Previously, we reported that the nucleoside analog/transcriptional inhibitor ARC was able to indu... more Previously, we reported that the nucleoside analog/transcriptional inhibitor ARC was able to induce p53-independent apoptosis in multiple cancer cell lines of different origin. This occurred at least in part by the suppression of Mcl-1 expression, which a short-lived, pro-survival member of the Bcl-2 family. Interestingly, pan-caspase inibitor Z-VAD-FMK protected leukemia cells from ARC-induced mitochondrial damage and apoptosis, and Mcl-1 mRNA and protein from down-regulation by ARC. At the same time treatment of human cancer cells with the pan-Bcl-2 inhibitor ABT-737 alone led to up-regulation of Mcl-1 protein expression. Combination of sub-apoptotic concentrations of ABT-737 and ARC suppressed Mcl-1 expression, induced mitochondrial injury and potent apoptosis in wide variety of human cancer cell lines. In prostate cancer cells co-treatment with ARC and ABT-737 resulted in synergistic induction of cell death with combination index 0.7-0.8. These data suggest that ABT-737/ARC combination that simultaneously targets Bcl-2 and Mcl-1 may be efficient against human cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1036.
Abllbract-Fourteen species representing nine genera of the Hydrangeeceee DumorlJer were surveyed ... more Abllbract-Fourteen species representing nine genera of the Hydrangeeceee DumorlJer were surveyed for their fievonoid pig ° ments. All tsxs exhibited profiles based upon common fiavonols. Myricetin was seen in two genera: Jamesia and Decumaria. Jamesia was further distinguished by the absence of keempferol or its glycosides. A complex array of 3-O-mono-, 3-O-di-end 3-O-triglycosides was observed, although not ell species had ell levels of glycosyletion. Decumaria barbara was unique within the species studied in its possession of 3,7-di-end 3,7-triglycosides. The overall pattern of flavonol glycosides observed for the Hydrangeaceee closely resembles that found in herbaceous genera of Saxifrageceee. The comparatively low frequency of myricetin contrasts with its high occurrence in herbaceous genera.
Background-A close association between periodontitis and diabetes has been demonstrated in human ... more Background-A close association between periodontitis and diabetes has been demonstrated in human cross-sectional studies, but an exact relationship between periodontitis and prediabetes has not been established. Previous studies using animal model systems consistently have shown that hyperinsulinemia occurs in animals with periodontitis compared to animals with healthy periodontium (while maintaining normoglycemia). Because bacterial lipopolysaccharide (LPS) plays an important role in the pathogenesis of periodontitis, we hypothesized that LPS may stimulate insulin secretion through a direct effect on β cell function. To test this hypothesis, pancreatic β cell line MIN6 cells were used to determine the effect of Porphyromonas gingivalis (Pg) LPS on insulin secretion. Furthermore, expression of genes altered by Pg LPS in innate immunity and insulin-signaling pathways was determined. Methods-MIN6 cells were grown in medium with glucose concentration of normoglycemia (5.5 mM). Pg LPS was added to each well at final concentrations of 50, 200, and 500 ng/mL. Insulin secretion was measured using enzyme-linked immunosorbent assay. Gene expression levels altered by Pg LPS were determined by polymerase chain reaction (PCR) array for mouse innate and adaptive immunity response and mouse insulin-signaling pathways, and results were confirmed for specific genes of interest by quantitative PCR. Results-Pg LPS stimulated insulin secretion in the normoglycemic condition by ≈1.5to 3.0fold depending on the concentration of LPS. Pg LPS treatment altered the expression of several genes involved in innate and adaptive immune response and insulin-signaling pathway. Pg LPS upregulated the expression of the immune response-related genes cluster of differentiation 8a (Cd8a), Cd14, and intercellular adhesion molecule-1 (Icam1) by about twofold. LPS also increased the expression of two insulin signaling-related genes, glucose-6-phosphatase catalytic subunit (G6pc) and insulin-like 3 (Insl3), by three-to four-fold.
DNA topoisomerases are critical enzymes involved in replication, transcription, chromatin assembl... more DNA topoisomerases are critical enzymes involved in replication, transcription, chromatin assembly and other aspects of DNA metabolism. They are also the targets of important anticancer drugs. The type II topoisomerases are specific targets of drug classes that comprise complex-stabilizing (epipodophyllotoxins, anthracyclines) and catalytic (merbarone, bisdioxopiperazines) inhibitors. In this review, we update our current knowledge of resistance to the antitumor inhibitors of the type II DNA topoisomerases, with special emphasis on the catalytic inhibitors, since novel catalytic inhibitor resistant cell lines have only recently been described. Resistance to topoisomerase II inhibitors can manifest as decreased or increased expression of or mutation in the topoisomerase II genes. However, the tumor cell's response to exposure to these inhibitors involves more than the target enzyme, and these other responses are a major focus of this review. Such cellular changes are associated with and may contribute to the drug resistance phenotype. They involve decreased drug accumulation due to expression of membrane 'pump' proteins, altered cytotoxic signaling through stress-activated protein kinases, and alterations in apoptosis and cell cycle proteins (e.g. Bcl-2, Bax, p53, Rb). While it is evident that mutation in or altered expression of the topoisomerase II genes are sufficient to confer resistance to topoisomerase inhibitors, it is not clear whether the other changes are a consequence of the selection or a response to the cytotoxic insult, nor is it clear how these other cellular changes contribute to the drug resistance phenotype. Copyright 1999 Harcourt Publishers Ltd.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, Nov 1, 1992
A new system is described to determine the mutational spectra of mutagens and carcinogens in Esch... more A new system is described to determine the mutational spectra of mutagens and carcinogens in Escherichia coil; data on a limited number (142) of spontaneous mutants is presented. The mutational assay employs a method to select (rather than screen) for mutations in a supF target gene carried on a plasmid. The E. coli host cells (ES87) are lacl-(am26), and carry the IocZAMI5 marker for a-complementation in /3-galactosidase. When these cells also carry a plasmid, such as pUB3, which contains a wild-type copy of supF and lacZ-a, the lactose operon is repressed (off). Furthermore, supF suppression of lacl "m26 results in a lactose repressor that has an uninducible, lacl s genotype, which makes the cells unable to grow on lactose minimal plates. In contrast, spontaneous or mutagen-induced supFmutations in pUB3 prevent suppression of lacl '."2~' and result in constitutive expression of the lactose operon, which permits growth on lactose minimal plates. The spontaneous mutation frequency in the supF gene is ~ 0.7 and ~ 1.0 x 10-~ without and with SOS induction, respectively. Spontaneous mutations are dominated by large insertions (67% in SOS-uninduced and 56% in SOS-induced cells), and their frequency of appearance is largely unaffected by SOS induction. These are identified by DNA sequencing to be Insertion Elements; IS1 dominates, but IS4, ISS, gamma-delta and ISl0 are also obtained. Large deletions also contribute significantly (19% and 15% for-SOS and +SOS, respectively), where a specific deletion between a 10 base pair direct repeat dominates; the frequency of appearance of these mutations also appears to be unaffected by SOS induction, in contrast, SOS induction increases base pairing mutations (13% and 27% for-SOS and + SOS, respectively). The ES87/pUB3 system has many advantages for determining mutational spectra, including the fact that mutant isolation is fast and simple, and the determination of mutational changes is rapid because of the small size of supF.
Forkhead box M1 (FoxM1) oncogenic transcription factor represents an attractive therapeutic targe... more Forkhead box M1 (FoxM1) oncogenic transcription factor represents an attractive therapeutic target in the fight against cancer, because it is overexpressed in a majority of human tumors. Recently, using a cell-based assay system we identified thiazole antibiotic Siomycin A as an inhibitor of FoxM1 transcriptional activity. Here, we report that structurally similar thiazole antibiotic, thiostrepton also inhibits the transcriptional activity of FoxM1. Furthermore, we found that these thiopeptides did not inhibit the transcriptional activity of other members of the Forkhead family or some non-related transcription factors. Further experiments revealed that thiazole antibiotics also inhibit FoxM1 expression, but not the expression of other members of the Forkhead box family. In addition, we found that the thiazole antibiotics efficiently inhibited the growth and induced potent apoptosis in human cancer cell lines of different origin. Thiopeptide-induced apoptosis correlated with the suppression of FoxM1 expression, while overexpression of FoxM1 partially protected cancer cells from the thiazole antibiotic-mediated cell death. These data suggest that Siomycin A and thiostrepton may specifically target FoxM1 to induce apoptosis in cancer cells and FoxM1 inhibitors/thiazole antibiotics could be potentially developed as novel anticancer drugs against human neoplasia.
Supplemental data Fig.1. Combination treatment of ARC and ABT-737 is synergistic in inducing mito... more Supplemental data Fig.1. Combination treatment of ARC and ABT-737 is synergistic in inducing mitochondrial injury in tumor cells. A. DM833 cells were treated with ARC, ABT-737 or both as indicated, stained with TMRE and subjected to flow cytometric analysis. Mean fluorescence of cells due to bound TMRE is shown. B. U2OS-C3 cells treated with the drugs for 24 hrs were stained with TMRE and analyzed by flow cytometry. Mean fluorescence of cells due to bound TMRE is shown. Fig.2. Combination treatment with ARC and ABT-737 does not affect Bcl-2 and Bax levels. A. DM366 melanoma cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin. B. HepG2 liver cancer cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin. C. SW480 colon cancer cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin.
Bergenia crassifolia accumulates 3-O-monoglycosides and 3-00diglycosides of kaempferol and querce... more Bergenia crassifolia accumulates 3-O-monoglycosides and 3-00diglycosides of kaempferol and quercetin and lacks both myricetin derivatives and triglycosides. This is one of the simpler patterns of flavonoids observed in the Saxifragaceae. Francoa sonchifolia also produces a very simple array of flavonoids. Myricetin is lacking but a 3-00monoglycoside, two 3-O-diglycosides and a 3-O-triglycoside were observed. This taxon is distinguishable from most others in the family by the presence of 3-0 ° methylquercetin 7-00glucoside. The flavonoid profile of Parnassia is markedly more complex than any other genus of the Saxifragaceae studied to date. Its profile comprises flavonol 3-O-monoglycosides and 3-O-diglycosides but is unique in possessing large numbers of flavonol 3,7-diglycosides and 3,7-triglycosides. Lepuropetalon spatulum resembles Parnassia in flavonoid pattern more than it resembles any other genus of the Saxifragaceae. The combination of flavonoid data with published information on a variety of other characters leads us to agree with the position of Bergenia as a member of Saxifragaceae in the restricted sense. The flavonoids of Francoa, Parnassia and Lepuropetalon agree with suggestions that these genera stand apart from the herbaceous Saxifragaceae.
The oncogenic transcription factor forkhead box M1 (FoxM1) is overexpressed in a number of differ... more The oncogenic transcription factor forkhead box M1 (FoxM1) is overexpressed in a number of different carcinomas, whereas its expression is turned off in terminally differentiated cells. For this reason, FoxM1 is an attractive target for therapeutic intervention in cancer treatment. As a first step toward realizing this goal, in this study, using a high-throughput, cell-based assay system, we screened for and isolated the antibiotic thiazole compound Siomycin A as an inhibitor of FoxM1. Interestingly, we observed that Siomycin A was able to down-regulate the transcriptional activity as well as the protein and mRNA abundance of FoxM1. Consequently, we found that the downstream target genes of FoxM1, such as Cdc25B, Survivin, and CENPB, were repressed. Also, we observed that consistent with earlier reports of FoxM1 inhibition, Siomycin A was able to reduce anchorage-independent growth of cells in soft agar. Furthermore, we found that Siomycin A was able to induce apoptosis selectively in transformed but not normal cells of the same origin. Taken together, our data suggest that FoxM1 inhibitor Siomycin A could represent a useful starting point for the development of anticancer therapeutics. (Cancer Res 2006; 66(19): 9731-5)
Recently, we identified a nucleoside analog named ARC (4-amino-6-hydrazino-7-beta-D-ribofuranosyl... more Recently, we identified a nucleoside analog named ARC (4-amino-6-hydrazino-7-beta-D-ribofuranosyl-7H-Pyrrolo[2,3-d]pyrimidine-5-carboxamide), which has the properties of a general transcriptional inhibitor. Here, we report the characterization of ARC on a panel of colorectal cancer (CRC) cell lines. Cell death induced by ARC in CRC cells was accompanied by caspase-3 cleavage and correlated with the downregulation of antiapoptotic proteins, survivin and Mcl-1 and with the inhibition of Akt phosphorylation. At the same time, colon cancer cell lines were resistant to the well-known nucleoside analog DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole), which failed to downregulate Mcl-1 or survivin. Overall, ARC could represent an attractive candidate for anti-cancer drug development that targets multiple survival pathways in colon cancer cells.
We previously described the identification of a nucleoside analog transcriptional inhibitor ARC (... more We previously described the identification of a nucleoside analog transcriptional inhibitor ARC (4-amino-6-hydrazino-7-beta-Dribofuranosyl-7H-Pyrrolo[2,3-d]-pyrimidine-5-carboxamide) and FoxM1 inhibitor, thiazole antibiotic Siomycin A that were able to induce apoptosis in cancer cell lines of different origin. Here, we report the characterization of these drugs on a panel of melanoma cell lines. We found that in contrast to the common anti-melanoma drug dacarbazine (DTIC), ARC and thiazole antibiotics, Siomycin A and thiostrepton, efficiently inhibited growth and induced cell death in melanoma cell lines in low concentrations. Overexpression of the antiapoptotic protein Mcl-1 protected melanoma cells from apoptosis induced by these compounds. Furthermore, we found that ARC and Siomycin A synergistically induce apoptosis in DM833 melanoma cell line suggesting that they may antagonize different anti-apoptotic pathways in melanoma cells. In general, these drugs may represent important candidates for anti-cancer drug development against melanoma.
PDF file - 13093K, Supplementary Fig.1: Loss of DDB2 expression in human colon cancer. Supplement... more PDF file - 13093K, Supplementary Fig.1: Loss of DDB2 expression in human colon cancer. Supplementary Fig.2: Low magnification IHC data showing reduced expression of DDB2 in high grade colon cancer samples. Supplementary Fig.3: Depletion of DDB2 increases invasivness of colon cancer cells. Supplementary Fig.4: Polyclonal DDB2 knockdown colon cancer cells exhibit increased tumorigenicity Supplementary Fig.5: Polyclonal DDB2 knockdown colon cancer cells exhibit EMT Supplementary Fig.6: DDB2 knockdown colon cancer cells generate aggressive tumors in nude mice Supplementary Fig.7: Low magnification picture showing EMT in xenografted tumors Supplementary Fig.8: DDB2 expression blocks EMT induced by hypoxia and TGF-beta Supplementary Fig.9: Chromatin-IP using T7-antibody demonstrating interaction of T7-DDB2 with the VEGF, Snail and Zeb1 promoters. Supplementary Fig.10: Transcription factor binding sites in the promoter fragments bound by DDB2 Supplementary Fig.11: Western blot analyses for the DDB2-associated proteins and XPC in DDB2-depleted cells. Supplementary Fig.12: Experimental metastasis assay with polyclonal DDB2 knockdown cells. Supplementary Fig.13: Expression of DDB2 does not inhibit proliferation in SW620 cells.
Colon cancer is one of the deadliest cancers worldwide because of its metastasis to other essenti... more Colon cancer is one of the deadliest cancers worldwide because of its metastasis to other essential organs. Metastasis of colon cancer involves a complex set of events, including epithelial-to-mesenchymal transition (EMT) that increases invasiveness of the tumor cells. Here, we show that the xeroderma pigmentosum group E (XPE) gene product, damaged DNA-binding protein (DDB)-2, is downregulated in high-grade colon cancers, and it plays a dominant role in the suppression of EMT of the colon cancer cells. Depletion of DDB2 promotes mesenchymal phenotype, whereas expression of DDB2 promotes epithelial phenotype. DDB2 constitutively represses genes that are the key activators of EMT, indicating that DDB2 is a master regulator of EMT of the colon cancer cells. Moreover, we observed evidence that DDB2 functions as a barrier for EMT induced by hypoxia and TGF-b. Also, we provide evidence that DDB2 inhibits metastasis of colon cancer. The results presented here identify a transcriptional regulatory pathway of DDB2 that is directly linked to the mechanisms that suppress metastasis of colon cancer. Cancer Res; 73(12); 3771-82. Ó2013 AACR.
Previously, we reported that the nucleoside analog/transcriptional inhibitor ARC was able to indu... more Previously, we reported that the nucleoside analog/transcriptional inhibitor ARC was able to induce p53-independent apoptosis in multiple cancer cell lines of different origin. This occurred at least in part by the suppression of Mcl-1 expression, which a short-lived, pro-survival member of the Bcl-2 family. Interestingly, pan-caspase inibitor Z-VAD-FMK protected leukemia cells from ARC-induced mitochondrial damage and apoptosis, and Mcl-1 mRNA and protein from down-regulation by ARC. At the same time treatment of human cancer cells with the pan-Bcl-2 inhibitor ABT-737 alone led to up-regulation of Mcl-1 protein expression. Combination of sub-apoptotic concentrations of ABT-737 and ARC suppressed Mcl-1 expression, induced mitochondrial injury and potent apoptosis in wide variety of human cancer cell lines. In prostate cancer cells co-treatment with ARC and ABT-737 resulted in synergistic induction of cell death with combination index 0.7-0.8. These data suggest that ABT-737/ARC combination that simultaneously targets Bcl-2 and Mcl-1 may be efficient against human cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1036.
Abllbract-Fourteen species representing nine genera of the Hydrangeeceee DumorlJer were surveyed ... more Abllbract-Fourteen species representing nine genera of the Hydrangeeceee DumorlJer were surveyed for their fievonoid pig ° ments. All tsxs exhibited profiles based upon common fiavonols. Myricetin was seen in two genera: Jamesia and Decumaria. Jamesia was further distinguished by the absence of keempferol or its glycosides. A complex array of 3-O-mono-, 3-O-di-end 3-O-triglycosides was observed, although not ell species had ell levels of glycosyletion. Decumaria barbara was unique within the species studied in its possession of 3,7-di-end 3,7-triglycosides. The overall pattern of flavonol glycosides observed for the Hydrangeaceee closely resembles that found in herbaceous genera of Saxifrageceee. The comparatively low frequency of myricetin contrasts with its high occurrence in herbaceous genera.
Background-A close association between periodontitis and diabetes has been demonstrated in human ... more Background-A close association between periodontitis and diabetes has been demonstrated in human cross-sectional studies, but an exact relationship between periodontitis and prediabetes has not been established. Previous studies using animal model systems consistently have shown that hyperinsulinemia occurs in animals with periodontitis compared to animals with healthy periodontium (while maintaining normoglycemia). Because bacterial lipopolysaccharide (LPS) plays an important role in the pathogenesis of periodontitis, we hypothesized that LPS may stimulate insulin secretion through a direct effect on β cell function. To test this hypothesis, pancreatic β cell line MIN6 cells were used to determine the effect of Porphyromonas gingivalis (Pg) LPS on insulin secretion. Furthermore, expression of genes altered by Pg LPS in innate immunity and insulin-signaling pathways was determined. Methods-MIN6 cells were grown in medium with glucose concentration of normoglycemia (5.5 mM). Pg LPS was added to each well at final concentrations of 50, 200, and 500 ng/mL. Insulin secretion was measured using enzyme-linked immunosorbent assay. Gene expression levels altered by Pg LPS were determined by polymerase chain reaction (PCR) array for mouse innate and adaptive immunity response and mouse insulin-signaling pathways, and results were confirmed for specific genes of interest by quantitative PCR. Results-Pg LPS stimulated insulin secretion in the normoglycemic condition by ≈1.5to 3.0fold depending on the concentration of LPS. Pg LPS treatment altered the expression of several genes involved in innate and adaptive immune response and insulin-signaling pathway. Pg LPS upregulated the expression of the immune response-related genes cluster of differentiation 8a (Cd8a), Cd14, and intercellular adhesion molecule-1 (Icam1) by about twofold. LPS also increased the expression of two insulin signaling-related genes, glucose-6-phosphatase catalytic subunit (G6pc) and insulin-like 3 (Insl3), by three-to four-fold.
DNA topoisomerases are critical enzymes involved in replication, transcription, chromatin assembl... more DNA topoisomerases are critical enzymes involved in replication, transcription, chromatin assembly and other aspects of DNA metabolism. They are also the targets of important anticancer drugs. The type II topoisomerases are specific targets of drug classes that comprise complex-stabilizing (epipodophyllotoxins, anthracyclines) and catalytic (merbarone, bisdioxopiperazines) inhibitors. In this review, we update our current knowledge of resistance to the antitumor inhibitors of the type II DNA topoisomerases, with special emphasis on the catalytic inhibitors, since novel catalytic inhibitor resistant cell lines have only recently been described. Resistance to topoisomerase II inhibitors can manifest as decreased or increased expression of or mutation in the topoisomerase II genes. However, the tumor cell's response to exposure to these inhibitors involves more than the target enzyme, and these other responses are a major focus of this review. Such cellular changes are associated with and may contribute to the drug resistance phenotype. They involve decreased drug accumulation due to expression of membrane 'pump' proteins, altered cytotoxic signaling through stress-activated protein kinases, and alterations in apoptosis and cell cycle proteins (e.g. Bcl-2, Bax, p53, Rb). While it is evident that mutation in or altered expression of the topoisomerase II genes are sufficient to confer resistance to topoisomerase inhibitors, it is not clear whether the other changes are a consequence of the selection or a response to the cytotoxic insult, nor is it clear how these other cellular changes contribute to the drug resistance phenotype. Copyright 1999 Harcourt Publishers Ltd.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, Nov 1, 1992
A new system is described to determine the mutational spectra of mutagens and carcinogens in Esch... more A new system is described to determine the mutational spectra of mutagens and carcinogens in Escherichia coil; data on a limited number (142) of spontaneous mutants is presented. The mutational assay employs a method to select (rather than screen) for mutations in a supF target gene carried on a plasmid. The E. coli host cells (ES87) are lacl-(am26), and carry the IocZAMI5 marker for a-complementation in /3-galactosidase. When these cells also carry a plasmid, such as pUB3, which contains a wild-type copy of supF and lacZ-a, the lactose operon is repressed (off). Furthermore, supF suppression of lacl "m26 results in a lactose repressor that has an uninducible, lacl s genotype, which makes the cells unable to grow on lactose minimal plates. In contrast, spontaneous or mutagen-induced supFmutations in pUB3 prevent suppression of lacl '."2~' and result in constitutive expression of the lactose operon, which permits growth on lactose minimal plates. The spontaneous mutation frequency in the supF gene is ~ 0.7 and ~ 1.0 x 10-~ without and with SOS induction, respectively. Spontaneous mutations are dominated by large insertions (67% in SOS-uninduced and 56% in SOS-induced cells), and their frequency of appearance is largely unaffected by SOS induction. These are identified by DNA sequencing to be Insertion Elements; IS1 dominates, but IS4, ISS, gamma-delta and ISl0 are also obtained. Large deletions also contribute significantly (19% and 15% for-SOS and +SOS, respectively), where a specific deletion between a 10 base pair direct repeat dominates; the frequency of appearance of these mutations also appears to be unaffected by SOS induction, in contrast, SOS induction increases base pairing mutations (13% and 27% for-SOS and + SOS, respectively). The ES87/pUB3 system has many advantages for determining mutational spectra, including the fact that mutant isolation is fast and simple, and the determination of mutational changes is rapid because of the small size of supF.
Forkhead box M1 (FoxM1) oncogenic transcription factor represents an attractive therapeutic targe... more Forkhead box M1 (FoxM1) oncogenic transcription factor represents an attractive therapeutic target in the fight against cancer, because it is overexpressed in a majority of human tumors. Recently, using a cell-based assay system we identified thiazole antibiotic Siomycin A as an inhibitor of FoxM1 transcriptional activity. Here, we report that structurally similar thiazole antibiotic, thiostrepton also inhibits the transcriptional activity of FoxM1. Furthermore, we found that these thiopeptides did not inhibit the transcriptional activity of other members of the Forkhead family or some non-related transcription factors. Further experiments revealed that thiazole antibiotics also inhibit FoxM1 expression, but not the expression of other members of the Forkhead box family. In addition, we found that the thiazole antibiotics efficiently inhibited the growth and induced potent apoptosis in human cancer cell lines of different origin. Thiopeptide-induced apoptosis correlated with the suppression of FoxM1 expression, while overexpression of FoxM1 partially protected cancer cells from the thiazole antibiotic-mediated cell death. These data suggest that Siomycin A and thiostrepton may specifically target FoxM1 to induce apoptosis in cancer cells and FoxM1 inhibitors/thiazole antibiotics could be potentially developed as novel anticancer drugs against human neoplasia.
Supplemental data Fig.1. Combination treatment of ARC and ABT-737 is synergistic in inducing mito... more Supplemental data Fig.1. Combination treatment of ARC and ABT-737 is synergistic in inducing mitochondrial injury in tumor cells. A. DM833 cells were treated with ARC, ABT-737 or both as indicated, stained with TMRE and subjected to flow cytometric analysis. Mean fluorescence of cells due to bound TMRE is shown. B. U2OS-C3 cells treated with the drugs for 24 hrs were stained with TMRE and analyzed by flow cytometry. Mean fluorescence of cells due to bound TMRE is shown. Fig.2. Combination treatment with ARC and ABT-737 does not affect Bcl-2 and Bax levels. A. DM366 melanoma cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin. B. HepG2 liver cancer cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin. C. SW480 colon cancer cells were treated with ARC, ABT-737 and combination of ARC/ABT-737 as shown and immunoblotted for Bcl-2, Bax and β-actin.
Bergenia crassifolia accumulates 3-O-monoglycosides and 3-00diglycosides of kaempferol and querce... more Bergenia crassifolia accumulates 3-O-monoglycosides and 3-00diglycosides of kaempferol and quercetin and lacks both myricetin derivatives and triglycosides. This is one of the simpler patterns of flavonoids observed in the Saxifragaceae. Francoa sonchifolia also produces a very simple array of flavonoids. Myricetin is lacking but a 3-00monoglycoside, two 3-O-diglycosides and a 3-O-triglycoside were observed. This taxon is distinguishable from most others in the family by the presence of 3-0 ° methylquercetin 7-00glucoside. The flavonoid profile of Parnassia is markedly more complex than any other genus of the Saxifragaceae studied to date. Its profile comprises flavonol 3-O-monoglycosides and 3-O-diglycosides but is unique in possessing large numbers of flavonol 3,7-diglycosides and 3,7-triglycosides. Lepuropetalon spatulum resembles Parnassia in flavonoid pattern more than it resembles any other genus of the Saxifragaceae. The combination of flavonoid data with published information on a variety of other characters leads us to agree with the position of Bergenia as a member of Saxifragaceae in the restricted sense. The flavonoids of Francoa, Parnassia and Lepuropetalon agree with suggestions that these genera stand apart from the herbaceous Saxifragaceae.
The oncogenic transcription factor forkhead box M1 (FoxM1) is overexpressed in a number of differ... more The oncogenic transcription factor forkhead box M1 (FoxM1) is overexpressed in a number of different carcinomas, whereas its expression is turned off in terminally differentiated cells. For this reason, FoxM1 is an attractive target for therapeutic intervention in cancer treatment. As a first step toward realizing this goal, in this study, using a high-throughput, cell-based assay system, we screened for and isolated the antibiotic thiazole compound Siomycin A as an inhibitor of FoxM1. Interestingly, we observed that Siomycin A was able to down-regulate the transcriptional activity as well as the protein and mRNA abundance of FoxM1. Consequently, we found that the downstream target genes of FoxM1, such as Cdc25B, Survivin, and CENPB, were repressed. Also, we observed that consistent with earlier reports of FoxM1 inhibition, Siomycin A was able to reduce anchorage-independent growth of cells in soft agar. Furthermore, we found that Siomycin A was able to induce apoptosis selectively in transformed but not normal cells of the same origin. Taken together, our data suggest that FoxM1 inhibitor Siomycin A could represent a useful starting point for the development of anticancer therapeutics. (Cancer Res 2006; 66(19): 9731-5)
Recently, we identified a nucleoside analog named ARC (4-amino-6-hydrazino-7-beta-D-ribofuranosyl... more Recently, we identified a nucleoside analog named ARC (4-amino-6-hydrazino-7-beta-D-ribofuranosyl-7H-Pyrrolo[2,3-d]pyrimidine-5-carboxamide), which has the properties of a general transcriptional inhibitor. Here, we report the characterization of ARC on a panel of colorectal cancer (CRC) cell lines. Cell death induced by ARC in CRC cells was accompanied by caspase-3 cleavage and correlated with the downregulation of antiapoptotic proteins, survivin and Mcl-1 and with the inhibition of Akt phosphorylation. At the same time, colon cancer cell lines were resistant to the well-known nucleoside analog DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole), which failed to downregulate Mcl-1 or survivin. Overall, ARC could represent an attractive candidate for anti-cancer drug development that targets multiple survival pathways in colon cancer cells.
We previously described the identification of a nucleoside analog transcriptional inhibitor ARC (... more We previously described the identification of a nucleoside analog transcriptional inhibitor ARC (4-amino-6-hydrazino-7-beta-Dribofuranosyl-7H-Pyrrolo[2,3-d]-pyrimidine-5-carboxamide) and FoxM1 inhibitor, thiazole antibiotic Siomycin A that were able to induce apoptosis in cancer cell lines of different origin. Here, we report the characterization of these drugs on a panel of melanoma cell lines. We found that in contrast to the common anti-melanoma drug dacarbazine (DTIC), ARC and thiazole antibiotics, Siomycin A and thiostrepton, efficiently inhibited growth and induced cell death in melanoma cell lines in low concentrations. Overexpression of the antiapoptotic protein Mcl-1 protected melanoma cells from apoptosis induced by these compounds. Furthermore, we found that ARC and Siomycin A synergistically induce apoptosis in DM833 melanoma cell line suggesting that they may antagonize different anti-apoptotic pathways in melanoma cells. In general, these drugs may represent important candidates for anti-cancer drug development against melanoma.
PDF file - 13093K, Supplementary Fig.1: Loss of DDB2 expression in human colon cancer. Supplement... more PDF file - 13093K, Supplementary Fig.1: Loss of DDB2 expression in human colon cancer. Supplementary Fig.2: Low magnification IHC data showing reduced expression of DDB2 in high grade colon cancer samples. Supplementary Fig.3: Depletion of DDB2 increases invasivness of colon cancer cells. Supplementary Fig.4: Polyclonal DDB2 knockdown colon cancer cells exhibit increased tumorigenicity Supplementary Fig.5: Polyclonal DDB2 knockdown colon cancer cells exhibit EMT Supplementary Fig.6: DDB2 knockdown colon cancer cells generate aggressive tumors in nude mice Supplementary Fig.7: Low magnification picture showing EMT in xenografted tumors Supplementary Fig.8: DDB2 expression blocks EMT induced by hypoxia and TGF-beta Supplementary Fig.9: Chromatin-IP using T7-antibody demonstrating interaction of T7-DDB2 with the VEGF, Snail and Zeb1 promoters. Supplementary Fig.10: Transcription factor binding sites in the promoter fragments bound by DDB2 Supplementary Fig.11: Western blot analyses for the DDB2-associated proteins and XPC in DDB2-depleted cells. Supplementary Fig.12: Experimental metastasis assay with polyclonal DDB2 knockdown cells. Supplementary Fig.13: Expression of DDB2 does not inhibit proliferation in SW620 cells.
Colon cancer is one of the deadliest cancers worldwide because of its metastasis to other essenti... more Colon cancer is one of the deadliest cancers worldwide because of its metastasis to other essential organs. Metastasis of colon cancer involves a complex set of events, including epithelial-to-mesenchymal transition (EMT) that increases invasiveness of the tumor cells. Here, we show that the xeroderma pigmentosum group E (XPE) gene product, damaged DNA-binding protein (DDB)-2, is downregulated in high-grade colon cancers, and it plays a dominant role in the suppression of EMT of the colon cancer cells. Depletion of DDB2 promotes mesenchymal phenotype, whereas expression of DDB2 promotes epithelial phenotype. DDB2 constitutively represses genes that are the key activators of EMT, indicating that DDB2 is a master regulator of EMT of the colon cancer cells. Moreover, we observed evidence that DDB2 functions as a barrier for EMT induced by hypoxia and TGF-b. Also, we provide evidence that DDB2 inhibits metastasis of colon cancer. The results presented here identify a transcriptional regulatory pathway of DDB2 that is directly linked to the mechanisms that suppress metastasis of colon cancer. Cancer Res; 73(12); 3771-82. Ó2013 AACR.
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Papers by Uppoor Bhat