Abstract
AbstractInsufficient expression of factor VIII (fVIII) is a major hurdle in the development of successful nucleic acid treatments for hemophilia. However, we recently showed that under myeloablative and reduced-intensity total body irradiation (TBI) conditioning, transplantation of hematopoietic stem cells (HSCs) transduced with recombinant retroviruses containing B domain–deleted porcine fVIII (BDDpfVIII) sequences provides curative fVIII levels in a hemophilia A mouse model. In the current study, we tested BDDpfVIII activity after nonmyeloablative conditioning with busulfan, cyclophosphamide, or fludarabine and immunosuppressive agents CTLA4-Ig + anti-CD40L or anti-(murine)thymocyte serum (ATS). ATS is similar in action to anti-(human)thymocyte globulin (ATG), which is used clinically with busulfan in bone marrow transplantations to increase donor cell engraftment. Mice conditioned with busulfan + ATS and that received a transplant of BDDpfVIII-transduced stem-cell antigen 1-positive cells exhibited moderate levels of donor cell chimerism (between 20% and 60%) and achieved sustained fVIII levels more than 1 U/mL. Similar results were observed in mice preimmunized with human fVIII and conditioned with 5 Gy TBI + ATS or busulfan + ATS. These data demonstrate that it is possible to achieve sufficient fVIII expression after transplantation of BDDpfVIII-transduced HSCs following low-toxicity pretransplantation conditioning with targeted immunosuppression, potentially even in the context of preexisting inhibitors.
Highlights
Hemophilia A is an X-linked recessive genetic disorder leading to a deficiency of functional clotting factor VIII
Consistent with these results, we showed that mesenchymal stem cells transduced with recombinant retroviruses containing BDDpfVIII sequences express factor VIII (fVIII) more efficiently than those transduced with recombinant retroviruses containing human fVIII (hfVIII) sequences.[4]
Genetic modification and transplantation of hematopoietic stem cells (HSCs), which offer a means to introduce the BDDpfVIII transgene into cells that are long-lived and undergo self-renewal and differentiation,[5] under myeloablative and reduced-intensity pretransplantation conditioning resulted in curative fVIII levels
Summary
Hemophilia A is an X-linked recessive genetic disorder leading to a deficiency of functional clotting factor VIII (fVIII). FVIII expression was sustained for greater than 1 year posttransplantation in a mouse model of hemophilia A, overcoming the barrier of low fVIII expression.[4] Alternatively, Moayeri et al achieved sustained levels of a bioengineered hfVIII under myeloablative conditions by transplanting high doses of genetically modified bone marrow cells.[6] Since prior studies have shown that in vivo transduction can lead to a host immune response and elimination of transduced cells,[7] these successes indicate that transduction of cells ex vivo and transplantation of the genetically modified cells under myelosuppressive conditioning may be necessary. We show that following nonmyeloablative transplantation of BDDpfVIII-modified HSCs into hemophilia A mice it is possible to achieve fVIII levels similar to that observed in healthy mice and humans
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