Genetically modified hematopoietic stem cells expressing o6methylguanine dna methyltransferase via a retrovirus vector protect the lymphoid and stem cell compartments in long-term engrafted animals treated with nitrosourea

R. Maze, Reuben Kapur, D. A. Williams

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Abstract

Bone marrow {BM) toxicity is a dose-limiting side effect of chloroethylnitrosoureas (CNU). A major determinant of CNU cytotoxidty is the methylation of O6-guanine and the formation of interstrand DNA crosslinks. Methylguanine DNA methyltransferase (MGMT) removes alkyl groups from the Opposition and has been shown to repair CNU-induced DNA damage. BM cells express a low level of MGMT. We have previously demonstrated that reconstitution of marine BM with genetically modified MGMT-expressing hematopoietic stem cells (MGMT-HSCs) significantly reduces CNU-related pancytopenia (Maze et al., PNAS, in press). In the present study, we examined the long-term effect of l,3-bis(2chloroctbylH-nitrosourea (BCNU) treatment and vector-derived expression of MGMT in hematopoietic stem cells (HSCs) on the lymphoid and HSC compartments following bone marrow transplantation (BMT). Mice were transplanted with MGMT or mock-infected (control) BM and treated for 5 weeks with BCNU (40 rag/kg). Five months post-BMT, bom the total thymic cellularity and the percentage of immature CD4+CD8+ double positive cells were significantly reduced in surviving BCNU-treated control mice vs. mice transplanted with MGMT-HSCs (7.3 ±3.6xl06 and 1.68% vs. 28.0i3.6×106 and 57.8%, respectively). Mice transplanted with MGMT-HSCs also had a normal frequency of mature B cells in the spleen (30.3%), based on B220 and IgD cell surface markers, whereas spleen cells from BCNU-treated control mice were depleted of mature B cells (1.63%). BCNU-treated mice transplanted with MGMT-HSCs demonstrated: a greater frequency of stem cells, based on serial transplants; BCNU resistance in vitro; and a higher level of DNA repair activity compared to BCNU-treated control mice. These studies demonstrate that MGMT-HSCs protect both the (ymphoid and HSC compartments from CNU-induced damage. Together with our previous data on myeloid protection, these data suggest an opportunity for CNU dose-intensification following autologous BMT in the treatment of CNU-sensitive tumors using gene therapy.

Original languageEnglish
JournalJournal of Investigative Medicine
Volume44
Issue number3
StatePublished - 1996

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Lymphoid Progenitor Cells
Methyltransferases
Retroviridae
Hematopoietic Stem Cells
Stem cells
Animals
DNA
Bone
Bone Marrow Transplantation
Bone Marrow
Cells
B-Lymphocytes
Spleen
Repair
Computer Security
Immunoglobulin D
Pancytopenia
Autologous Transplantation
Guanine
Gene therapy

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

@article{a7ed35f7991b43dabe1a86cbc565bc66,
title = "Genetically modified hematopoietic stem cells expressing o6methylguanine dna methyltransferase via a retrovirus vector protect the lymphoid and stem cell compartments in long-term engrafted animals treated with nitrosourea",
abstract = "Bone marrow {BM) toxicity is a dose-limiting side effect of chloroethylnitrosoureas (CNU). A major determinant of CNU cytotoxidty is the methylation of O6-guanine and the formation of interstrand DNA crosslinks. Methylguanine DNA methyltransferase (MGMT) removes alkyl groups from the Opposition and has been shown to repair CNU-induced DNA damage. BM cells express a low level of MGMT. We have previously demonstrated that reconstitution of marine BM with genetically modified MGMT-expressing hematopoietic stem cells (MGMT-HSCs) significantly reduces CNU-related pancytopenia (Maze et al., PNAS, in press). In the present study, we examined the long-term effect of l,3-bis(2chloroctbylH-nitrosourea (BCNU) treatment and vector-derived expression of MGMT in hematopoietic stem cells (HSCs) on the lymphoid and HSC compartments following bone marrow transplantation (BMT). Mice were transplanted with MGMT or mock-infected (control) BM and treated for 5 weeks with BCNU (40 rag/kg). Five months post-BMT, bom the total thymic cellularity and the percentage of immature CD4+CD8+ double positive cells were significantly reduced in surviving BCNU-treated control mice vs. mice transplanted with MGMT-HSCs (7.3 ±3.6xl06 and 1.68{\%} vs. 28.0i3.6×106 and 57.8{\%}, respectively). Mice transplanted with MGMT-HSCs also had a normal frequency of mature B cells in the spleen (30.3{\%}), based on B220 and IgD cell surface markers, whereas spleen cells from BCNU-treated control mice were depleted of mature B cells (1.63{\%}). BCNU-treated mice transplanted with MGMT-HSCs demonstrated: a greater frequency of stem cells, based on serial transplants; BCNU resistance in vitro; and a higher level of DNA repair activity compared to BCNU-treated control mice. These studies demonstrate that MGMT-HSCs protect both the (ymphoid and HSC compartments from CNU-induced damage. Together with our previous data on myeloid protection, these data suggest an opportunity for CNU dose-intensification following autologous BMT in the treatment of CNU-sensitive tumors using gene therapy.",
author = "R. Maze and Reuben Kapur and Williams, {D. A.}",
year = "1996",
language = "English",
volume = "44",
journal = "Journal of Investigative Medicine",
issn = "1081-5589",
publisher = "Lippincott Williams and Wilkins",
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T1 - Genetically modified hematopoietic stem cells expressing o6methylguanine dna methyltransferase via a retrovirus vector protect the lymphoid and stem cell compartments in long-term engrafted animals treated with nitrosourea

AU - Maze, R.

AU - Kapur, Reuben

AU - Williams, D. A.

PY - 1996

Y1 - 1996

N2 - Bone marrow {BM) toxicity is a dose-limiting side effect of chloroethylnitrosoureas (CNU). A major determinant of CNU cytotoxidty is the methylation of O6-guanine and the formation of interstrand DNA crosslinks. Methylguanine DNA methyltransferase (MGMT) removes alkyl groups from the Opposition and has been shown to repair CNU-induced DNA damage. BM cells express a low level of MGMT. We have previously demonstrated that reconstitution of marine BM with genetically modified MGMT-expressing hematopoietic stem cells (MGMT-HSCs) significantly reduces CNU-related pancytopenia (Maze et al., PNAS, in press). In the present study, we examined the long-term effect of l,3-bis(2chloroctbylH-nitrosourea (BCNU) treatment and vector-derived expression of MGMT in hematopoietic stem cells (HSCs) on the lymphoid and HSC compartments following bone marrow transplantation (BMT). Mice were transplanted with MGMT or mock-infected (control) BM and treated for 5 weeks with BCNU (40 rag/kg). Five months post-BMT, bom the total thymic cellularity and the percentage of immature CD4+CD8+ double positive cells were significantly reduced in surviving BCNU-treated control mice vs. mice transplanted with MGMT-HSCs (7.3 ±3.6xl06 and 1.68% vs. 28.0i3.6×106 and 57.8%, respectively). Mice transplanted with MGMT-HSCs also had a normal frequency of mature B cells in the spleen (30.3%), based on B220 and IgD cell surface markers, whereas spleen cells from BCNU-treated control mice were depleted of mature B cells (1.63%). BCNU-treated mice transplanted with MGMT-HSCs demonstrated: a greater frequency of stem cells, based on serial transplants; BCNU resistance in vitro; and a higher level of DNA repair activity compared to BCNU-treated control mice. These studies demonstrate that MGMT-HSCs protect both the (ymphoid and HSC compartments from CNU-induced damage. Together with our previous data on myeloid protection, these data suggest an opportunity for CNU dose-intensification following autologous BMT in the treatment of CNU-sensitive tumors using gene therapy.

AB - Bone marrow {BM) toxicity is a dose-limiting side effect of chloroethylnitrosoureas (CNU). A major determinant of CNU cytotoxidty is the methylation of O6-guanine and the formation of interstrand DNA crosslinks. Methylguanine DNA methyltransferase (MGMT) removes alkyl groups from the Opposition and has been shown to repair CNU-induced DNA damage. BM cells express a low level of MGMT. We have previously demonstrated that reconstitution of marine BM with genetically modified MGMT-expressing hematopoietic stem cells (MGMT-HSCs) significantly reduces CNU-related pancytopenia (Maze et al., PNAS, in press). In the present study, we examined the long-term effect of l,3-bis(2chloroctbylH-nitrosourea (BCNU) treatment and vector-derived expression of MGMT in hematopoietic stem cells (HSCs) on the lymphoid and HSC compartments following bone marrow transplantation (BMT). Mice were transplanted with MGMT or mock-infected (control) BM and treated for 5 weeks with BCNU (40 rag/kg). Five months post-BMT, bom the total thymic cellularity and the percentage of immature CD4+CD8+ double positive cells were significantly reduced in surviving BCNU-treated control mice vs. mice transplanted with MGMT-HSCs (7.3 ±3.6xl06 and 1.68% vs. 28.0i3.6×106 and 57.8%, respectively). Mice transplanted with MGMT-HSCs also had a normal frequency of mature B cells in the spleen (30.3%), based on B220 and IgD cell surface markers, whereas spleen cells from BCNU-treated control mice were depleted of mature B cells (1.63%). BCNU-treated mice transplanted with MGMT-HSCs demonstrated: a greater frequency of stem cells, based on serial transplants; BCNU resistance in vitro; and a higher level of DNA repair activity compared to BCNU-treated control mice. These studies demonstrate that MGMT-HSCs protect both the (ymphoid and HSC compartments from CNU-induced damage. Together with our previous data on myeloid protection, these data suggest an opportunity for CNU dose-intensification following autologous BMT in the treatment of CNU-sensitive tumors using gene therapy.

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