Non-myeloablative conditioning with busulfan before hematopoietic stem cell transplantation leads to phenotypic correction of murine Bernard-Soulier syndrome

J Thromb Haemost. 2014 Oct;12(10):1726-32. doi: 10.1111/jth.12673. Epub 2014 Aug 26.


Background: Bernard-Soulier syndrome (BSS) is an inherited bleeding disorder characterized by macrothrombocytopenia. Platelet transfusion is used for the management of bleeding, but repeated transfusion often results in alloimmunization. We have recently shown phenotypic correction of murine BSS (GPIbα(null) ) using lethal radiation conditioning followed by hematopoietic lentivirus-mediated gene transfer.

Objectives: For application of gene therapy to treatment of human patients, it is important to minimize treatment-related side effects. The objective of this study is to model a clinically relevant non-myeloablative hematopoietic stem cell (HSC) transplantation strategy.

Methods: Using transplantation of bone marrow (BM) HSCs from transgenic mice that express hGPIbα (hGPIbα(tg+/+) ), we sought to (i) determine the percentage of hGPIbα(tg+/+) HSCs required for therapeutic benefit, (ii) evaluate the efficacy of non-myeloablative conditioning using busulfan, and (iii) test the ability of anti-thymocyte globulin (ATG) to prevent/reduce undesirable immune responses.

Results: Transplantation of 10-20% hGPIbα(tg+/+) BM HSCs mixed with GPIbα(null) BM HSCs into irradiated GPIbα(null) mice was sufficient to correct bleeding time (n = 5). Transplantation of hGPIbα(tg+/+) BM HSCs into busulfan-conditioned GPIbα(null) mice corrected bleeding time in 21 of 27 recipients. Antibody response to hGPIbα and immune-mediated thrombocytopenia was documented in eight of 27 recipients, suggesting immunogenicity of hGPIbα in busulfan-conditioned GPIbα(null) mice. However, these antibodies disappeared without treatment within 30 weeks after transplantation. A combination of busulfan plus ATG conditioning successfully prevented antibody development and significantly increased therapeutic engraftment.

Conclusion: A conditioning regimen of busulfan in combination with ATG could potentially be used in non-myeloablative autologous gene therapy in human BSS.

Keywords: Bernard-Soulier syndrome; bleeding time; blood platelets; bone marrow transplantation; genetic therapy.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bernard-Soulier Syndrome / genetics
  • Bernard-Soulier Syndrome / therapy*
  • Bleeding Time
  • Busulfan / administration & dosage*
  • Disease Models, Animal
  • Gene Transfer Techniques
  • Genetic Therapy
  • Hematopoietic Stem Cell Transplantation
  • Humans
  • Immune System
  • Immunosuppressive Agents / chemistry
  • Lentivirus
  • Mice
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Phenotype
  • Thrombocytopenia / metabolism
  • Transgenes
  • Transplantation Conditioning*


  • Immunosuppressive Agents
  • Busulfan