title: In vivo repopulation ability of genetically corrected bone marrow cells from Fanconi anemia patients.
abstract: Fanconi anemia (FA) is a rare inherited genomic instability syndrome representing one of the best examples of hematopoietic stem cell deficiency. Although FA might be an excellent candidate for bone marrow (BM) genetic correction ex vivo, knockout animal models are not sufficient to guide preclinical steps, and gene therapy attempts have proven disappointing so far. Contributing to these poor results is a characteristic and dramatic early BM-cells die-off when placed in culture. We show here that human primary FA BM cell survival can be ameliorated by using specific culture conditions that limit oxidative stress. When coupled with retrovirus-mediated transfer of the main complementation group FANCA-cDNA, we could achieve long-term reconstitution of the stem cell compartment both in vitro and in vivo. Gene-corrected BM cultures grew for >120 days, and after cultured cell transplantation into NOD/SCID mice, clonogenic human cells carrying the FANCA transgene could be detected 6 months after transduction. By comparison, untransduced cells died in culture by 15 days. Of necessity for ethical reasons, experiments were conducted on a very limited number of primary BM cells. By using low cytokine regimen and conditions matching regulatory requirements, a contingent of gene-corrected cells slowly emerges with an unmet potential for in vivo engraftment. Future therapeutic applications of stem cells might be expanding from these data. In addition, we provide a model of gene-corrected human primary cell growth that carries the potential to better delineate the combined role of both DNA damage and oxidative stress in the pathogenesis of FA.
authors.affiliation: Laboratory of Biotechnology and Applied Pharmacogenetics, Ecole Normale Supérieure de Cachan, 94235 Cachan Cedex, France. odile.haguenauer@lbpa.ens-cachan.fr
authors.forename: Odile
authors.initials: O
authors.lastname: Cohen-Haguenauer
authors.forename: Bruno
authors.initials: B
authors.lastname: Péault
authors.forename: Cécile
authors.initials: C
authors.lastname: Bauche
authors.forename: Marie-Thérèse
authors.initials: MT
authors.lastname: Daniel
authors.forename: Ibrahim
authors.initials: I
authors.lastname: Casal
authors.forename: Vincent
authors.initials: V
authors.lastname: Levy
authors.forename: Jean
authors.initials: J
authors.lastname: Dausset
authors.forename: Michel
authors.initials: M
authors.lastname: Boiron
authors.forename: Christian
authors.initials: C
authors.lastname: Auclair
authors.forename: Eliane
authors.initials: E
authors.lastname: Gluckman
authors.forename: Michel
authors.initials: M
authors.lastname: Marty
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001854:Bone Marrow Cells / Q000166:cytology / Q000502:physiology*; D016026:Bone Marrow Transplantation; D002648:Child; D018076:DNA, Complementary / Q000235:genetics; D005199:Fanconi Anemia / Q000235:genetics / Q000628:therapy*; D052217:Fanconi Anemia Complementation Group A Protein / Q000235:genetics*; D005260:Female; D015316:Genetic Therapy; D006801:Humans; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D018384:Oxidative Stress / Q000187:drug effects; D014162:Transfection
chemical_list: D000975:Antioxidants; D018076:DNA, Complementary; C498471:FANCA protein, human; D052217:Fanconi Anemia Complementation Group A Protein