Retrovirus-mediated gene transfer into rhesus monkey hematopoietic stem cells: The effect of viral titers on transduction efficiency
article
We have generated a cell line, designated POAM-P1, shedding amphotropic recombinant retroviruses carrying the human adenosine deaminase (hADA) gene. It exhibits a 1 log increased retrovirus titer on NIH-3T3 cells and a five-fold more efficient transduction of human ADA-deficient T lymphocytes, as compared to the previously generated cell line POC-1 which produces the same recombinant hADA retrovirus. To study whether the titer of retrovirus-producing cell lines influences the transduction efficiency of hematopoietic stem cells in a co-culture setting, we compared the POAM-P1 and POC-1 cell lines with respect to their gene transfer efficiency on rhesus monkey bone marrow. Following co-cultivation of rhesus monkey bone marrow with POAM-P1 cells, successful transduction could be demonstrated in approximately 10% of myeloid progenitor colonies (CFU-C) and 0.1% of peripheral blood mononuclear cells (PBMC) and granulocytes in vivo until > 1 year after autologous transplantation. In addition, the presence of functional hADA enzyme was detected in red blood cells, PBMC, and granulocytes. Monkeys receiving POC-1 co-cultured bone marrow carried transduced blood cells for > 2 years after transplantation. Despite the higher retrovirus titer of POAM-P1 cells as compared to POC-1 cells, no difference was observed in gene transfer efficiency into CFU-C and long-term repopulating stem cells. This shows that in our co-cultivation procedure the retrovirus titer was not limiting the transduction efficiency of primate hematopoietic stem cells.
Topics
Cell enzymeGranulocyte enzymeAnimal cellAnimal experimentAutotransplantationBlood cellBone marrow cellCell strain 3t3Enzyme localizationErythrocyteGene expressionGene transferGenetic transductionGranulocyteHematopoietic stem cellHumanHuman cellMononuclear cellNonhumanPromyelocyteRetrovirusRhesus monkeyT lymphocyteVirus recombinant3T3 CellsAnimalBone Marrow TransplantationCells, CulturedGenetic VectorsHematopoietic Stem CellsMacaca mulattaMiceRetroviridaeSupport, Non-U.S. Gov'tTransduction, GeneticTransfectionTransplantation, AutologousMacaca mulattaPrimatesunidentified retrovirusAdenosine deaminase, 9026-93-1Adenosine Deaminase, EC 3.5.4.4Genetic Vectors
TNO Identifier
232076
ISSN
1043-0342
Source
Human Gene Therapy, 4(3), pp. 239-247.
Pages
239-247
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