In vivo selection of hematopoietic stem cells transduced at a low multiplicity-of-infection with a foamy viral MGMTP140K vector

Shanbao Cai, Aaron Ernstberger, Haiyan Wang, Barbara J. Bailey, Jennifer R. Hartwell, Anthony L. Sinn, Olaf Eckermann, Yvonne Linka, W. Goebel, Helmut Hanenberg, Karen Pollok

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Abstract

Objective: Using a clinically relevant transduction strategy, we investigated to what extent hematopoietic stem cells in lineage-negative bone marrow (Linneg BM) could be genetically modified with an foamy virus (FV) vector that expresses the DNA repair protein, O6-methylguanine DNA methyltransferase (MGMTP140K) and selected in vivo with submyeloablative or myeloablative alkylator therapy. Materials and Methods: Linneg BM was transduced at a low multiplicity-of-infection with the FV vector, MD9-P140K, which coexpresses MGMTP140K and the enhanced green fluorescent protein, transplanted into C57BL/6 mice, and mice treated with submyeloablative or myeloablative alkylator therapy. The BM was analyzed for the presence of in vivo selected, MD9-P140K-transduced cells at 6 months post-transplantation and subsequently transplanted into secondary recipient animals. Results: Following submyeloablative therapy, 55% of the mice expressed MGMTP140K in the BM. Proviral integration was observed in ∼50% of committed BM-derived progenitors and analysis of proviral insertion sites indicated up to two integrations per transduced progenitor colony. Transduced BM cells selected with submyeloablative therapy reconstituted secondary recipient mice for up to 6 months post-transplantation. In contrast, after delivery of myeloablative therapy to primary recipient mice, only 25% survived. Hematopoietic stem cells were transduced because BM cells from the surviving animals reconstituted secondary recipients with MGMTP140K-positive cells for 5 to 6 months. Conclusions: In vivo selection of MD9-P140K-transduced BM cells was more efficient following submyeloablative than myeloablative therapy. These data indicate that a critical number of transduced stem cells must be present to produce sufficient numbers of genetically modified progeny to protect against acute toxicity associated with myeloablative therapy.

Original languageEnglish
Pages (from-to)283-292
Number of pages10
JournalExperimental Hematology
Volume36
Issue number3
DOIs
StatePublished - Mar 2008

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Hematopoietic Stem Cells
Infection
Spumavirus
Alkylating Agents
Therapeutics
Transplantation
Bone Marrow
Methyltransferases
Cell Lineage
Inbred C57BL Mouse
DNA Repair
Stem Cells
DNA
Proteins

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

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In vivo selection of hematopoietic stem cells transduced at a low multiplicity-of-infection with a foamy viral MGMTP140K vector. / Cai, Shanbao; Ernstberger, Aaron; Wang, Haiyan; Bailey, Barbara J.; Hartwell, Jennifer R.; Sinn, Anthony L.; Eckermann, Olaf; Linka, Yvonne; Goebel, W.; Hanenberg, Helmut; Pollok, Karen.

In: Experimental Hematology, Vol. 36, No. 3, 03.2008, p. 283-292.

Research output: Contribution to journalArticle

Cai, S, Ernstberger, A, Wang, H, Bailey, BJ, Hartwell, JR, Sinn, AL, Eckermann, O, Linka, Y, Goebel, W, Hanenberg, H & Pollok, K 2008, 'In vivo selection of hematopoietic stem cells transduced at a low multiplicity-of-infection with a foamy viral MGMTP140K vector', Experimental Hematology, vol. 36, no. 3, pp. 283-292. https://doi.org/10.1016/j.exphem.2007.11.009
Cai, Shanbao ; Ernstberger, Aaron ; Wang, Haiyan ; Bailey, Barbara J. ; Hartwell, Jennifer R. ; Sinn, Anthony L. ; Eckermann, Olaf ; Linka, Yvonne ; Goebel, W. ; Hanenberg, Helmut ; Pollok, Karen. / In vivo selection of hematopoietic stem cells transduced at a low multiplicity-of-infection with a foamy viral MGMTP140K vector. In: Experimental Hematology. 2008 ; Vol. 36, No. 3. pp. 283-292.
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abstract = "Objective: Using a clinically relevant transduction strategy, we investigated to what extent hematopoietic stem cells in lineage-negative bone marrow (Linneg BM) could be genetically modified with an foamy virus (FV) vector that expresses the DNA repair protein, O6-methylguanine DNA methyltransferase (MGMTP140K) and selected in vivo with submyeloablative or myeloablative alkylator therapy. Materials and Methods: Linneg BM was transduced at a low multiplicity-of-infection with the FV vector, MD9-P140K, which coexpresses MGMTP140K and the enhanced green fluorescent protein, transplanted into C57BL/6 mice, and mice treated with submyeloablative or myeloablative alkylator therapy. The BM was analyzed for the presence of in vivo selected, MD9-P140K-transduced cells at 6 months post-transplantation and subsequently transplanted into secondary recipient animals. Results: Following submyeloablative therapy, 55{\%} of the mice expressed MGMTP140K in the BM. Proviral integration was observed in ∼50{\%} of committed BM-derived progenitors and analysis of proviral insertion sites indicated up to two integrations per transduced progenitor colony. Transduced BM cells selected with submyeloablative therapy reconstituted secondary recipient mice for up to 6 months post-transplantation. In contrast, after delivery of myeloablative therapy to primary recipient mice, only 25{\%} survived. Hematopoietic stem cells were transduced because BM cells from the surviving animals reconstituted secondary recipients with MGMTP140K-positive cells for 5 to 6 months. Conclusions: In vivo selection of MD9-P140K-transduced BM cells was more efficient following submyeloablative than myeloablative therapy. These data indicate that a critical number of transduced stem cells must be present to produce sufficient numbers of genetically modified progeny to protect against acute toxicity associated with myeloablative therapy.",
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T1 - In vivo selection of hematopoietic stem cells transduced at a low multiplicity-of-infection with a foamy viral MGMTP140K vector

AU - Cai, Shanbao

AU - Ernstberger, Aaron

AU - Wang, Haiyan

AU - Bailey, Barbara J.

AU - Hartwell, Jennifer R.

AU - Sinn, Anthony L.

AU - Eckermann, Olaf

AU - Linka, Yvonne

AU - Goebel, W.

AU - Hanenberg, Helmut

AU - Pollok, Karen

PY - 2008/3

Y1 - 2008/3

N2 - Objective: Using a clinically relevant transduction strategy, we investigated to what extent hematopoietic stem cells in lineage-negative bone marrow (Linneg BM) could be genetically modified with an foamy virus (FV) vector that expresses the DNA repair protein, O6-methylguanine DNA methyltransferase (MGMTP140K) and selected in vivo with submyeloablative or myeloablative alkylator therapy. Materials and Methods: Linneg BM was transduced at a low multiplicity-of-infection with the FV vector, MD9-P140K, which coexpresses MGMTP140K and the enhanced green fluorescent protein, transplanted into C57BL/6 mice, and mice treated with submyeloablative or myeloablative alkylator therapy. The BM was analyzed for the presence of in vivo selected, MD9-P140K-transduced cells at 6 months post-transplantation and subsequently transplanted into secondary recipient animals. Results: Following submyeloablative therapy, 55% of the mice expressed MGMTP140K in the BM. Proviral integration was observed in ∼50% of committed BM-derived progenitors and analysis of proviral insertion sites indicated up to two integrations per transduced progenitor colony. Transduced BM cells selected with submyeloablative therapy reconstituted secondary recipient mice for up to 6 months post-transplantation. In contrast, after delivery of myeloablative therapy to primary recipient mice, only 25% survived. Hematopoietic stem cells were transduced because BM cells from the surviving animals reconstituted secondary recipients with MGMTP140K-positive cells for 5 to 6 months. Conclusions: In vivo selection of MD9-P140K-transduced BM cells was more efficient following submyeloablative than myeloablative therapy. These data indicate that a critical number of transduced stem cells must be present to produce sufficient numbers of genetically modified progeny to protect against acute toxicity associated with myeloablative therapy.

AB - Objective: Using a clinically relevant transduction strategy, we investigated to what extent hematopoietic stem cells in lineage-negative bone marrow (Linneg BM) could be genetically modified with an foamy virus (FV) vector that expresses the DNA repair protein, O6-methylguanine DNA methyltransferase (MGMTP140K) and selected in vivo with submyeloablative or myeloablative alkylator therapy. Materials and Methods: Linneg BM was transduced at a low multiplicity-of-infection with the FV vector, MD9-P140K, which coexpresses MGMTP140K and the enhanced green fluorescent protein, transplanted into C57BL/6 mice, and mice treated with submyeloablative or myeloablative alkylator therapy. The BM was analyzed for the presence of in vivo selected, MD9-P140K-transduced cells at 6 months post-transplantation and subsequently transplanted into secondary recipient animals. Results: Following submyeloablative therapy, 55% of the mice expressed MGMTP140K in the BM. Proviral integration was observed in ∼50% of committed BM-derived progenitors and analysis of proviral insertion sites indicated up to two integrations per transduced progenitor colony. Transduced BM cells selected with submyeloablative therapy reconstituted secondary recipient mice for up to 6 months post-transplantation. In contrast, after delivery of myeloablative therapy to primary recipient mice, only 25% survived. Hematopoietic stem cells were transduced because BM cells from the surviving animals reconstituted secondary recipients with MGMTP140K-positive cells for 5 to 6 months. Conclusions: In vivo selection of MD9-P140K-transduced BM cells was more efficient following submyeloablative than myeloablative therapy. These data indicate that a critical number of transduced stem cells must be present to produce sufficient numbers of genetically modified progeny to protect against acute toxicity associated with myeloablative therapy.

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