Cyclophosphamide promotes engraftment of gene-modified cells in a mouse model of Fanconi anemia without causing cytogenetic abnormalities.
A major hurdle for hematopoietic stem cell (HSC) gene therapy for inherited bone marrow disorders, including Fanconi anemia (FA), is adequate engraftment of gene-modified cells. A phenotypic defect in DNA repair renders FA patients sensitive to alkylating agents such as cyclophosphamide (Cy); however, at lower doses, Cy is well tolerated in the FA transplant setting. We tested whether non-alkylating agents could replace Cy for pretransplant conditioning to enhance engraftment of FANCA gene-modified hematopoietic cells. We compared Cy preconditioning with fludarabine (Flu) or cytarabine (AraC) or no conditioning as a control in fanca ( -/- ) mutant mice receiving gene-modified bone marrow cells. Only mice conditioned with Cy exhibited appreciable engraftment of gene-modified cells by PCR and resistance to mitomycin C (MMC). Cy administration following transplantation increased gene marking levels in all animals treated, but highest gene marking and corresponding MMC resistance were observed in mice receiving Cy pre- and posttransplantation. Importantly, no cytogenetic abnormalities were observed in Cy-treated mice. We conclude that Cy is an effective and superior preparative regimen with respect to engraftment of lentivirus-transduced cells and functional correction in fanca ( -/- ) mice. Thus, appropriately dosed Cy may provide a suitable conditioning regimen for FA patients undergoing HSC gene therapy.
Division of Clinical Research, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue North, Seattle, WA 98109-1024, USA.
Jennifer E
JE
Adair
Xin
X
Zhao
Sylvia
S
Chien
Min
M
Fang
Martin E
ME
Wohlfahrt
Grant D
GD
Trobridge
Jason A
JA
Taylor
Brian C
BC
Beard
Hans-Peter
HP
Kiem
Pamela S
PS
Becker
D000818:Animals; D001854:Bone Marrow Cells; D003520:Cyclophosphamide / Q000494:pharmacology*; D003582:Cytogenetics; D004195:Disease Models, Animal; D005199:Fanconi Anemia / Q000235:genetics* / Q000628:therapy; D005434:Flow Cytometry / Q000379:methods; D015316:Genetic Therapy / Q000379:methods; D057809:HEK293 Cells; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008747:Methylcellulose / Q000737:chemistry; D051379:Mice; D008822:Mice, Transgenic; D016685:Mitomycin / Q000494:pharmacology; D019653:Myeloablative Agonists / Q000494:pharmacology; D016133:Polymerase Chain Reaction / Q000379:methods; D019172:Transplantation Conditioning / Q000379:methods*
D019653:Myeloablative Agonists; D016685:Mitomycin; D003520:Cyclophosphamide; D008747:Methylcellulose