Selection against expression of the Escherichia coli gene gpt in hprt+ mouse teratocarcinoma and hybrid cells.

Besnard, C; Monthioux, E; Jami, J

doi:10.1128/mcb.7.11.4139

Cited by 27 publications

(7 citation statements)

References 10 publications

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“…In early-generation retroviral vectors, this gene proved useful as a selectable marker by virtue of its ability to impart resistance to a mycophenolic acid/xanthine/ hypoxanthine (MXH) regimen, in which XGPRT-catalyzed xanthine incorporation permits cells to circumvent a mycophenolic acid-induced block in de novo guanine nucleotide synthesis, and hypoxanthine may aid selection by inhibiting guanine salvage pathways Berg, 1980, 1981;Perkins et al, 1983). On the other hand, as transfected DNA, the same gene was recently reported capable of imparting sensitivity to a xanthine analog, 6-thioxanthine (6TX, Besnard et al, 1987)-a consequence of XGPRT-catalyzed incorporation of this analog into the same de novo synthetic pathways. Therefore, this study was conducted first to determine in vitro whether the inclusion of the gpt gene in a retroviral vector might permit that vector to mediate both selectability and sensitivity in the same cell population, thereby obviating a need to use two genes for this purpose, and second to determine whether the "suicide" potential of the gene could be translated into tumor eradication in vivo.…”

Section: Introductionmentioning

confidence: 97%

Retrovirally TransducedEscherichia coli gptGenes Combine Selectability with Chemosensitivity Capable of Mediating Tumor Eradication

Mroz

Moolten²

1993

Human Gene Therapy

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"Suicide genes" encoding exceptional sensitivity to chemotherapeutic agents can potentially improve the selectivity of cancer therapy. As a component of a retroviral gene therapy vector, a suicide gene might also improve the safety of gene therapy by permitting the subsequent ablation of transduced cells exhibiting neoplastic or other aberrant behavior. An extra gene, however, cannot easily be added to vectors already carrying a therapeutic gene and a selectable drug resistance marker without compromising gene expression. To circumvent this obstacle, we have investigated a retrovirally transduced Escherichia coli gpt gene on the basis of evidence that it might subserve a dual sensitivity/resistance function. A gpt vector was used to transduce the gene into murine K3T3 sarcoma cells. In vitro, gpt-positive K3T3 clones could be selected on the basis of resistance to a regimen containing mycophenolic acid and xanthine; the same clones were 18 to 86 times as sensitive to 6-thioxanthine (6TX) as their gpt-negative counterparts. In mice, systemic 6TX therapy induced durable regressions in 19/20 gpt-positive K3T3 sarcomas without affecting gpt-negative tumors. These results indicate that selectability and in vivo chemosensitivity can be expressed in the same cell population from a single retrovirally transduced gene and imply the additional possibility of fusing the gpt gene with a therapeutic gene to create vectors expressing three important functions from a single gene.

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Section: Introductionmentioning

confidence: 97%

Retrovirally TransducedEscherichia coli gptGenes Combine Selectability with Chemosensitivity Capable of Mediating Tumor Eradication

Mroz

Moolten²

1993

Human Gene Therapy

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“…By this method, a large Most mutation assays based on a transgenic marker suitable for forward and reverse selection have used the bacterial gpt gene. In hprf recipients, positive selection is effected with mycophenolic acid (plus xanthine) and negative selection with 6-thioguanine [39]. Transgenic cell lines carrying a single, stably integrated copy of the gene have been found to be responsive to induced inactivation of the transgene by both point mutation and deletion.…”

Section: Discussionmentioning

confidence: 99%

Novel use of a selectable fusion gene as an “In‐Out” marker for studying genetic loss in mammalian cells

Trott

Cuthbert

Todd

et al. 1995

Molecular Carcinogenesis

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Recent demonstrations of loss of heterozygosity in a wide variety of human cancers suggest that large multilocus genetic deletions (presumably including tumor suppressor genes) constitute a major class of genetic alteration in human carcinogenesis. Here we show that a bifunctional fusion gene (Hytk), suitable for both positive and negative selection, is an effective marker for studying genetic loss in mammalian cells with minimal interference from point-mutational changes. Studies with a transgenic V79 cell line in which a single functional copy of Hytk was stably inserted into the genome in a retroviral vector showed that loss of the marker (and presumably flanking cellular genetic material) could be induced efficiently by ionizing radiation (gamma-rays and fast neutrons) but only weakly by the powerful point-mutagen benzo[a]pyrene diol epoxide. In a first application of the system, we provide evidence that radiation-induced loss can occur through an indirect mechanism after a high-frequency event. Collectively, our results suggest that the Hytk marker should be a valuable tool for studying genome position effects on the tolerance of genetic loss in cultured human cells that represent different stages in clonal evolution and tumor progression.

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“…ECO-GPT is a useful counter-selection marker gene applicable for both positive selection (with Mycophenolic acid (MPA)) and negative selection (with 6-thioxyanthine (6-TX)) 18 . Clones carrying ECO-GPT marker at TOPBP1 locus were first selected with MPA, and the resultant TOPBP1 +/+/ECO-GPT clones (Supplementary Fig.…”

Section: Correction Of Chromosome 2 Trisomy In Dt40 Cellsmentioning

confidence: 99%

Targeting Chromosome Trisomy for Chromosome Editing

Abe

Suzuki

Hirota

2021

Preprint

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A trisomy is a type of aneuploidy characterised by an additional chromosome. The additional chromosome theoretically accepts any kind of changes since it is not necessary for cellular proliferation. This advantage led us to apply two chromosome manipulation methods to autosomal trisomy in chicken DT40 cells. We first corrected chromosome 2 trisomy to disomy by employing counter-selection makers. Upon construction of cells carrying makers targeted in one of the trisomic chromosome 2s, cells that have lost makers integrated in chromosome 2 were subsequently selected. The loss of one of the chromosome 2s had little impacts on the proliferative capacity, indicating unsubstantial role of the additional chromosome 2 in DT40 cells. We next tested large-scale truncations of chromosome 2 to make a mini-chromosome for the assessment of chromosome stability by introducing telomere repeat sequences to delete most of p-arm or q-arm of chromosome 2. The obtained cell lines had 0.7 Mb mini-chromosome, and approximately 0.2% of mini-chromosome was lost per cell division in wild-type background while the rate of chromosome loss was significantly increased by the depletion of DDX11, a cohesin regulatory protein. Collectively, our findings propose that trisomic chromosomes are good targets to make unique artificial chromosomes. (197 words)

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Selection against expression of the Escherichia coli gene gpt in hprt+ mouse teratocarcinoma and hybrid cells.

Cited by 27 publications

References 10 publications

Retrovirally TransducedEscherichia coli gptGenes Combine Selectability with Chemosensitivity Capable of Mediating Tumor Eradication

Retrovirally TransducedEscherichia coli gptGenes Combine Selectability with Chemosensitivity Capable of Mediating Tumor Eradication

Novel use of a selectable fusion gene as an “In‐Out” marker for studying genetic loss in mammalian cells

Targeting Chromosome Trisomy for Chromosome Editing

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