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Role of SLC5A8, a plasma membrane transporter and a tumor suppressor, in the antitumor activity of dichloroacetate.

Babu E, Ramachandran S, CoothanKandaswamy V, Elangovan S, Prasad PD, Ganapathy V, Thangaraju M - Oncogene (2011)

Bottom Line: Credible evidence exists for the antitumor activity of this compound, but high concentrations are needed for significant therapeutic effect.Here we show that SLC5A8 transports DCA very effectively with high affinity.The mechanism of the compound's antitumor activity still remains its ability to inhibit pyruvate dehydrogenase kinase and force mitochondrial oxidation of pyruvate.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Medical College of Georgia, Georgia Health Sciences University, Augusta, GA 30912, USA.

ABSTRACT
There has been growing interest among the public and scientists in dichloroacetate (DCA) as a potential anticancer drug. Credible evidence exists for the antitumor activity of this compound, but high concentrations are needed for significant therapeutic effect. Unfortunately, these high concentrations produce detrimental side effects involving the nervous system, thereby precluding its use for cancer treatment. The mechanistic basis of the compound's antitumor activity is its ability to activate the pyruvate dehydrogenase complex through inhibition of pyruvate dehydrogenase kinase. As the compound inhibits the kinase at micromolar concentrations, it is not known why therapeutically prohibitive high doses are needed for suppression of tumor growth. We hypothesized that lack of effective mechanisms for the entry of DCA into tumor cells may underlie this phenomenon. Here we show that SLC5A8 transports DCA very effectively with high affinity. This transporter is expressed in normal cells, but expression is silenced in tumor cells by epigenetic mechanisms. The lack of the transporter makes tumor cells resistant to the antitumor activity of DCA. However, if the transporter is expressed in tumor cells ectopically, the cells become sensitive to the drug at low concentrations. This is evident in breast cancer cells, colon cancer cells and prostate cancer cells. Normal cells, which constitutively express the transporter, are however not affected by the compound, indicating tumor cell-selective therapeutic activity. The mechanism of the compound's antitumor activity still remains its ability to inhibit pyruvate dehydrogenase kinase and force mitochondrial oxidation of pyruvate. As silencing of SLC5A8 in tumors involves DNA methylation and its expression can be induced by treatment with DNA methylation inhibitors, our findings suggest that combining DCA with a DNA methylation inhibitor would offer a means to reduce the doses of DCA to avoid detrimental effects associated with high doses but without compromising antitumor activity.

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Differential effect of dichloroacetate (DCA) on intracellular levels of pyruvate in colon cancer cells and its dependence on SLC5A8Three human colon cancer cell lines (HT29, SW620, and HCT116) were transfected with either vector alone or SLC5A8 cDNA, and then cultured in the absence or presence of acetate, monocholoroacetate (MCA), dichloroacetate (DCA), or trichloroacetate (TCA) for 48 h. Concentration of the fatty acids was 1 mM. Cells were then lysed and used for determination of pyruvate and protein. Data for pyruvate were normalized with protein levels. (A) HT29; (B) SW620; (C) HCT116. *, p < 0.05, **, p < 0.01 and ***, p < 0.001.
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Figure 6: Differential effect of dichloroacetate (DCA) on intracellular levels of pyruvate in colon cancer cells and its dependence on SLC5A8Three human colon cancer cell lines (HT29, SW620, and HCT116) were transfected with either vector alone or SLC5A8 cDNA, and then cultured in the absence or presence of acetate, monocholoroacetate (MCA), dichloroacetate (DCA), or trichloroacetate (TCA) for 48 h. Concentration of the fatty acids was 1 mM. Cells were then lysed and used for determination of pyruvate and protein. Data for pyruvate were normalized with protein levels. (A) HT29; (B) SW620; (C) HCT116. *, p < 0.05, **, p < 0.01 and ***, p < 0.001.

Mentions: What is the mechanism responsible for the induction of apoptosis by SLC5A8/dichloroacetate in cancer cells? SLC5A8 is an active transporter for dichloroacetate with a Kt of 36 ± 7μM. Therefore, the compound will be concentrated in cancer cells if the transporter is expressed. This would explain the obligatory requirement for SLC5A8 for low concentrations of dichloroacetate to induce apoptosis in these cells. If the transporter is not expressed, cells may not accumulate the compound to levels sufficient enough to cause apoptosis. Once accumulated inside the cancer cells, how does dichloroacetate act to induce apoptosis? The only known mechanism of action of dichloroacetate is its ability to activate PDC through inhibition of PDK (Stacpoole et al., 1998, 2003, 2008). This would result in stimulation of mitochondrial oxidation, generation of reactive oxygen species, depolarization of mitochondrial membrane potential, and induction of apoptosis. Activation of PDC by dichloroacetate in intact cells would decrease intracellular levels of pyruvate. Based on this, we hypothesized that dichloroacetate would decrease pyruvate levels in cancer cells and that the effect would be dependent obligatorily on SLC5A8 expression. We tested this hypothesis in three different cancer cell lines (HT29, SW620, and HCT116) with and without exogenous expression of SLC5A8 (Fig. 6). The expression of the transporter by itself did not have any effect on intracellular levels of pyruvate. Dichloroacetate was able to decrease the levels of pyruvate to a significant extent in all three cancer cell lines, but only when the transporter was expressed. Interestingly, monochloroacetate also had a similar effect, again only in the presence of SLC5A8; in contrast, trichloroacetate and acetate were not able to affect pyruvate levels.


Role of SLC5A8, a plasma membrane transporter and a tumor suppressor, in the antitumor activity of dichloroacetate.

Babu E, Ramachandran S, CoothanKandaswamy V, Elangovan S, Prasad PD, Ganapathy V, Thangaraju M - Oncogene (2011)

Differential effect of dichloroacetate (DCA) on intracellular levels of pyruvate in colon cancer cells and its dependence on SLC5A8Three human colon cancer cell lines (HT29, SW620, and HCT116) were transfected with either vector alone or SLC5A8 cDNA, and then cultured in the absence or presence of acetate, monocholoroacetate (MCA), dichloroacetate (DCA), or trichloroacetate (TCA) for 48 h. Concentration of the fatty acids was 1 mM. Cells were then lysed and used for determination of pyruvate and protein. Data for pyruvate were normalized with protein levels. (A) HT29; (B) SW620; (C) HCT116. *, p < 0.05, **, p < 0.01 and ***, p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3140604&req=5

Figure 6: Differential effect of dichloroacetate (DCA) on intracellular levels of pyruvate in colon cancer cells and its dependence on SLC5A8Three human colon cancer cell lines (HT29, SW620, and HCT116) were transfected with either vector alone or SLC5A8 cDNA, and then cultured in the absence or presence of acetate, monocholoroacetate (MCA), dichloroacetate (DCA), or trichloroacetate (TCA) for 48 h. Concentration of the fatty acids was 1 mM. Cells were then lysed and used for determination of pyruvate and protein. Data for pyruvate were normalized with protein levels. (A) HT29; (B) SW620; (C) HCT116. *, p < 0.05, **, p < 0.01 and ***, p < 0.001.
Mentions: What is the mechanism responsible for the induction of apoptosis by SLC5A8/dichloroacetate in cancer cells? SLC5A8 is an active transporter for dichloroacetate with a Kt of 36 ± 7μM. Therefore, the compound will be concentrated in cancer cells if the transporter is expressed. This would explain the obligatory requirement for SLC5A8 for low concentrations of dichloroacetate to induce apoptosis in these cells. If the transporter is not expressed, cells may not accumulate the compound to levels sufficient enough to cause apoptosis. Once accumulated inside the cancer cells, how does dichloroacetate act to induce apoptosis? The only known mechanism of action of dichloroacetate is its ability to activate PDC through inhibition of PDK (Stacpoole et al., 1998, 2003, 2008). This would result in stimulation of mitochondrial oxidation, generation of reactive oxygen species, depolarization of mitochondrial membrane potential, and induction of apoptosis. Activation of PDC by dichloroacetate in intact cells would decrease intracellular levels of pyruvate. Based on this, we hypothesized that dichloroacetate would decrease pyruvate levels in cancer cells and that the effect would be dependent obligatorily on SLC5A8 expression. We tested this hypothesis in three different cancer cell lines (HT29, SW620, and HCT116) with and without exogenous expression of SLC5A8 (Fig. 6). The expression of the transporter by itself did not have any effect on intracellular levels of pyruvate. Dichloroacetate was able to decrease the levels of pyruvate to a significant extent in all three cancer cell lines, but only when the transporter was expressed. Interestingly, monochloroacetate also had a similar effect, again only in the presence of SLC5A8; in contrast, trichloroacetate and acetate were not able to affect pyruvate levels.

Bottom Line: Credible evidence exists for the antitumor activity of this compound, but high concentrations are needed for significant therapeutic effect.Here we show that SLC5A8 transports DCA very effectively with high affinity.The mechanism of the compound's antitumor activity still remains its ability to inhibit pyruvate dehydrogenase kinase and force mitochondrial oxidation of pyruvate.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Medical College of Georgia, Georgia Health Sciences University, Augusta, GA 30912, USA.

ABSTRACT
There has been growing interest among the public and scientists in dichloroacetate (DCA) as a potential anticancer drug. Credible evidence exists for the antitumor activity of this compound, but high concentrations are needed for significant therapeutic effect. Unfortunately, these high concentrations produce detrimental side effects involving the nervous system, thereby precluding its use for cancer treatment. The mechanistic basis of the compound's antitumor activity is its ability to activate the pyruvate dehydrogenase complex through inhibition of pyruvate dehydrogenase kinase. As the compound inhibits the kinase at micromolar concentrations, it is not known why therapeutically prohibitive high doses are needed for suppression of tumor growth. We hypothesized that lack of effective mechanisms for the entry of DCA into tumor cells may underlie this phenomenon. Here we show that SLC5A8 transports DCA very effectively with high affinity. This transporter is expressed in normal cells, but expression is silenced in tumor cells by epigenetic mechanisms. The lack of the transporter makes tumor cells resistant to the antitumor activity of DCA. However, if the transporter is expressed in tumor cells ectopically, the cells become sensitive to the drug at low concentrations. This is evident in breast cancer cells, colon cancer cells and prostate cancer cells. Normal cells, which constitutively express the transporter, are however not affected by the compound, indicating tumor cell-selective therapeutic activity. The mechanism of the compound's antitumor activity still remains its ability to inhibit pyruvate dehydrogenase kinase and force mitochondrial oxidation of pyruvate. As silencing of SLC5A8 in tumors involves DNA methylation and its expression can be induced by treatment with DNA methylation inhibitors, our findings suggest that combining DCA with a DNA methylation inhibitor would offer a means to reduce the doses of DCA to avoid detrimental effects associated with high doses but without compromising antitumor activity.

Show MeSH
Related in: MedlinePlus