Gene therapy of Diamond Blackfan anemia CD34+ cells leads to improved erythroid development and engraftment following transplantation
Received 21 May 2008; received in revised form 16 June 2008; accepted 24 June 2008. published online 21 August 2008.
Objective
Diamond-Blackfan anemia (DBA) is a rare congenital hypoplastic anemia caused by mutations in ribosomal protein (RP) genes. Our aim is to develop gene therapy for DBA patients with mutations in RPS19. We previously demonstrated that RPS19 gene transfer partially corrects erythroid development in vitro. In this study, we asked if RPS19 gene transfer corrects erythroid development in unsorted cells transplanted to immunodeficient mice and if the RPS19-corrected fraction has a proliferative advantage after transplantation. We further determined if high level of RPS19 expression is required for correction.
Material and Methods
Mobilized peripheral blood CD34+ cells were transduced by oncoretroviral vector particles pseudotyped with the feline endogenous retrovirus envelope. Vectors containing two different promoters with different RPS19 transgene expression levels were compared. Transduced cells were transplanted to immunocompromised nonobese diabetic/severe combined immunodeficient–β2 microglobulin null mice in order to assess therapeutic effects of RPS19 gene transfer in vivo.
Results
We show that correction of erythroid development requires high RPS19 expression. The corrected fraction of unselected DBA cells have a survival advantage in vivo, suggesting that successful gene therapy may only require correction of a fraction of the patient cells.
Conclusion
Our findings are fundamental for development of clinical gene therapy for DBA because they demonstrate increased engraftment of RPS19-transduced cells without selection of gene-corrected cells prior to transplantation, an essential prelude to studying long-term therapeutic effects in emerging animal models for DBA.
aDepartment of Molecular Medicine and Gene Therapy, Lund Stem Cell Center, Lund University Hospital, Lund, Sweden
bWhitehead Institute for Biomedical Research, Massachusetts Institute of Technology, Cambridge, Mass., USA
Offprint requests to: Johan Flygare, M.D., Ph.D., Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology, 9 Cambridge Center, Cambridge, MA 02142
ABSTRACT