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- Karlsson, Stefan, et al.
(författare)
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Development of gene therapy for blood disorders by gene transfer into haematopoietic stem cells.
- 2002
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Ingår i: Haemophilia. - : Wiley. - 1351-8216. ; 8:3, s. 255-260
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Tidskriftsartikel (refereegranskat)abstract
- Haematopoietic stem cells (HSCs) are important target cells for gene therapy of blood disorders due to their pluripotency and ability to reconstitute haematopoiesis following myeloablation and transplantation. HSCs can 'self-renew' and generate new stem cells. Genetically modified stem cells are therefore expected to last a lifetime in the recipient following blood and marrow transplantation, and can potentially cure haematological disorders. Oncoretroviral vectors have been the main vectors used for HSCs because of their ability to integrate into the chromosomes of their target cells. Because oncoretroviral vectors require dividing target cells for successful localization of the preintegration complex and subsequent chromosomal integration of the provirus, only the dividing fraction of the target cells can be transduced. As only a small fraction of haematopoietic stem cells is dividing at any one time, oncoretroviral vector transduction of human HSCs has been low in clinical trials. However, patients with severe combined immune deficiency-X1 (SCID-X1) have recently been treated successfully by gene therapy of autologous bone marrow cells using oncoretroviral vectors containing the common gamma chain gene. While several additional disorders may potentially be treated successfully using oncoretroviral gene transfer to HSCs, many disorders may require much higher gene transfer efficiency than was achieved in the SCID-X1 study. Therefore, lentiviral vectors have recently emerged as promising vectors for human HSCs because they can transduce dividing and nondividing HSCs efficiently, and may become the vectors of choice in the future for treatment of blood disorders where a large fraction of HSCs has to be corrected.
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| 2. |
- Woods, Niels-Bjarne, et al.
(författare)
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Development of Gene Therapy for Hematopoietic Stem Cells using Lentiviral Vectors
- 2002
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Ingår i: Leukemia. - 1476-5551. ; 16:4, s. 563-569
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Hematopoietic stem cells are an ideal target for genetic manipulation for the purpose of curing hematological disorders as they have the ability to develop into all blood lineages and to self-renew. In this study we demonstrate that lentiviral vectors, based on HIV-1, can efficiently transfer genes into human hematopoietic progenitor and stem cells as assessed following stem cell transplantation in immune compromised mice. High efficiency transduction of repopulating cells was achieved in both primary (48±23%, n=6) and secondary transplant recipients (64±13%). These results demonstrate the ability of lentiviral vectors to efficiently transduce human pluripotent candidate stem cells. Modifications to the vector design were performed to optimize the vector for high-level transgene expression in the progeny of the repopulating cells. Eight-fold higher expression levels were achieved in mice in both the lymphoid and myeloid progeny cells compared to the original lentiviral vectors. We also analyzed the vector copy number in the bone marrow cells by semi-quantitative PCR. To our surprise we found that on average there were multiple vector copies integrated per transduced cell (5.6±3.3 n=12). While the multiple vector copy integration into stem cells is efficient in terms of transduction and expression, it may increase the risk for insertional mutagenesis. Transduction of murine embryonic stem cells was performed to study transgene expression throughout in vitro differentiation to hematopoietic cells. In addition, we optimized the conditions for gene transfer into murine hematopoietic progenitors in order to be able to test hematopoietic stem cell gene therapy in animal models of human disease. These studies demonstrate the ability of lentiviral vectors to efficiently transduce human hematopoietic stem cells, and advance lentiviral vectors as a tool for the treatment of hematological disorders.
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