Biomedical subjects

G D Frenkel

Publications and source records attributed to G D Frenkel.

At least 19 recordsLinked to original sources

PubMed2001

A prevention strategy for circumventing drug resistance in cancer chemotherapy.

The development of drug resistance is considered to be a major cause for the failure of chemotherapy in a number of types of cancer, including ovarian, breast and lung. Most previous research has focused on approaches to reverse drug resistance once it has arisen, that is, on the use of agents which can make drug-resistant tumors more sensitive to chemotherapy. Unfortunately, this approach has thus far met with only limited clinical success. Because of the prevalence of drug resistance in cases of advanced cancer, there exists an urgent need to develop new approaches to dealing with this problem. We have hypothesized the feasibility of an alternative approach: the use of specific agents to prevent the development of resistance before it arises. Our initial studies to examine this hypothesis have focused on ovarian cancer. We have designed both in vitro and in vivo systems in which resistance develops rapidly after exposure of tumor cells or xenografts to melphalan or cisplatin. Using these systems we have shown that two selenium compounds, selenite and selenomethionine are able to prevent the induction of resistance. Furthermore, inclusion of selenite in a chemotherapeutic protocol can result in a significant enhancement of the efficacy of cisplatin in suppressing the growth of human ovarian tumor xenografts. These results have supported the idea that prevention may be a useful new approach to the problem of drug resistance in cancer chemotherapy.

Animals↗

PubMed2000

Selenium compounds prevent the induction of drug resistance by cisplatin in human ovarian tumor xenografts in vivo.

PURPOSE: The development of drug resistance is a major cause for the failure of chemotherapy, particularly in ovarian cancer. Most previous research has focused on approaches to reverse drug resistance once it has arisen, that is, on the use of agents which can make drug-resistant tumors more sensitive to chemotherapy. We have suggested the feasibility of an alternative approach: the use of specific agents to prevent the development of resistance. METHODS: We designed an in vivo system to assay for the ability of compounds to prevent the induction of resistance by cisplatin. In this system, mice bearing tumors (which originated from A2780 human ovarian tumor cells) were treated with a low dose (2.6 mg/kg) of cisplatin and the tumors rapidly developed resistance to subsequent cisplatin treatment. Cell lines initiated from these tumors retained the resistant phenotype even after several months in culture. RESULTS: When either selenite or selenomethionine were administered (i.p., 1.5 mg/kg) close to the time of the initial cisplatin treatment, the induction of resistance was prevented. Similar treatments with sulfite or methionine had no effect on the induction of resistance by cisplatin. Studies in cells from treated tumors have indicated that the selenium compounds may prevent the induction of resistance by preventing a cisplatin-induced increase in glutathione level. CONCLUSIONS: Selenium compounds specifically prevent the induction by cisplatin of drug resistance in human ovarian tumors in vivo.

Animals↗

PubMed1999

Rapid development of glutathione-S-transferase-dependent drug resistance in vitro and its prevention by ethacrynic acid.

Exposure of A2780 human ovarian tumor cells to a low concentration of melphalan in vitro for 7 days resulted in the development of melphalan resistance. This resistance was not a stable characteristic of the cells since it was lost after 2 weeks in culture in the absence of drug. The melphalan-resistant cells exhibited significant cross-resistance to cisplatin but only minor cross-resistance to doxorubicin. The resistant cells had elevated levels of glutathione-S-transferase activity and mRNA. Exposure of the cells to the ethacrynic acid resulted in a decrease in enzyme activity as well as a reversal of their drug-resistant phenotype, indicating that the enzyme is involved in the resistance. When ethacrynic acid was present during the 7-day exposure of the cells to melphalan, the development of drug resistance was prevented. This system may serve as a useful preliminary step in screening for agents which can prevent the development of chemotherapy-induced drug resistance in human cancer.

Antineoplastic Agents, Alkylating↗

PubMed1998

Studies on the mechanism of the selenite-induced decrease in cell attachment: effect of selenite on the levels of fibronectin receptor (alpha5beta1 integrin) mRNAs.

We previously reported that exposure of HeLa cells to selenite for 2 h results in a decrease in their ability to attach to fibronectin (Yan and Frenkel, Cancer Res. 52, 5803-5807 [1992]), as well as a decrease in the level of fibronectin receptor (alpha5beta1 integrin) at the cell surface (Yan and Frenkel, Biol. Trace Element Res. 46, 79-89 [1994]). We have now found that after exposure to selenite, there was a decrease in the total cellular content of the receptor protein, as well as in the level of the mRNAs for both of the subunits. Exposure of cells to actinomycin D (an inhibitor of RNA synthesis) also resulted in a decrease in the level of these mRNAs, suggesting that the effect of selenite is the result of its known inhibitory effect on RNA synthesis (Frenkel, Toxicol. Lett. 25, 219-223 [1985]). Exposure of cells to actinomycin D for 2 h also resulted in a decrease in the ability of cells to attach to fibronectin. Furthermore, both selenite and actinomycin D caused a decrease in integrin mRNA levels and in cell attachment to fibronectin only when high-density cells were exposed to the agents. In contrast, when low-density cells were exposed,neither agent had any detectable effect on mRNA levels or on cell attachment. These results have suggested the following scheme for the mechanism of the inhibition of cell attachment by selenite: After exposure to selenite for 2 h, there is a significant inhibition of cellular RNA synthesis, which results in a general decrease in the cellular level of those mRNAs with relatively short half-lives, including in particular those of the fibronectin receptor. This leads to a decrease in the intracellular level of the receptor protein and, consequently, in its level at the cell surface, which in turn causes a decrease in the rate of cell attachment to fibronectin.

Blotting, Northern↗

PubMed1998

Prevention of the development of melphalan resistance in vitro by selenite.

Exposure of A2780 human ovarian tumor cells to a low concentration of melphalan in vitro for 7 d results in the development of melphalan resistance, which is dependent on elevated cellular levels of glutathione and glutathione S-transferase. The inclusion of selenite (at concentrations as low as 0.2 microM) during the exposure to melphalan completely prevented the development of resistance. Selenite did not prevent the melphalan-induced increase in glutathione, but it did prevent the increase in the activity of glutathione S-transferase. It also prevented the increase in the expression of the glutathione S-transferase gene, suggesting that this may be the mechanism by which it prevents the development of melphalan resistance. The results of this in vitro study suggest that selenite may prove to be useful in preventing the development of drug resistance in vivo.

Antineoplastic Agents, Alkylating↗

PubMed1997

Sensitivity of melphalan-resistant tumors to selenite in vivo.

Previous studies have demonstrated that melphalan-resistant human ovarian tumor cells exhibit a higher degree of sensitivity to the cytotoxic effects of selenite in vitro than comparable drug-sensitive cells (P.B. Caffrey, G.D. Frenkel, Selenite cytotoxicity in drug resistant and non-resistant human ovarian tumor cells, Cancer Res. 52 (1992) 4812-4816; P.B. Caffrey, G.D. Frenkel, The development of drug resistance by tumor cells in vitro is accompanied by the development of sensitivity to selenite, Cancer Lett. 81 (1994) 59-65). We have now examined the sensitivity of drug-resistant tumors to selenite in vivo. A2780 human ovarian tumor cells, or their melphalan-resistant derivative (A2780ME) cells were injected subcutaneously into nude mice and the resulting tumors were found to be melphalan-sensitive and -resistant, respectively, in vivo. Treatment with selenite (2 mg/kg Se s.c.), which had no overt toxic effect on the animals, resulted in a significant decrease in the rate of growth of the melphalan-resistant tumors, but not on the rate of growth of the drug-sensitive tumors. Thus, melphalan-resistant ovarian tumors are also more sensitive to selenite treatment in vivo.

Animals↗

PubMed1994

Time-course of inhibition of cellular nucleic acid synthesis by selenite.

The relationship between intracellular sulfhydryl(SH) compounds and the kinetics of the inhibitory effect of selenite on cellular nucleic acid synthesis has been examined. In A549 cells, with a relatively high SH level, exposure to low concentrations of selenite caused inhibition even after short exposure times. In contrast, in VA cells, with a relatively low level of SH compounds, selenite had no significant effect at short exposure times, but inhibited significantly with longer exposures. Selenodicysteine, the product of the reaction of selenite with cysteine (an important intracellular SH compound), inhibited synthesis in both cell types at short exposure times. Exposure of cells to diethylmaleate, which decreased the level of intracellular SH compounds, reduced the inhibitory effect of a short exposure to selenite but did not affect a long exposure. These results indicate that the reaction of selenite with intracellular SH compounds may be a determining factor in the kinetics of its inhibitory effect on cellular DNA and RNA synthesis.

Adenocarcinoma↗

PubMed1994

The development of drug resistance by tumor cells in vitro is accompanied by the development of sensitivity to selenite.

The effects of selenite on cell viability and proliferation in a line of drug-sensitive human ovarian tumor (A2780) cells were compared with its effects on a melphalan-resistant derivative of these cells (A2780-ME) which had been developed in vitro (Hamilton et al. (1985) Biochemical Pharmacol., 34, 2583-2586). With the A2780-ME cells there was a 50% decrease in the number of viable cells (i.e. which exclude Trypan Blue dye) after exposure to less than 100 microM selenite for 6 h. In contrast, exposure to more than 300 microM selenite was required to achieve the same effect in the parent line. Similarly, exposure to 10 microM selenite resulted in a 50% decrease in A2780-ME cell proliferation, whereas this treatment had only a small inhibitory effect on proliferation of the parent cells. Thus, the development of melphalan resistance in vitro was accompanied by the development of selenite sensitivity. Pre-exposure of the two cell types to buthionine sulfoximine eliminated the difference in their intracellular glutathione levels, as well as most of their differential sensitivity to selenite. Furthermore, the two cell types did not exhibit a difference in sensitivity to selenodiglutathione, the product of the reaction of selenite with glutathione. Thus, the increase in intracellular glutathione, which has been shown to be responsible for the development of drug resistance in these cells is also responsible for the development of selenite sensitivity.

Cell Division↗

PubMed1994

Effect of selenite on tumor cell invasiveness.

Pre-exposure of HeLa or NIH:OVCAR-3 cells to selenite resulted in a dose-dependent decrease in the ability of the cells to invade a layer of Matrigel, a reconstituted basement membrane preparation. In contrast, selenate, selenomethionine and sulfite had no significant effect on cell invasiveness. Exposure of HeLa cells to selenite also resulted in a decrease in two of the necessary steps of the invasion process, attachment and mobility; in contrast, exposure of OVCAR cells decreased attachment but not mobility. There was an apparent correlation between the processes that are affected by selenite and those that involve the cellular fibronectin receptor (alpha 5 beta 1 integrin).

3T3 Cells↗

PubMed1994

Protein synthesis is not required for the inhibitory effect of selenite on cell colony formation and RNA synthesis.

Selenite has been shown to undergo intracellular metabolism that results in its conversion to other low molecular weight Se-containing species and also to its incorporation into a selenocysteine residue in selenoprotein. In order to investigate whether the incorporation into protein is required for the cytotoxic effects of selenite, we have examined whether inhibition of protein synthesis prevents the inhibitory effect of selenite on the ability of cells to form colonies or to synthesize RNA. We have found that treatment of HeLa cells with cycloheximide inhibited protein synthesis by > 90% but had no effect on the inhibitory effect of selenite on cell colony formation or RNA synthesis. Since protein synthesis is not necessary for these cytotoxic effects of selenite they are unlikely to result from an increase in the synthesis of selenoproteins.

Clone Cells↗

PubMed1992

Inhibition of cell attachment by selenite.

Brief pre-exposure of HeLa cells to micromolar concentrations of selenite resulted in a dose-dependent decrease in the rate of their subsequent attachment to a solid matrix (tissue culture dish). Similar low concentrations of selenite also inhibited colony formation, but only when the cells were exposed prior to their attaching to the dish, not when they were exposed after attachment. This indicates that inhibition of cell proliferation by selenite requires exposure to higher concentrations for longer periods of time. In contrast, selenate, selenomethionine, selenocystine, and sulfite did not affect cell attachment, even at significantly higher concentrations. Thus, the inhibition of cell attachment is a specific effect of selenite. Selenite also inhibited the attachment of cells to bacteriological dishes coated with fibronectin, laminin, or collagen, proteins that are components of the extracellular matrix. There was no inhibition when the tissue culture dishes or the protein-coated dishes were pre-exposed to selenite. There was also no inhibition when the cells were exposed to selenite during the attachment process. Thus, pre-exposure of the cells to selenite was necessary for inhibition of attachment. Since cell attachment has been shown to be an important early step in tumor cell invasion and metastasis, these results suggest a novel mechanism of the anticarcinogenic effect of selenite: inhibition of the attachment of tumor cells to the extracellular matrix.

Anticarcinogenic Agents↗

PubMed1992

Selenite cytotoxicity in drug resistant and nonresistant human ovarian tumor cells.

Our previous studies on selenite cytotoxicity led us to hypothesize that drug resistant tumor cells with high intracellular glutathione will exhibit a high degree of sensitivity to selenite. To examine this we studied the effects of selenite on drug resistant human ovarian tumor (NIH:OVCAR-3) cells in three assays of cytotoxicity: proliferation; cell viability (trypan blue exclusion); and attachment to a solid matrix. The cells were sensitive to low levels of selenite: concentrations as low as 5 microM inhibited cell proliferation and attachment; and viability was decreased by concentrations as low as 20 microM. In each of these assays the NIH:OVCAR-3 cells were more sensitive to selenite than were drug sensitive human ovarian tumor (A2780) cells. These results suggest the potential for the utilization of selenite in the treatment of some drug resistant tumors.

Cell Adhesion↗

PubMed1992

Inhibition by selenium of DNA and RNA synthesis in normal and malignant human cells in vitro.

Several studies have demonstrated differences between normal and malignant cells in their sensitivity to various effects of selenite. We have compared the effect of selenite on DNA and RNA synthesis in two pairs of normal and malignant human cell lines. One pair of cells, CCL-210 (normal lung fibroblasts) and A549 (lung adenocarcinoma cells), exhibited a large difference in their sensitivity to selenite but no significant difference in their sensitivity to selenodiglutathione. They also had a large difference in the level of intracellular sulfhydryl (SH) compounds. In contrast the other pair of cells, WI-38 (normal fetal lung fibroblasts) and WI-38VA (SV-40 transformed WI-38 cells) both had low levels of intracellular SH compounds and exhibited similar (low) sensitivity to selenite. Our results indicate that differences between normal and malignant cells in their sensitivity to selenite could be due to a difference in the reaction of selenite with intracellular sulfhydryl compounds to form selenotrisulfides.

Cell Line↗

PubMed1992

Selenite-induced inhibition of colony formation by buthionine sulfoximine-sensitive and resistant cell lines.

We previously demonstrated that treatment of HeLa cells with buthionine sulfoximine (BSO), which decreases the level of cellular glutathione, resulted in a decrease in the potency of selenite in inhibiting cell colony formation. We have now examined the effect of selenite on normal human lung fibroblast (CCL-210) cells, which resemble HeLa cells in their sensitivity to BSO, and on human lung adenocarcinoma (A549) cells, which are relatively insensitive to BSO. We have found that BSO treatment caused an approximately fourfold decrease in selenite potency in the CCL-210 cells, but had no significant effect on its potency in A549 cells. These results support the hypothesis that for selenite to exert its cytotoxic effect, it must undergo the reaction with an SH compound to form the selenotrisulfide. As a result of the lower sensitivity of the tumor cells to BSO, it was possible to achieve a large differential sensitivity to the cytotoxic effect of selenite.

Antimetabolites, Antineoplastic↗

PubMed1992

The effect of saffron on intracellular DNA, RNA and protein synthesis in malignant and non-malignant human cells.

Extract of saffron (Crocus sativis) has previously been shown to inhibit colony formation and cellular DNA and RNA synthesis by HeLa cells in vitro. In order to compare the sensitivity of malignant and non-malignant cells to saffron, we examined the effect of the extract on macromolecular synthesis in three human cell lines: A549 cells (derived from a lung tumor), WI-38 cells (normal lung fibroblasts) and VA-13 cells (WI-38 cells transformed in vitro by SV40 tumor virus). We found that the malignant cells were more sensitive than the normal cells to the inhibitory effects of saffron on both DNA and RNA synthesis. There was no effect on protein synthesis in any of the cells.

Cell Line↗

PubMed1992

Effect of saffron on cell colony formation and cellular nucleic acid and protein synthesis.

A concentrated extract of saffron was prepared from the flowers of Crocus sativis. The effect of this extract on the ability of HeLa cells to form colonies, and on cellular DNA, RNA and protein synthesis was examined. Incubation of cells with extract for 3 h resulted in significant inhibition of colony formation and cellular nucleic acid synthesis with 50% inhibition at concentrations of approximately 100-150 micrograms/ml. In contrast there was no inhibition of cellular protein synthesis at concentrations of extract as high as 400 micrograms/ml.

Carotenoids↗

PubMed1991

Products of the reaction of selenite with intracellular sulfhydryl compounds.

The usual first step in the intracellular metabolism of exogenous selenite is its chemical reaction with glutathione to form selenodiglutathione (1). We have investigated whether selenite also reacts intracellularly with other SH compounds. HeLa cells were exposed to [75Se]selenite and lysed with SDS. Cellular proteins and nucleic acids were precipitated with trichloroacetic acid, and the acid-soluble fraction was analyzed by ion-exchange thin-layer chromatography (ion-exchange TLC) and autoradiography. In control cells, the major [75Se]-containing species detected can be identified by its mobility as selenodiglutathione. Two other species were detected, which can be identified as selenodimercaptoethylamine and the mixed selenotrisulfide of mercaptoethylamine and glutathione. In contrast, in cells that were depleted of glutathione (by treatment with buthionine sulfoximine), very little, if any, selenodiglutathione was detected. However, new [75Se]-containing species were detected, which can be identified as selenodicysteine and the mixed selenotrisulfide of cysteine and glutathione. The same species were detected when [75Se]selenite was added to the acid-soluble fraction of a cell extract (as opposed to living cells), confirming that these compounds can be formed by nonenzymatic reactions.

Antimetabolites↗

PubMed1991

Inhibition of cell colony formation by selenite: involvement of glutathione.

Selenium is an essential trace element that has been shown to have anticarcinogenic activity. One mechanism that has been proposed for this activity is a cytotoxic effect of selenium on tumor cells. As a means of assessing its cytotoxicity, we have examined the effect of selenite on tumor cell viability, using as an assay the ability of the cells to form colonies. We have found that brief exposure of HeLa cells to micromolar concentrations of selenite resulted in significant inhibition of colony formation, indicating that this is an assay for selenite cytotoxicity that is more sensitive than those that have been employed previously. In order to investigate the involvement of cellular glutathione in selenite cytotoxicity, we treated cells with buthionine sulfoximine (BSO) before selenite exposure. This treatment, which resulted in a 7-fold reduction in the level of intracellular glutathione, also caused a significant decrease in the inhibitory effect of selenite on colony formation. However, when cells were exposed to selenite that had previously been reacted with glutathione, the BSO-induced decrease in cytotoxicity was eliminated. In contrast, reaction of selenite with other sulfhydryl compounds, such as cysteine and mercaptoethylamine, did not restore its potency in BSO-treated cells. The simplest explanation for these results is that, for selenite to exert its inhibitory effect, it must react with intracellular glutathione to form the selenodiglutathione derivative.

Buthionine Sulfoximine↗