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Dysregulation of Catalase and Glutathione Peroxidase1 Impacts Diffuse Large B-Cell Lymphoma (DLBCL) Patient Survival
phosphorylated forms of Akt, Erk1/2, Jnk and p38 were lower in EGCG pre-treated HCT116 cells which were treated then with 200μM H2O2 for 24h and 48h. Furthermore, our findings suggested that treatment of HCT116 cells with 200μM EGCG led DQLQFUHDVHLQ7*)ȕSURWHLQOHYHOVLQDtime dependent manner starting after 6 hours of treatment. Nrf2 and its downstream heme oxygenase 1 (HMOX1) both mRNA and protein levels found to be increased in the same fashion following EGCG treatment in these cells. Our studies are ongoing to explore the role of 7*)ȕ signaling on Nrf2 levels. doi: 10.1016/j.freeradbiomed.2014.10.217
321 Reactive Oxygen Species Involved Cancer Cellular Specific 5-Aminolevulinic Acid Uptake in Gastric Epithelial Cells Masahiko Terasaki1, Hiromu Ito1, Tsuyoshi Kaneko1, Hirofumi Matsui1, and Ichinosuke Hyodo1 1 Faculty of medicine, University of Tsukuba, Japan Photodynamic therapy (PDT) and photodynamic diagnosis (PDD) using 5-aminolevulinic acid (ALA) are clinically useful for cancer treatments. Cancer cells have been reported that ALA is incorporated via peptide transporter 1 (PEPT1), which is one of the membrane transport proteins, and has been reported to be significantly expressed in various gastrointestinal cancer cells such as Caco-2. However, the mechanism of this protein expression has not been elucidated. Concentration of Reactive oxygen species (ROS) is higher in cancer cells in comparison with that of normal cells. We have previously reported that ROS derived from mitochondria (mitROS) is likely related to invasions and proliferations of cancer cells (Tamura M. et al. J Clin Biochem Nutr 2014). Since ALA is the most important precursor of heme which is necessary protein for cellular proliferations, mitROS may be also related to PEPT1 expressions. In this study, we used a rat gastric mucosal cell line RGM1 and its cancer-like mutated cell line RGK1, and we clarified the ALA uptake mechanism and its relations between mitROS and PEPT1 expression in RGK1. We also used our self-established stable clone of cell which overexpresses manganese superoxide dismutase, a mitROS scavenger. We studied differences of the PDT effects in these cells after ALA administrations to clear the influence of mitROS. We have succeeded to demonstrate following two conclusions: 1) PEPT1 is most likely to be a membrane transporter for cancer specific ALA incorporation. 2) cancer specific high mitROS concentration plays an important role for this phenomenon. doi: 10.1016/j.freeradbiomed.2014.10.218
322 Dysregulation of Catalase and Glutathione Peroxidase1 Impacts Diffuse Large B-Cell Lymphoma (DLBCL) Patient Survival Kristy Lee1, Heather S. Baldwin1, Gregory R. Martin2, Florence A. Scholl2, Lisa M. Rimsza1, and Margaret E. Tome1 1 University of Arizona, USA, 2Ventana Medical Systems, Inc., USA Tumor cells have a more oxidized set point than normal cells. Consequences of a higher level of oxidants include increased proliferation and mutation frequency each of which can enhance tumorigenesis. Decreased antioxidant defense (AOD) enzymes
S134
can contribute to this effect. One chemotherapeutic strategy for cancer treatment is to use drugs that increase oxidants in the tumor cells to an even greater degree resulting in apoptosis. Elevated AOD enzymes can decrease drug toxicity in tumor cell culture and mouse model systems. Modulating the levels of catalase and glutathione peroxidase (GPX1) alters intracellular H2O2 and the response to oxidants. Although the data from model systems suggest that the levels of catalase and GPX1 should influence lymphoma patient survival, previous analyses of patient data have produced discrepant results. The goal of this study was to combine gene expression profile (GEP) dataset analysis with immunohistochemical (IHC) staining of tumor tissue to: 1) better define the role of catalase and GPX1 in lymphoma biology; and 2) test our prediction that moderate expression of these enzymes would result in better lymphoma patient survival than low (expected to contribute to lymphomagenesis) or high (expected to contribute to chemoresistance) expression. Analysis of GEP datasets showed that catalase and GPX1 exhibited a wide range of expression levels in three types of lymphoma. IHC staining indicated that lymphoma tumor tissue could have greater or less protein that their normal cell counterparts. We found that neither catalase nor GPX1 was linearly correlated with patient survival in DLBCL. However, Kaplan-Meier curves indicated that patients with moderate expression (middle two quartiles) for both catalase and GPX1 had better survival than those in either the top or bottom quartiles. IHC staining of DLBCL tumor samples from 12 patients for whom survival was known showed a trend toward high and low GPX1 expression correlating with worse survival. These data indicate that: 1) catalase and GPX1 are dysregulated in lymphoma; and 2) DLBCL patients with moderate catalase and GPX1 expression have longer survival than those with high or low expression of either enzyme. doi: 10.1016/j.freeradbiomed.2014.10.219
323 Alterations in Epigenetic Markers Related to 5Hydroxymethylcytosine in Brain Cancer Carolyne Lowry1, Marie-Belle Poirier1, David Fortin1, and J. Richard Wagner1 1 Université de Sherbrooke, Canada 5-Hydroxymethylcytosine (5hmC) is an intermediate in the enzymatic oxidation of 5-methylcytosine (5mC) by ten eleven translocation (TET) D±ketoglutarate (D-KG) and Fe(II)-dependent dioxygenases. The level 5hmC is relatively high in brain compared to other organs, and these levels are significantly reduced in various types of cancer, particularly in brain cancer. Here, we measured 5mC, 5hmC, and related modifications in the DNA of tumor specimens from glioblastoma multiforme (GBM, n=37) and early stage astrocytomas (Grade I-III; n=12) using mass spectrometry (LC-MS/MS). The average level of 5mC was 4.5 and that of 5hmC was 0.80 in units of % cytosine. The level of 5hmC in the DNA of GBM was several-fold lower than that obtained from lower grade tumors. In contrast, the level of 5formylcytosine (5fC), 5-hydroxymethyluracil (5hmU) and 5formyluracil (5fU) were at least 10-fold lower compared to that of 5hmC in the DNA of GBM specimens. Interestingly, there was a strong correlation between the level of 5fC and 5fU (r2= 0.6), whereas other correlations involving 5hmC and 5fC, and 5hmU and 5fU were weak (r2<0.2). TET 1 and 2 gene expression analysis indicated a strong correlation between TET 1 and TET 2 (r2= 0.7), a 2.2-fold decrease in gene expression in GBM compared to grade II and III specimens, but no apparent correlation with 5mC or other modifications. Our studies confirm and report additional epigenetic changes in brain cancer.
SFRBM 2014
321 Reactive Oxygen Species Involved Cancer Cellular Specific 5-Aminolevulinic Acid Uptake in Gastric Epithelial Cells Masahiko Terasaki1, Hiromu Ito1, Tsuyoshi Kaneko1, Hirofumi Matsui1, and Ichinosuke Hyodo1 1 Faculty of medicine, University of Tsukuba, Japan Photodynamic therapy (PDT) and photodynamic diagnosis (PDD) using 5-aminolevulinic acid (ALA) are clinically useful for cancer treatments. Cancer cells have been reported that ALA is incorporated via peptide transporter 1 (PEPT1), which is one of the membrane transport proteins, and has been reported to be significantly expressed in various gastrointestinal cancer cells such as Caco-2. However, the mechanism of this protein expression has not been elucidated. Concentration of Reactive oxygen species (ROS) is higher in cancer cells in comparison with that of normal cells. We have previously reported that ROS derived from mitochondria (mitROS) is likely related to invasions and proliferations of cancer cells (Tamura M. et al. J Clin Biochem Nutr 2014). Since ALA is the most important precursor of heme which is necessary protein for cellular proliferations, mitROS may be also related to PEPT1 expressions. In this study, we used a rat gastric mucosal cell line RGM1 and its cancer-like mutated cell line RGK1, and we clarified the ALA uptake mechanism and its relations between mitROS and PEPT1 expression in RGK1. We also used our self-established stable clone of cell which overexpresses manganese superoxide dismutase, a mitROS scavenger. We studied differences of the PDT effects in these cells after ALA administrations to clear the influence of mitROS. We have succeeded to demonstrate following two conclusions: 1) PEPT1 is most likely to be a membrane transporter for cancer specific ALA incorporation. 2) cancer specific high mitROS concentration plays an important role for this phenomenon. doi: 10.1016/j.freeradbiomed.2014.10.218
322 Dysregulation of Catalase and Glutathione Peroxidase1 Impacts Diffuse Large B-Cell Lymphoma (DLBCL) Patient Survival Kristy Lee1, Heather S. Baldwin1, Gregory R. Martin2, Florence A. Scholl2, Lisa M. Rimsza1, and Margaret E. Tome1 1 University of Arizona, USA, 2Ventana Medical Systems, Inc., USA Tumor cells have a more oxidized set point than normal cells. Consequences of a higher level of oxidants include increased proliferation and mutation frequency each of which can enhance tumorigenesis. Decreased antioxidant defense (AOD) enzymes
S134
can contribute to this effect. One chemotherapeutic strategy for cancer treatment is to use drugs that increase oxidants in the tumor cells to an even greater degree resulting in apoptosis. Elevated AOD enzymes can decrease drug toxicity in tumor cell culture and mouse model systems. Modulating the levels of catalase and glutathione peroxidase (GPX1) alters intracellular H2O2 and the response to oxidants. Although the data from model systems suggest that the levels of catalase and GPX1 should influence lymphoma patient survival, previous analyses of patient data have produced discrepant results. The goal of this study was to combine gene expression profile (GEP) dataset analysis with immunohistochemical (IHC) staining of tumor tissue to: 1) better define the role of catalase and GPX1 in lymphoma biology; and 2) test our prediction that moderate expression of these enzymes would result in better lymphoma patient survival than low (expected to contribute to lymphomagenesis) or high (expected to contribute to chemoresistance) expression. Analysis of GEP datasets showed that catalase and GPX1 exhibited a wide range of expression levels in three types of lymphoma. IHC staining indicated that lymphoma tumor tissue could have greater or less protein that their normal cell counterparts. We found that neither catalase nor GPX1 was linearly correlated with patient survival in DLBCL. However, Kaplan-Meier curves indicated that patients with moderate expression (middle two quartiles) for both catalase and GPX1 had better survival than those in either the top or bottom quartiles. IHC staining of DLBCL tumor samples from 12 patients for whom survival was known showed a trend toward high and low GPX1 expression correlating with worse survival. These data indicate that: 1) catalase and GPX1 are dysregulated in lymphoma; and 2) DLBCL patients with moderate catalase and GPX1 expression have longer survival than those with high or low expression of either enzyme. doi: 10.1016/j.freeradbiomed.2014.10.219
323 Alterations in Epigenetic Markers Related to 5Hydroxymethylcytosine in Brain Cancer Carolyne Lowry1, Marie-Belle Poirier1, David Fortin1, and J. Richard Wagner1 1 Université de Sherbrooke, Canada 5-Hydroxymethylcytosine (5hmC) is an intermediate in the enzymatic oxidation of 5-methylcytosine (5mC) by ten eleven translocation (TET) D±ketoglutarate (D-KG) and Fe(II)-dependent dioxygenases. The level 5hmC is relatively high in brain compared to other organs, and these levels are significantly reduced in various types of cancer, particularly in brain cancer. Here, we measured 5mC, 5hmC, and related modifications in the DNA of tumor specimens from glioblastoma multiforme (GBM, n=37) and early stage astrocytomas (Grade I-III; n=12) using mass spectrometry (LC-MS/MS). The average level of 5mC was 4.5 and that of 5hmC was 0.80 in units of % cytosine. The level of 5hmC in the DNA of GBM was several-fold lower than that obtained from lower grade tumors. In contrast, the level of 5formylcytosine (5fC), 5-hydroxymethyluracil (5hmU) and 5formyluracil (5fU) were at least 10-fold lower compared to that of 5hmC in the DNA of GBM specimens. Interestingly, there was a strong correlation between the level of 5fC and 5fU (r2= 0.6), whereas other correlations involving 5hmC and 5fC, and 5hmU and 5fU were weak (r2<0.2). TET 1 and 2 gene expression analysis indicated a strong correlation between TET 1 and TET 2 (r2= 0.7), a 2.2-fold decrease in gene expression in GBM compared to grade II and III specimens, but no apparent correlation with 5mC or other modifications. Our studies confirm and report additional epigenetic changes in brain cancer.
SFRBM 2014