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OP18 Utilization of hydrogen sulfide donor as a cancer therapy
Abstracts / Nitric Oxide 31 (2013) S11–S65
contributes to their proliferation, migration and invasion. Together with additional in vitroand in vivo data presented at the current meeting, these data support the view that CBS-derived H2S serves as an endogenous cancer cell survival factor, and identify CBS as a potential future antitumor drug target. http://dx.doi.org/10.1016/j.niox.2013.06.047
OP18 Utilization of hydrogen sulfide donor as a cancer therapy Zheng-Wei Lee a, Evon Yi-Wen Tay a, Choon-Hong Tan b, Thilo Hagen a, Philip Keith Moore c, Lih-Wen Deng a a Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore b Division of Chemistry and Biological Chemistry, Nanyang Technological University, Singapore 637371, Singapore c Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore Very little is known regarding role of hydrogen sulfide (H2S) in cancer biology. We showed that slow H2S donor, GYY4137 [1] exhibited anti-cancer effect both in vitro and in vivo. Using conventional H2S detection method methylene blue assay, we demonstrated the different H2S-releasing manner of sodium hydrosulfide (NaHS) and GYY4137. NaHS releases H2S acutely and in high amount as contrast to GYY4137 which releases H2S slowly and in low yet detectable level. Surprisingly, anti-proliferative effect of H2S was observed across a broad range of cancer cell lines (HeLa, HCT-116, HepG2, U2OS, MCF7, HL-60 and MV4–11) in GYY4137 treatment but was of little effect or not significant in NaHS treatment. This suggested that prolonged exposure to low concentration of H2S may serve as novel anti-cancer strategy. Furthermore, normal cell lines (WI38 and IMR90) tested were not affected by GYY4137 nor NaHS. Tumor xenograft mice model too showed a significant shrinkage of tumor volume in a GYY4137 dose-dependent manner [2]. Further analysis on this selective phenomena suggested that H2S killing effect may be glucose metabolism related. We identified that H2S induced an altered glucose metabolism rate and interfered with pH homeostasis that led to significant pH disruption in cancer cells. This eventually drives cancer cells to death. On the other hand, pH of normal cells was not affected by H2S. Selectivity of H2S targeting on cancer but not normal cells would serve as a novel and effective cancer therapy. References [1] Li et al., Circulation 117 (2008) 2351–2360. [2] Lee et al., PloS One 6 (2011) e21077. http://dx.doi.org/10.1016/j.niox.2013.06.048
OP019 H2S attenuates DNA damage by targeting at MEK1 and PAR Kexin Zhao a,b, Rui Wang a,c, Guangdong Yang a,b a Cardiovascular and Metabolic Research Unit, Lakehead University, Thunder Bay, Ontario, Canada b The School of Kinesiology, Lakehead University, Thunder Bay, Ontario, Canada c Department of Biology, Lakehead University, Thunder Bay, Ontario, Canada
S27
Hydrogen sulfide (H2S), a novel member of gasotransmitter family along with nitric oxide and carbon monoxide, exerts a wide range of cellular and molecular actions in our body. Cystathionine gammalyase (CSE) is a major H2S-generating enzyme in mammalian. The maintenance of genome integrity is essential for the proper function and survival of all organisms, and DNA damage may result in blockages of transcription and replication, mutagenesis, cellular senescence, and carcinogenesis. In this study, we demonstrated that more DNA damages occur in mouse embryonic fibroblasts (MEFs) from CSE knockout mice in comparison with that from wild-type mice, and H2S at physiologically relevant concentration attenuates DNA damage and cellular senescence in human umbilical vein endothelial cells (HUVECs). Treatment of HUVECs with 10 lM NaHS stimulated poly (ADP-ribosyl) ation (PAR) activation, while knockdown of CSE expression by RNA interference or inhibition of CSE activity by 5 mM DL-propargyglycine attenuated PAR activation. We also observed that H2S stimulates more PAR, XRCC-1 and DNA ligase III complex in HUVECs, while H2S had little effect on the release of apoptosis inducing factor from mitochondria. We further found that PAR levels are significantly higher in liver, kidney and MEFs from wild-type mice when compared with those from CSE knockout mice. H2S induced the interaction of PAR and phosphorylated ERK1/2, and U0126 (an inhibitor of MEK1/2) reversed H2S-induced PAR activation. MEK1, but not MEK2 and PARP-1 were S-sulfhydrated by H2S, following increased ERK1 phosphorylation and PAR activation. These results provide mechanistic insight into how H2S signalling mediates DNA damage and cellular senescence (supported by a grant-in-aid from Heart and Stroke Foundation of Canada to G.Y.). http://dx.doi.org/10.1016/j.niox.2013.06.049
OP20 Inhibition of recombinant T-type Ca2+ channels by hydrogen sulfide J. Elies, M. Dallas, J.L. Scragg, J.P. Boyle, C. Peers Leeds Institute of Genetics, Health and Therapeutics, Faculty of Medicine and Health, University of Leeds, and School of Pharmacy, University of Reading, UK Awareness of H2S as an important, widespread signalling molecule is growing rapidly. It is known to influence, for example, proliferation of vascular smooth muscle cells and pain sensation [1]. Interestingly, both of these processes involve T-type Ca2+ channel activity [2,3], and ion channels are emerging as an important family of target proteins for modulation by H2S [4]. We have therefore investigated whether H2S modulates T-type Ca2+ channels, using whole-cell patch clamp recordings from HEK293 cells over-expressing, separately, the three known type of T-type Ca2+ channels, Cav3.1, Cav3.2 and Cav3.3 [5]. Cells were exposed to H2S by bath application of NaHS. This agent caused a concentration-dependent (10 lM to 1 mM) inhibition of currents in Cav3.2-expressing cells. Inhibition was observed at all activating test potentials applied, and all subsequent values detailed are taken from currents recorded at a test potential of 20 mV (holding potential 80 mV). Maximal inhibition of 35.3 ± 2.2%, n = 6, P < 0.0001, student’s paired t-test was observed at the NaHS concentration of 1 mM. By contrast, current carried by Cav3.1 channels were unaffected over the same concentration range, and those carried by Cav3.3 were only modestly inhibited (13.2 ± 3.6%, n = 5, P = 0.022) by 1 mM NaHS. Thus, H2S appeared to inhibit Cav3.2 channels selectively. Cav3.2 inhibition was only poorly reversible, but current amplitudes could be recovered by dithiothreitol (2 mM; n = 7), suggesting H2S acts via channel redox modulation.
contributes to their proliferation, migration and invasion. Together with additional in vitroand in vivo data presented at the current meeting, these data support the view that CBS-derived H2S serves as an endogenous cancer cell survival factor, and identify CBS as a potential future antitumor drug target. http://dx.doi.org/10.1016/j.niox.2013.06.047
OP18 Utilization of hydrogen sulfide donor as a cancer therapy Zheng-Wei Lee a, Evon Yi-Wen Tay a, Choon-Hong Tan b, Thilo Hagen a, Philip Keith Moore c, Lih-Wen Deng a a Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore b Division of Chemistry and Biological Chemistry, Nanyang Technological University, Singapore 637371, Singapore c Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore Very little is known regarding role of hydrogen sulfide (H2S) in cancer biology. We showed that slow H2S donor, GYY4137 [1] exhibited anti-cancer effect both in vitro and in vivo. Using conventional H2S detection method methylene blue assay, we demonstrated the different H2S-releasing manner of sodium hydrosulfide (NaHS) and GYY4137. NaHS releases H2S acutely and in high amount as contrast to GYY4137 which releases H2S slowly and in low yet detectable level. Surprisingly, anti-proliferative effect of H2S was observed across a broad range of cancer cell lines (HeLa, HCT-116, HepG2, U2OS, MCF7, HL-60 and MV4–11) in GYY4137 treatment but was of little effect or not significant in NaHS treatment. This suggested that prolonged exposure to low concentration of H2S may serve as novel anti-cancer strategy. Furthermore, normal cell lines (WI38 and IMR90) tested were not affected by GYY4137 nor NaHS. Tumor xenograft mice model too showed a significant shrinkage of tumor volume in a GYY4137 dose-dependent manner [2]. Further analysis on this selective phenomena suggested that H2S killing effect may be glucose metabolism related. We identified that H2S induced an altered glucose metabolism rate and interfered with pH homeostasis that led to significant pH disruption in cancer cells. This eventually drives cancer cells to death. On the other hand, pH of normal cells was not affected by H2S. Selectivity of H2S targeting on cancer but not normal cells would serve as a novel and effective cancer therapy. References [1] Li et al., Circulation 117 (2008) 2351–2360. [2] Lee et al., PloS One 6 (2011) e21077. http://dx.doi.org/10.1016/j.niox.2013.06.048
OP019 H2S attenuates DNA damage by targeting at MEK1 and PAR Kexin Zhao a,b, Rui Wang a,c, Guangdong Yang a,b a Cardiovascular and Metabolic Research Unit, Lakehead University, Thunder Bay, Ontario, Canada b The School of Kinesiology, Lakehead University, Thunder Bay, Ontario, Canada c Department of Biology, Lakehead University, Thunder Bay, Ontario, Canada
S27
Hydrogen sulfide (H2S), a novel member of gasotransmitter family along with nitric oxide and carbon monoxide, exerts a wide range of cellular and molecular actions in our body. Cystathionine gammalyase (CSE) is a major H2S-generating enzyme in mammalian. The maintenance of genome integrity is essential for the proper function and survival of all organisms, and DNA damage may result in blockages of transcription and replication, mutagenesis, cellular senescence, and carcinogenesis. In this study, we demonstrated that more DNA damages occur in mouse embryonic fibroblasts (MEFs) from CSE knockout mice in comparison with that from wild-type mice, and H2S at physiologically relevant concentration attenuates DNA damage and cellular senescence in human umbilical vein endothelial cells (HUVECs). Treatment of HUVECs with 10 lM NaHS stimulated poly (ADP-ribosyl) ation (PAR) activation, while knockdown of CSE expression by RNA interference or inhibition of CSE activity by 5 mM DL-propargyglycine attenuated PAR activation. We also observed that H2S stimulates more PAR, XRCC-1 and DNA ligase III complex in HUVECs, while H2S had little effect on the release of apoptosis inducing factor from mitochondria. We further found that PAR levels are significantly higher in liver, kidney and MEFs from wild-type mice when compared with those from CSE knockout mice. H2S induced the interaction of PAR and phosphorylated ERK1/2, and U0126 (an inhibitor of MEK1/2) reversed H2S-induced PAR activation. MEK1, but not MEK2 and PARP-1 were S-sulfhydrated by H2S, following increased ERK1 phosphorylation and PAR activation. These results provide mechanistic insight into how H2S signalling mediates DNA damage and cellular senescence (supported by a grant-in-aid from Heart and Stroke Foundation of Canada to G.Y.). http://dx.doi.org/10.1016/j.niox.2013.06.049
OP20 Inhibition of recombinant T-type Ca2+ channels by hydrogen sulfide J. Elies, M. Dallas, J.L. Scragg, J.P. Boyle, C. Peers Leeds Institute of Genetics, Health and Therapeutics, Faculty of Medicine and Health, University of Leeds, and School of Pharmacy, University of Reading, UK Awareness of H2S as an important, widespread signalling molecule is growing rapidly. It is known to influence, for example, proliferation of vascular smooth muscle cells and pain sensation [1]. Interestingly, both of these processes involve T-type Ca2+ channel activity [2,3], and ion channels are emerging as an important family of target proteins for modulation by H2S [4]. We have therefore investigated whether H2S modulates T-type Ca2+ channels, using whole-cell patch clamp recordings from HEK293 cells over-expressing, separately, the three known type of T-type Ca2+ channels, Cav3.1, Cav3.2 and Cav3.3 [5]. Cells were exposed to H2S by bath application of NaHS. This agent caused a concentration-dependent (10 lM to 1 mM) inhibition of currents in Cav3.2-expressing cells. Inhibition was observed at all activating test potentials applied, and all subsequent values detailed are taken from currents recorded at a test potential of 20 mV (holding potential 80 mV). Maximal inhibition of 35.3 ± 2.2%, n = 6, P < 0.0001, student’s paired t-test was observed at the NaHS concentration of 1 mM. By contrast, current carried by Cav3.1 channels were unaffected over the same concentration range, and those carried by Cav3.3 were only modestly inhibited (13.2 ± 3.6%, n = 5, P = 0.022) by 1 mM NaHS. Thus, H2S appeared to inhibit Cav3.2 channels selectively. Cav3.2 inhibition was only poorly reversible, but current amplitudes could be recovered by dithiothreitol (2 mM; n = 7), suggesting H2S acts via channel redox modulation.