Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells.
BACKGROUND:Hematopoietic stem cells (HSC), in particular mobilized peripheral blood stem cells, represent an attractive target for cell and gene therapy. Efficient gene delivery into these target cells without compromising self-renewal and multi-potency is crucial for the success of gene therapy. We...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2009-07-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC2714083?pdf=render |
| _version_ | 1818136829781606400 |
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| author | Michelle Millington Allison Arndt Maureen Boyd Tanya Applegate Sylvie Shen |
| author_facet | Michelle Millington Allison Arndt Maureen Boyd Tanya Applegate Sylvie Shen |
| author_sort | Michelle Millington |
| collection | DOAJ |
| description | BACKGROUND:Hematopoietic stem cells (HSC), in particular mobilized peripheral blood stem cells, represent an attractive target for cell and gene therapy. Efficient gene delivery into these target cells without compromising self-renewal and multi-potency is crucial for the success of gene therapy. We investigated factors involved in the ex vivo transduction of CD34(+) HSCs in order to develop a clinically relevant transduction protocol for gene delivery. Specifically sought was a protocol that allows for efficient transduction with minimal ex vivo manipulation without serum or other reagents of animal origin. METHODOLOGY/PRINCIPAL FINDINGS:Using commercially available G-CSF mobilized peripheral blood (PB) CD34(+) cells as the most clinically relevant target, we systematically examined factors including the use of serum, cytokine combinations, pre-stimulation time, multiplicity of infection (MOI), transduction duration and the use of spinoculation and/or retronectin. A self-inactivating lentiviral vector (SIN-LV) carrying enhanced green fluorescent protein (GFP) was used as the gene delivery vehicle. HSCs were monitored for transduction efficiency, surface marker expression and cellular function. We were able to demonstrate that efficient gene transduction can be achieved with minimal ex vivo manipulation while maintaining the cellular function of transduced HSCs without serum or other reagents of animal origin. CONCLUSIONS/SIGNIFICANCE:This study helps to better define factors relevant towards developing a standard clinical protocol for the delivery of SIN-LV into CD34(+) cells. |
| first_indexed | 2024-12-11T09:46:38Z |
| format | Article |
| id | doaj.art-1e821a1cae8546abb6f954d7bd4e0f55 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-11T09:46:38Z |
| publishDate | 2009-07-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-1e821a1cae8546abb6f954d7bd4e0f552022-12-22T01:12:33ZengPublic Library of Science (PLoS)PLoS ONE1932-62032009-07-0147e646110.1371/journal.pone.0006461Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells.Michelle MillingtonAllison ArndtMaureen BoydTanya ApplegateSylvie ShenBACKGROUND:Hematopoietic stem cells (HSC), in particular mobilized peripheral blood stem cells, represent an attractive target for cell and gene therapy. Efficient gene delivery into these target cells without compromising self-renewal and multi-potency is crucial for the success of gene therapy. We investigated factors involved in the ex vivo transduction of CD34(+) HSCs in order to develop a clinically relevant transduction protocol for gene delivery. Specifically sought was a protocol that allows for efficient transduction with minimal ex vivo manipulation without serum or other reagents of animal origin. METHODOLOGY/PRINCIPAL FINDINGS:Using commercially available G-CSF mobilized peripheral blood (PB) CD34(+) cells as the most clinically relevant target, we systematically examined factors including the use of serum, cytokine combinations, pre-stimulation time, multiplicity of infection (MOI), transduction duration and the use of spinoculation and/or retronectin. A self-inactivating lentiviral vector (SIN-LV) carrying enhanced green fluorescent protein (GFP) was used as the gene delivery vehicle. HSCs were monitored for transduction efficiency, surface marker expression and cellular function. We were able to demonstrate that efficient gene transduction can be achieved with minimal ex vivo manipulation while maintaining the cellular function of transduced HSCs without serum or other reagents of animal origin. CONCLUSIONS/SIGNIFICANCE:This study helps to better define factors relevant towards developing a standard clinical protocol for the delivery of SIN-LV into CD34(+) cells.http://europepmc.org/articles/PMC2714083?pdf=render |
| spellingShingle | Michelle Millington Allison Arndt Maureen Boyd Tanya Applegate Sylvie Shen Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. PLoS ONE |
| title | Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. |
| title_full | Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. |
| title_fullStr | Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. |
| title_full_unstemmed | Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. |
| title_short | Towards a clinically relevant lentiviral transduction protocol for primary human CD34 hematopoietic stem/progenitor cells. |
| title_sort | towards a clinically relevant lentiviral transduction protocol for primary human cd34 hematopoietic stem progenitor cells |
| url | http://europepmc.org/articles/PMC2714083?pdf=render |
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