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High-titer foamy virus vector transduction and integration sites of human CD34(+) cell-derived SCID-repopulating cells.

Nasimuzzaman M, Kim YS, Wang YD, Persons DA - Mol Ther Methods Clin Dev (2014)

Bottom Line: Engraftment frequencies of FV vector-transduced cells were significantly higher than those of LV vector-transduced cells.Linear amplification-mediated PCR with Illumina paired-end runs showed that all human chromosomes contained FV provirus.Repopulating lymphoid and myeloid cells contained common integration sites, suggesting that FV vector could transduce multilineage hematopoietic stem/progenitor populations.

View Article: PubMed Central - PubMed

Affiliation: Division of Experimental Hematology, Department of Hematology, St. Jude Children's Research Hospital , Memphis, Tennessee, USA.

ABSTRACT
Foamy virus (FV) vectors are promising tools for gene therapy, but low titer is a major challenge for large-scale clinical trials. Here, we increased FV vector titer 50-fold by constructing novel vector plasmids and using polyethylenimine-mediated transfection. FV and lentiviral (LV) vectors were used separately to transduce human CD34(+) cells at multiplicities of infection of 25, and those cells were transplanted into immunodeficient mice. FV vector transduction frequencies of repopulating human cells were 37.1 ± 1.9% in unstimulated cells and 36.9 ± 2.2% in prestimulated cells, and engraftment frequencies were 40.9 ± 4.9% in unstimulated cells and 47.1 ± 3.3% in prestimulated cells. Engraftment frequencies of FV vector-transduced cells were significantly higher than those of LV vector-transduced cells. Linear amplification-mediated PCR with Illumina paired-end runs showed that all human chromosomes contained FV provirus. FV had an integration preference near transcriptional start sites and CpG islands of RefSeq genes but not within genes. Repopulating lymphoid and myeloid cells contained common integration sites, suggesting that FV vector could transduce multilineage hematopoietic stem/progenitor populations. Our new FV vector backbone may be a suitable candidate for developing therapeutic FV vectors for use in clinical trials.

No MeSH data available.


Related in: MedlinePlus

Human CD34+ cells in bone marrow of NSG mice. (a) Percentages of CD34+ cells in the engrafted human CD45+ cells. (b) Proportion of GFP+ cells in the CD34+ population. Data represent the mean and SEM of three individual experiments.
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fig4: Human CD34+ cells in bone marrow of NSG mice. (a) Percentages of CD34+ cells in the engrafted human CD45+ cells. (b) Proportion of GFP+ cells in the CD34+ population. Data represent the mean and SEM of three individual experiments.

Mentions: CD34+ cells within human CD45+ population revealed similar percentages for both the FV and LV vector-transduced cells (10.7–13.1%; Figure 4a). GFP marking was consistent with the CD34+ population for both transduction conditions for FV and LV vectors (Figure 4b).


High-titer foamy virus vector transduction and integration sites of human CD34(+) cell-derived SCID-repopulating cells.

Nasimuzzaman M, Kim YS, Wang YD, Persons DA - Mol Ther Methods Clin Dev (2014)

Human CD34+ cells in bone marrow of NSG mice. (a) Percentages of CD34+ cells in the engrafted human CD45+ cells. (b) Proportion of GFP+ cells in the CD34+ population. Data represent the mean and SEM of three individual experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4441046&req=5

fig4: Human CD34+ cells in bone marrow of NSG mice. (a) Percentages of CD34+ cells in the engrafted human CD45+ cells. (b) Proportion of GFP+ cells in the CD34+ population. Data represent the mean and SEM of three individual experiments.
Mentions: CD34+ cells within human CD45+ population revealed similar percentages for both the FV and LV vector-transduced cells (10.7–13.1%; Figure 4a). GFP marking was consistent with the CD34+ population for both transduction conditions for FV and LV vectors (Figure 4b).

Bottom Line: Engraftment frequencies of FV vector-transduced cells were significantly higher than those of LV vector-transduced cells.Linear amplification-mediated PCR with Illumina paired-end runs showed that all human chromosomes contained FV provirus.Repopulating lymphoid and myeloid cells contained common integration sites, suggesting that FV vector could transduce multilineage hematopoietic stem/progenitor populations.

View Article: PubMed Central - PubMed

Affiliation: Division of Experimental Hematology, Department of Hematology, St. Jude Children's Research Hospital , Memphis, Tennessee, USA.

ABSTRACT
Foamy virus (FV) vectors are promising tools for gene therapy, but low titer is a major challenge for large-scale clinical trials. Here, we increased FV vector titer 50-fold by constructing novel vector plasmids and using polyethylenimine-mediated transfection. FV and lentiviral (LV) vectors were used separately to transduce human CD34(+) cells at multiplicities of infection of 25, and those cells were transplanted into immunodeficient mice. FV vector transduction frequencies of repopulating human cells were 37.1 ± 1.9% in unstimulated cells and 36.9 ± 2.2% in prestimulated cells, and engraftment frequencies were 40.9 ± 4.9% in unstimulated cells and 47.1 ± 3.3% in prestimulated cells. Engraftment frequencies of FV vector-transduced cells were significantly higher than those of LV vector-transduced cells. Linear amplification-mediated PCR with Illumina paired-end runs showed that all human chromosomes contained FV provirus. FV had an integration preference near transcriptional start sites and CpG islands of RefSeq genes but not within genes. Repopulating lymphoid and myeloid cells contained common integration sites, suggesting that FV vector could transduce multilineage hematopoietic stem/progenitor populations. Our new FV vector backbone may be a suitable candidate for developing therapeutic FV vectors for use in clinical trials.

No MeSH data available.


Related in: MedlinePlus