, forward-looking GT strategy for β-globin defects to overcome these limitations, based on 1) gene-cassette optimization for enhanced production of a potent AS β-globin; 2) the usage of novel hyper-functional transposon-vectors to safely integrate multiple copies of complex gene-cassettes into HSPCs; 3) the implementation of a new, cost-effective, scalable and standardized vector-production generating ready-to-use minicircle DNAs, overcoming the major drawbacks of LV production for clinical application, expensive and variable in infectivity. Production of therapeutic AS Hb and the overall GT strategy will be tested in vitro in donor- and patient-derived HSCs, and in vivo in humanized immunodeficient mice in an efficacy/safety preclinical study. This GT for β-globin defects has a solid potential for clinical efficacy and translation, combining clinical evidences with a strategic design to maximize its accessibility to the multitude of neglected children born with these lethal and devastating diseases in the poorest, and most affected, countries of Africa, Middle East and Indian subcontinent.
Linda Bucciarelli, Graduated student
Poletti V., Charrier S., Corre G., Gjata B., Vignaud A., Zhang F., Rothe M., Schambach A., Gaspar H.B., Thrasher A.J., and Mavilio F. Preclinical Development of a Lentiviral Vector for Gene Therapy of X-Linked Severe Combined Immunodeficiency Mol Ther Methods Clin Dev. 2018 Mar 10;9:257-269;
Poletti V., Urbinati F., Charrier S., Corre G., Hollis R.P., Martin S., Rothe M., Schambach A., Kohn D.B and Mavilio F. Pre-clinical development of a lentiviral vector expressing the anti-sickling βAS3 globin for gene therapy for sickle-cell disease Mol Ther Methods Clin Dev. 2018 Nov 1;11:167-179;
Weber L., Poletti V., Magrin E., Antoniani C., Martin S., Bayard C., Sadek H., Felix T., Meneghini V., Antoniou M., El-Nemer W., Mavilio F, Cavazzana M., Andre-Schmutz I. and Miccio A. An optimized lentiviral vector corrects efficiently the human sickle cell disease phenotype Mol Ther Methods Clin Dev. 2018 Aug 4; 10:268-280;
Urbinati F., Campo Fernandez B., Masuik K., Poletti V., Hollis R.P, Koziol C, Kaufman M.L., Brown D., Mavilio F. and Kohn D.B. Gene Therapy for Sickle Cell Disease: a lentiviral vector comparison study Hum Gene Ther. 2018 Sep 10;
Miccio A., Poletti V., Tiboni F., Rossi C., Antonelli A., Mavilio F. and Ferrari G. The GATA1 HS2 enhancer allows persistent and position-independent expression of a beta-globin transgene PLoS ONE 6(12): e27955; Dec 2, 2011;
Poletti V., Delli Carri A., Tagliazucchi G.M, Faedo A., Petiti L., Mazza E.M.C, Peano C., De Bellis G., Bicciato S., Miccio A., Cattaneo E., Mavilio F.. Genome-Wide Definition of Promoter and Enhancer Usage during Neural Induction of Human Embryonic Stem Cells PLoS ONE 10(5): e0126590; May 15, 2015
Holstein M., Mesa-Nuñez C., Miskey C., Almarza E., Poletti V., Schmeer M., Grueso E., Ordóñez Flores J.C., Kobelt D., Walther W., Kumar Aneja M., JGeiger J., B. Bonig H., Izsvák Z., Schleef M., Rudolph C., Mavilio F., Bueren J.A., Guenechea G. and Ivics Z. Efficient Non-Viral Gene Delivery into Human Hematopoietic Stem Cells by Minicircle Sleeping Beauty Transposon Vectors Mol Ther. 2018 Jan 31.
Lidonnici M.R., Paleari Y., Tiboni F., Mandelli G., Rossi C., Vezzoli M., Aprile A., Lederer C., Ambrosi A., Chanut F., Sanvito F., Calabria A., Poletti V., Mavilio F., Montini E., Naldini L., Cristofori P. and Ferrari G. Multiple integrated non-clinical studies predict safety of lentiviral mediated gene therapy for beta thalassemia Mol Ther Methods Clin Dev. 2018 Sep 13;11:9-28;
Charrier S., Lagresle C., Poletti V., Rothe M., Cédrone G., Gjata B., Mavilio F., Fischer A., Schambach A., de Villartay J-P, Cavazzana M., Hacein-Bey Abina S., Galy A. Biosafety Studies of a clinically applicable lentiviral vector for the gene therapy of Artemis-SCID Mol Ther Methods Clin Dev, Dec 2019,15:232-245;
Poletti V. and Biffi A. Gene-Based Approaches to Inherited Neurometabolic Diseases. Hum Gene Ther. 2019 Sep 10;
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