1. ¹Division of Experimental Hematology, Cincinnati Children's Research Foundation, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA
2. ²Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA
3. ³Cincinnati Children's Research Foundation, Center for Epidemiology and Biostatistics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA
4. ⁴Department of Experimental Hematology, Hannover Medical School, Hannover, Germany

Correspondence: David A. Williams, Children's Hospital Boston, 300 Longwood Avenue, Karp 07212, Boston, Massachusetts 02115, USA. E-mail: DAWilliams@childrens.harvard.edu

Received 31 December 2007; Accepted 10 March 2008; Published online 8 April 2008.

Abstract

Fanconi anemia (FA) is a rare recessive syndrome, characterized by congenital anomalies, bone marrow failure, and predisposition to cancer. Two earlier clinical trials utilizing -retroviral vectors for the transduction of autologous FA hematopoietic stem cells (HSCs) required extensive in vitro manipulation and failed to achieve detectable long-term engraftment of transduced HSCs. As a strategy for minimizing ex vivo manipulation, we investigated the use of a "rapid" lentiviral transduction protocol in a murine Fanca^–/– model. Importantly, while this and most murine models of FA fail to completely mimic the human hematopoietic phenotype, we observed a high incidence of HSC transplant engraftment failure and low donor chimerism after conventional transduction (CT) of Fanca^–/– donor cells. In contrast, rapid transduction (RT) of Fanca^–/– HSCs preserved engraftment to the level achieved in wild-type cells, resulting in long-term multilineage engraftment of gene-modified cells. We also demonstrate the correction of the characteristic hypersensitivity of FA cells against the cross-linking agent mitomycin C (MMC), and provide evidence for the advantage of using pharmacoselection as a means of further increasing gene-modified cells after RT. Collectively, these data support the use of rapid lentiviral transduction for gene therapy in FA.