Insights into the clonal inventory of gene corrected human SCID-XI hematopoiesis

The semi-random nature of retroviral integration introduces a stable molecular marker into the cellular genome in the form of a genomic-proviral fusion sequence which can be passed on unmodified to all the progenies of a transduced cell. Retroviral integration site analysis provides a powerful method to study the clonal composition of in vivo hematopoiesis after hematopoietic stem cell transplantation. <br>In the present study, a highly sensitive and specific LAM-PCR method was employed to characterize the in vivo hematopoiesis on DNA material from patient No. 4 of the French SCID-X1 gene therapy clinical trial, who has developed a T-cell leukemia like disease at 31 months after gene therapy. Multiple integration sites could be identified in posttransplantation samples, demonstrating multiple transduced clones contributing to hematopoiesis after transplantation. LAM-PCR analysis performed on 37 months posttransplantation samples, however, displayed an oligoclonal repopulation profile, suggesting that only a few predominant transduced clones were contributing to post-chemotherapy hematopoiesis. Two clones identified from 37 months post-chemotherapy samples could be traced back in 6 months and 24 months post-transplantation samples, which provided the first <br>molecular evidence of prolonged activity of individual transduced clones even after clinical interventions such as chemotherapy. Altogether 148 RISs (97 appable RISs) could be identified in this patient. To address the question whether human gene coding regions are favored targets for retroviral integration, all the identified RISs were analyzed with relationship to human RefSeq genes. 64.7% (22/34) of the pre-transplantation RISs as well as 38.1% of the post-transplantation RISs had landed within RefSeq genes. This finding is significant different from a set of 10,000 computer simulated random integration, whereby only 22.4% of integrations has landed within RefSeq genes, indicating a strong tendency of retroviral integration towards gene coding regions of the human genome. Further development of safer designed vectors to be used in human <br>hematopoietic stem cell gene therapy trials is therefore justified.

Author dc:creator

• Hu, Jingqing

Contributors dc:contributor

• Kalle, Christof von

• SCID-X1
• Integrationsstellenanalyse
• integration site analysis

Repository record source_url

https://freidok.uni-freiburg.de/data/1534

OAI identifier oai:identifier

oai:freidok.uni-freiburg.de:1534