Cellular heterogeneity and drug resistance in two ovarian adenocarcinoma cell lines derived from a single patient

Wolf, C. Roland; Hayward, Ian Philip; Lawrie, Sandra S.; Buckton, K. E.; McIntyre, Margaret; Adams, Daniel J.; Lewis, Alexander D.; Scott, Angela; Smyth, John F.

doi:10.1002/ijc.2910390607

Cited by 128 publications

(86 citation statements)

References 29 publications

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“…Kinetic analyses done in vitro (461, and enhanced anti-BPDE cytotoxicity of 1.7-fold, relative to controls, caused by depletion of intracellular glutathione levels (17) to 10-60% of GST's K, for glutathione (0.1 mM) (22,461, were predictive of recombinant GST .rr + 'S ability to confer resistance to anti-BPDE. Significantly, values of 1.5-fold resistance relative to controls are identical with resistance values to anticancer alkylating agents observed clinically (30,31,58).…”

Section: Discussionsupporting

confidence: 74%

Recombinant glutathione S‐transferase (GST) expressing cells purified by flow cytometry on the basis of a GST‐catalyzed intracellular conjugation of glutathione to monochlorobimane

et al. 1991

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COS cells transiently expressing glutathione S-transferase (GST) m, Ya, or Yb, (human Pi, rat Alpha or Mu, cytosolic classes) were purified by flow cytometry and used in colony-forming assays to show that GST confers cellular resistance to the carcinogen benzoblpyrene (*)-anti-diol epoxide (anti-BPDE). We developed a sorting technique to viably separate recombinant GST+ cells (20%) from the nonexpressing electroporated population (80%) on the basis of a GST-catalyzed intracellular conjugation of glutathione to the fluorescent labeling reagent monochlorobimane (mClB). The concentration of mClB, length of time cells are exposed to mClB, and activity of the expressed GST isozyme determined the degree to which recombinant GST+ cells fluoresced more intensely than controls. On-line reagent addition ensured that all cells were exposed to 25 p M mClB for 3035 s during transit before being analyzed for fluorescence intensity and sorted. The apparent K , for mClB of the endogenous COS cell GST-catalyzed intracellular reaction was 88 pM. Stained GST Ya+ or Yb,+ cells catalyzed the conjugation 2 or 5 times more effectively than GST n+ cells. Enzyme activity in cytosolic fractions prepared from sorted recombinant GST+ cells was 1.8 -+ 0.3-fold greater than that of the control (80 f 4 nmoYmin/mg protein). Upon a 5-fold purification of GST n+ cells in the electroporated population, resistance to anti-BPDE in colony-forming assays increased 5 times, from 1.1-fold (unsorted) to 1.5-fold (sorted) (P <0.001).Key terms: Resistance to carcinogens, reversion of transient expression, COS cells Glutathione S-transferases (GST,3 EC 2.5.1.18) are a family of isozymes that can catalyze the covalent addition of the tripeptide glutathione, present at 0.5-10 mM in mammalian cells (35), to a structurally diverse array of physiological and xenobiotic electrophiles (21,25,32,37,41,52,53). GSTs are ubiquitous in the animal and plant kingdoms (53), and Mannervik (32) has classified the mammalian cytosolic GSTs into three classes (Pi, Alpha, and Mu) based on structural, immunological, and enzymatic properties. Mammalian cytosolic GSTs constitute 1-10% of total cytosolic protein (21) and are composed of two subunits (23-28 kilodaltons each) (32) that dimerize by noncovalent interactions (51). As demonstrated with in vitro kinetic analyses, GSTs can conjugate glutathione to anticancer alkylating agents (2,3,9,49,59) [1606][1607][1608][1609][1610][1611][1612] 1991) and Ublacker et al. (Cancer Res 51: 1783-1788, 1991, based upon in vitro kinetic analyses, support our findings done in whole cells which show that the rat )I and a class GSTs catalyze the conjugation of GSH to monochlorobimane more efficiently than 71 class GSTs.

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Section: Discussionsupporting

confidence: 74%

Recombinant glutathione S‐transferase (GST) expressing cells purified by flow cytometry on the basis of a GST‐catalyzed intracellular conjugation of glutathione to monochlorobimane

et al. 1991

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“…The HEY ovarian cancer cell line was maintained in MEMα medium with 10% FCS (Buick et al, 1985). The remainder of the ovarian cancer cell lines -OAW 42 (Wilson, 1984), OAW 28+53 (Wilson et al, 1987), PEO1 and PEO4 (Wolf et al, 1987), PEO14 (Langdon et al, 1988), JAM (Ward et al, 1987), SKOV3 (Fogh and Trempe, 1975), COLO316 (Woods et al, 1979), CAOV3 , OVCAR-3 (Hamilton et al, 1983), and 27/87 (derived by T Hurst) -were all maintained in RPMI 1640 with 10% FCS. Cells were harvested for RNA and protein isolation at about 80% confluence.…”

Section: Cell Lines Ovarian Surface Epithelium Cultures and Primary mentioning

confidence: 99%

Decreased expression of the Id3 gene at 1p36.1 in ovarian adenocarcinomas

et al. 2001

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SummaryThe molecular events that drive the initiation and progression of ovarian adenocarcinoma are not well defined. We have investigated changes in gene expression in ovarian cancer cell lines compared to an immortalized human ovarian surface epithelial cell line (HOSE) using a cDNA array. We identified 17 genes that were under-expressed and 10 genes that were over-expressed in the cell lines compared to the HOSE cells. One of the genes under-expressed in the ovarian cancer cell lines, Id3, a transcriptional inactivator, was selected for further investigation. Id3 mRNA was expressed at reduced levels in 6 out of 9 ovarian cancer cell lines compared to the HOSE cells while at the protein level, all 7 ovarian cancer cell lines examined expressed the Id3 protein at greatly reduced levels. Expression of Id3 mRNA was also examined in primary ovarian tumours and was found in only 12/38 (32%) cases. A search was conducted for mutations of Id3 in primary ovarian cancers using single stranded conformation polymorphism (SSCP) analysis. Only one nucleotide substitution, present also in the corresponding constitutional DNA, was found in 94 ovarian tumours. Furthermore no association was found between LOH at 1p36 and lack of expression of Id3. These data suggest that Id3 is not the target of LOH at 1p36.

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“…For example, certain compounds which inhibit chemical carcinogenesis are often inducers of the GST in the target tissue (Benson et al, 1978;Benson & Barretto, 1985). These proteins are also overexpressed in preneoplastic lesions (Kitahara et al, 1984;Pickett et al, 1984; Buchmann et al, 1985) and, in addition, have been demonstrated to be increased in both normal and tumour cells exposed to cytotoxic drugs (Adams et al, 1985;Wang & Tew, 1985;Carmichael et al, 1986;Batist et al, 1986;Evans et al, 1987;Robson et al, 1987;Wolf et al, 1987a;.It is likely that glutathione S-transferase levels and isoenzyme composition will play a role in both the intrinsic and acquired resistance to cancer chemotherapy (McGowan & Fox, 1986;Wolf et al, 1987b;Buller et al, 1987;Lewis et al, 1988a). Surprisingly little is known about the GST isoenzyme content of cell lines and work on solid tumours is still limited.…”

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confidence: 99%

Glutathione S-transferase isoenzymes in human tumours and tumour derived cell lines

Lewis

Forrester²,

Hayes³

et al. 1989

Br J Cancer

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Edinburgh, Edinburgh Royal Infirmary, Edinburgh EH3 9 YW; and 3Department of Clinical Oncology, Newcastle General Hospital, Newcastle-upon-Tyne, UK.Summary An increasing body of evidence indicates that glutathione S-transferases play a role in the intrinsic and acquired resistance of tumours to anticancer drugs. In view of the wide use of tumour cell lines to understand the factors which confer either sensitivity or resistance to chemotherapeutic agents we have determined glutathione S-transferase (GST) activity and isozyme composition in nine human cell lines. These data have been compared with the values obtained in solid tumours. In most cases overall GST activity was higher in the tumours than in the cell lines. This was most pronounced for the breast tumour samples relative to MCF7 cell line. The pi class GST subunit was present at similar concentration in the cell lines and the tumours, and in most cases was the most abundant subunit present. The alpha and mu class GST were expressed in most of the cell lines but at much lower concentration than the pi class subunit. Also considerable variability particularly in the expression of the mu subunits was observed. This was also the case for the expression of these subunits in the solid tumour samples. The levels of these GSTs (when expressed) in the solid tumours was invariably higher than that observed in the cell lines. There are therefore several similarities but also some significant differences in GST expression in solid tumours and cell lines. Whether the differences are because expression is lost during the generation of the cell lines or whether it reflects the individuality of human tumours remains to be clearly established.The glutathione S-transferases (GST) are a multigene family of dimeric proteins which play a central role in the detoxification of electrophilic xenobiotics (Chasseaud, 1979). In man, cytosolic GSTs have been divided into three major classes termed alpha (basic), mu (neutral) and pi (acidic) (Mannervik, 1985;Stockman et al., 1987). Proteins within these groups have marked differences in their substrate specificity. There is a growing body of evidence which indicates that GST play an important role in both carcinogenesis and drug resistance. For example, certain compounds which inhibit chemical carcinogenesis are often inducers of the GST in the target tissue (Benson et al., 1978;Benson & Barretto, 1985). These proteins are also overexpressed in preneoplastic lesions (Kitahara et al., 1984;Pickett et al., 1984; Buchmann et al., 1985) and, in addition, have been demonstrated to be increased in both normal and tumour cells exposed to cytotoxic drugs (Adams et al., 1985;Wang & Tew, 1985;Carmichael et al., 1986;Batist et al., 1986;Evans et al., 1987;Robson et al., 1987;Wolf et al., 1987a;.It is likely that glutathione S-transferase levels and isoenzyme composition will play a role in both the intrinsic and acquired resistance to cancer chemotherapy (McGowan & Fox, 1986;Wolf et al., 1987b;Buller et al., 1987;Lewis et al., 1988a). Surpris...

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Cellular heterogeneity and drug resistance in two ovarian adenocarcinoma cell lines derived from a single patient

Cited by 128 publications

References 29 publications

Recombinant glutathione S‐transferase (GST) expressing cells purified by flow cytometry on the basis of a GST‐catalyzed intracellular conjugation of glutathione to monochlorobimane

Recombinant glutathione S‐transferase (GST) expressing cells purified by flow cytometry on the basis of a GST‐catalyzed intracellular conjugation of glutathione to monochlorobimane

Decreased expression of the Id3 gene at 1p36.1 in ovarian adenocarcinomas

Glutathione S-transferase isoenzymes in human tumours and tumour derived cell lines

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