Correlation of unstable multidrug cross resistance in Chinese hamster ovary cells with a homogeneously staining region on chromosome 1.

Grund, S.; Patil, Sandeep; Shah, Harshit; Pauw, Peter G.; Stadler, Janice

doi:10.1128/mcb.3.9.1634

Cited by 29 publications

(5 citation statements)

References 32 publications

(31 reference statements)

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“…The presence of double minute chromosomes and homogeneously staining regions in multidrug-resistant human and rodent cells is consistent with this mechanism (1,8,11). More recently, the development of in-gel renaturation techniques by Roninson (19) has allowed the direct visualization of amplified genomic DNA.…”

supporting

confidence: 61%

Chromosome-mediated gene transfer of multidrug resistance.

Gros¹,

Fallows²,

Croop³

et al. 1986

Mol. Cell. Biol.

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supporting

confidence: 61%

Chromosome-mediated gene transfer of multidrug resistance.

Gros¹,

Fallows²,

Croop³

et al. 1986

Mol. Cell. Biol.

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“…It has been suggested that multidrug resistance is due to a decreased intracellular drug accumulation (2,3) caused by alterations of the plasma membrane (4)(5)(6). Multidrug-resistant cells often contain double minute chromosomes or homogeneously staining chromosomal regions, suggesting that gene amplification at least in part underlies multidrug resistance (7)(8)(9)(10)(11).…”

mentioning

confidence: 99%

Isolation and characterization of DNA sequences amplified in multidrug-resistant hamster cells.

Gros¹,

Croop²,

Roninson³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

191

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The mechanism by which mammalian cells acquire resistance to chemotherapeutic agents has been investigated by using molecular genetic techniques. LZ and C5, two independently derived multidrug-resistant Chinese hamster cell lines, share specific amplified DNA sequences. We demonstrate that commonly amplified DNA sequences reside in a contiguous domain of :120 kilobases (kb). We report the isolation of this DNA domain in cosmid clones and show that the level of amplification of the domain is correlated with the level of resistance in multidrug-resistant cell lines. The organization of the amplified domain was deduced by a unique approach utilizing in-gel hybridization of cloned DNA with amplified genomic DNA. We show that the entire cloned region is amplified in adriamycin-resistant LZ cells and independently derived, colchicine-resistant C5 cells. A mRNA species of '-5 kb is encoded by a gene located within the boundaries of this region. Genomic sequences homologous to the 5-kb mRNA span over 75 kb of the amplified DNA segment. The level of expression of this mRNA in multidrug-resistant cells is correlated with the degree of gene amplification and the degree of drug resistance. Our results strongly suggest that the 5-kb mRNA species plays a role in the mechanism of multidrug resistance common to the LZ and C5 cell lines.The use of cytotoxic agents in cancer therapy is often complicated by the emergence of multidrug-resistant tumor cells. Cultured mammalian cells that exhibit analogous multidrug-resistance phenotypes present an attractive model to study this phenomenon (1). It has been suggested that multidrug resistance is due to a decreased intracellular drug accumulation (2, 3) caused by alterations of the plasma membrane (4-6). Multidrug-resistant cells often contain double minute chromosomes or homogeneously staining chromosomal regions, suggesting that gene amplification at least in part underlies multidrug resistance (7-11).Two independently derived multidrug-resistant Chinese hamster cell lines, C5 (4) and LZ (11), were used in our studies. LZ, a highly resistant line, and 77A, a weakly resistant line, were derived by stepwise selection with increasing concentrations of adriamycin from drug-sensitive V79 cells (11). C5, a highly colchicine-resistant line, was derived from drug-sensitive Chinese hamster ovary cells (12). CS cells are cross-resistant to a number of cytotoxic agents, including adriamycin and colchicine. In our previous study, we used the technique of in-gel renaturation (13) to directly demonstrate the presence of amplified DNA sequences in the LZ and C5 genomes (14). It was also shown that a subset of amplified LZ DNA sequences was amplified in common between the two cell lines. One of the commonly amplified DNA fragments was cloned, and the degree of its amplification in the cells was shown to correlate with the degree of their drug resistance (14).In the present study, using phage and cosmid vectors, we have cloned a contiguous domain of DNA sequences that spans ==120 kilobases (k...

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“…Homogeneously staining regions or double minute chromosomes, both known to carry amplified DNA, have been observed in a variety of multidrug-resistant cell lines (3,5,12,29). The degree of amplification, reflected by the size of the homogeneously staining region or the number of double minute chromosomes, can be correlated with the level of P-glycoprotein overproduction (5,29,31).…”

mentioning

confidence: 99%

Overexpression and amplification of five genes in a multidrug-resistant Chinese hamster ovary cell line.

Bliek¹,

Velde-Koerts²,

Ling³

et al. 1986

Mol. Cell. Biol.

246

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(25,26). Their overproduction depends on the drug used for selection, however. In addition, the putative mouse homolog (21 kDa) of V19 has recently been reported to disappear after prolonged culture, even though multidrug resistance was retained (24). The loss of proteins (72 and 75 kDa) has also been found in a multidrug-resistant cell line (27), completing the spectrum of altered protein levels which have been associated with * Corresponding author. multidrug resistance. How each of these proteins contributes to the ability of the cell to maintain a low intracellular drug concentration is unknown.The overproduction of proteins, and in particular those involved in drug resistance in mammalian cells, is often the result of gene amplification (see references 7, 14, 21, 33, 36). Homogeneously staining regions or double minute chromosomes, both known to carry amplified DNA, have been observed in a variety of multidrug-resistant cell lines (3,5,12,29). The degree of amplification, reflected by the size of the homogeneously staining region or the number of double minute chromosomes, can be correlated with the level of P-glycoprotein overproduction (5,29,31). Recently, a cDNA clone encoding part of the P-glycoprotein has been isolated from an expression library (29). This clone hybridizes to a homogeneously staining region, and, in line with the cytological observations, the degree of amplification corresponds to the level of resistance. Although overproduction of the 170-kDa P-glycoprotein remains the most consistent feature, homogeneously staining regions have also been observed in multidrug-resistant cell lines without an apparent overproduction of the P-glycoprotein (24).The disparity in altered protein levels and gene amplification suggests that several proteins contribute to the multidrug-resistant phenotype. To identify genes encoding proteins involved in multidrug resistance, we constructed a cDNA library from the resistant chinese hamster ovary (CHO) cell line CH"C5 and isolated cDNA clones derived from overexpressed transcripts by differential hybridization. In this paper we describe the initial characterization of cDNAs corresponding to at least five genes that are overexpressed and amplified in the resistant cell line.MATERIALS AND METHODS Cell culture. The CHO cell lines AUXB1 and CHRC5 were cultured as described previously (22) without colchicine.

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Correlation of unstable multidrug cross resistance in Chinese hamster ovary cells with a homogeneously staining region on chromosome 1.

Abstract: An enrichment selection method using repeated pulses of low drug concentration (1 ,ug/ml) was

Cited by 29 publications

References 32 publications

Chromosome-mediated gene transfer of multidrug resistance.

Chromosome-mediated gene transfer of multidrug resistance.

Isolation and characterization of DNA sequences amplified in multidrug-resistant hamster cells.

Overexpression and amplification of five genes in a multidrug-resistant Chinese hamster ovary cell line.

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