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Sa1690 Proteomic and Metabolic Signaling Pathways in Colorectal Adenoma
between polyamines and MAT2A. Increased MAT2A expression will provide more SAMe for polyamines biosynthesis; increased polyamine (putrescine in this case) can activate MAT2A at the transcriptional level. This along with increased ODC expression in colon cancer all feed forward to further enhance the proliferative capacity of the cancer cell. This work was supported by NIH grants R01DK51719 and R01AT004896 (SC Lu, JM Mato), and F32AA020150 (ML Tomasi). RKO cells were provided by the Cell Culture Core of the USC Research Center for Liver Diseases (P30DK48522). Sa1689 Microsatellite Instability, BRAF and KRAS Mutation in Postcolonoscopy Cancers: An Explorative Study Chantal le Clercq, Robert Riedl, Mariëlle Bouwens, Beatriz Carvalho, Gerrit A. Meijer, Manon van Engeland, Bjorn Winkens, Ad Masclee, Silvia Sanduleanu A subset of postcolonoscopy colorectal cancers (PC-CRCs) may evolve through alternative molecular mechanisms, yet the biologic features of these cancers are largely unknown. We therefore retrospectively examined the histopathologic and molecular characteristics (i.e. MSI, BRAF, KRAS) of PC-CRCs diagnosed at our university hospital. Methods: We reviewed clinicopathologic data of all patients who were diagnosed with CRC at our institution from Jan 2001 to Dec 2010. Digital colonoscopy and histopathology records were collected and verified using the national pathology database (PALGA) as well as data from the Netherlands Cancer Registry. We excluded patients with hereditary forms of CRC, inflammatory bowel disease or previous history of CRC. We defined PC-CRCs as cancers occurring within 5 years after a complete index colonoscopy. According to location, CRCs were categorized into proximal or distal to the splenic flexure. We defined advanced adenomas, as adenomatous polyps ≥1cm, containing high grade dysplasia or a villous component. Histology of PCCRCs was revised by an experienced GI-pathologist and microsatellite instability (MSI), mismatch repair gene expression, hypermethylation of MLH1 and mutation analyses of KRAS (exon 2) and BRAF (V600E) genes were performed. Results: Out of a total of 1,218 patients with 1,267 CRCs, we identified 28 patients (mean age, 73.3 years; 71% men) with PC-CRCs. Overall, 68% of PC-CRCs were proximally located and all of them were adenocarcinomas. Of the 19 proximal PC-CRCs, 5 were MSI, showing loss of expression of both MLH1 and PMS2 proteins by immunohistochemistry, all in combination with hypermethylation of MLH1. Five out of these 19 PC-CRCs were BRAF mutated (2 of them being MSI), while in 3 cases KRAS mutations were found (all of them being MSS). Of the 9 distal PC-CRCs, 1 was BRAF mutated and 1 was KRAS mutated, and all of them were MSS [Figure]. Mean (range) size of PC-CRCs was 2.9 (0.5-5.5) cm, 48% of them were poorly differentiated, 11% were mucinous cancers and 32% contained lymphoid aggregates. TNM-stage of PC-CRCs was I, II, III and IV in 9, 6, 9 and 3 cases respectively, while in 1 case was unclear. At the time of the index colonoscopy, 12 patients had adenomas with 9 having advanced adenomas, 4 had hyperplastic polyps only, while 12 had no abnormalities. No SSA/Ps or TSAs were identified. Interestingly, 3 out of the 4 patients with hyperplastic polyps only developed BRAF mutated PC-CRC, one of which was MSI. Conclusion: In this population, 21% of the postcolonoscopy cancers were BRAF mutated MSS/MSI cancers suggesting an alternative biology might contribute to a minor proportion of these cancers. Comprehensive studies on the (epi)genetic molecular alterations of PC-CRC are needed to clarify their biology, as this information might provide the basis for personalized surveillance in the future.
Sa1687 SPARC Modulates Autophagy via Caspase 8 in the Presence of 5-Fluorouracil in Colorectal Cancer Mahbuba Rahman, Sharon Gorski, Isabella T. Tai Background and aims: Autophagy and apoptosis are two evolutionarily conserved processes regulating cellular fate in response to cytotoxic stress. SPARC, a matricellular protein, acts as a chemosensitizer in the presence of 5-fluorouracil (5FU) by activating the extrinsic apoptotic pathway protein caspase-8 in colorectal cancer (CRC). However, the functional relationship between autophagy and apoptosis in the presence of SPARC is largely unknown. Here we demonstrate that SPARC activation of caspase-8 in the presence of 5FU leads to a reduction in autophagic flux. Methods: The SPARC over-expressing colorectal cancer cell line MIP/SP and its empty vector control MIP/ZEO were treated with 5FU (5μM) for 72 hours. A MTT cell viability assay was performed to verify the effect of 5FU in the two cell lines. Apoptosis was assessed using TUNEL, the caspase 3/7 activity assay and western blot analysis of caspase-8 cleavage. Autophagosome accumulation was evaluated using LC3B immunofluorescence, and autophagic flux was assessed by western blot analysis of LC3BII in the presence and absence of Bafilomycin A1 (Baf). To investigate the role of SPARC or caspase-8 in autophagy modulation, the respective proteins were knocked down using SPARC-siRNA or caspase 8-siRNA. The effects on autophagy were monitored by the LC3BII flux assay and western blot analysis of autophagy proteins. Results: Treatment with 5FU resulted in reduced cell viability (MTT) in both cell lines, though was reduced significantly further in MIP/SP compared to MIP/ZEO. 5FU also led to increased TUNEL staining and reduced LC3B IF in both cell lines. However, the level of apoptosis (as assessed by casapse 8 and caspase 3/7 assay) was higher, and the reduction in autophagic flux appeared greater in the MIP/SP cell line compared to its control cell line MIP/ZEO. To investigate the potential role of SPARC in autophagy modulation, siRNA was used to knockdown the SPARC protein. Autophagic flux increased in both cell lines following SPARC knockdown. Concomitantly, the level of active caspase 8 and caspase 3/7 activity was reduced in MIP/SP even in the presence of 5FU. Knockdown of caspase 8 similarly resulted in a higher level of autophagy flux and also increased levels of ATG7 in the presence of 5FU in the MIP/SP cell line, but not in MIP/ZEO. Conclusion: These observations demonstrate that knockdown of caspase 8 in MIP/SP leads to elevated Atg7 and can partially rescue autophagic flux, suggesting that SPARC inhibits autophagy via a caspase 8-mediated mechanism. High levels of SPARC upregulate caspase 8, which not only affects the extrinsic pathway of apoptosis but also reduces autophagic flux. Our findings indicate that SPARC can augment chemosensitivity by affecting both the apoptosis and the autophagy pathways. Sa1688 Polyamine and Methionine Adenosyltransferase 2A Crosstalk in Human Colon Cancer Maria Lauda Tomasi, Minjung Ryoo, Anna Skay, Ivan Tomasi, Pasquale Giordano, Jose M. Mato, Shelly C. Lu ACKGROUND & AIMS: Colon cancer is one of the most common forms of cancer and a leading cause of cancer associated deaths in the United States. Methionine adenosyltransferase 2A (MAT2A) encodes for the enzyme that catalyzes the formation of S-adenosylmethionine (SAMe), the principal biological methyl donor and precursor of polyamines. We reported that MAT2A expression is increased in human colon cancer and in colon cancer cells treated with mitogens. Induction of MAT2A was required in order for several growth factors to exert the mitogenic effect. We also observed that silencing MAT2A resulted in apoptosis. The aim of the current work was to examine the mechanism responsible for MAT2Adependent growth and apoptosis. METHODS: Studies were conducted using human colon cancer cell line RKO and colon cancer specimens. Real-time PCR and Western blotting measured gene and protein expression, respectively. Expression of MAT2A and ornithine decarboxylase (ODC) was varied using siRNA or overexpression vector. RESULTS: In RKO cells, MAT2A siRNA treatment for 24 hours reduced MAT2A mRNA level by 75% and resulted in depletion of cellular SAMe and putrescine levels by 70 and 75%, respectively, increased apoptosis and growth suppression. Spermidine and spermine levels were unchanged at this time point but fell by 48 to 72 hours afterwards. Putrescine supplementation (100pmol/L) blunted significantly MAT2A siRNA-induced apoptosis and growth suppression. Putrescine treatment raised MAT2A mRNA level by 4.3-fold. In addition, putrescine treatment also increased the expression of AP-1 family members c-Jun and c-Fos, which are known to trans-activate the human MAT2A promoter and increase its expression. In human colon cancer specimens, the mRNA and protein levels of MAT2A, ODC, c-Jun and c-Fos are all elevated as compared to adjacent non-tumorous tissues. Overexpression of ODC (the ratelimiting enzyme in polyamine biosynthesis that catalyzes the formation of putrescine) in RKO cells also raised MAT2A mRNA level by 50%. CONCLUSIONS: There is a cross-talk
Sa1690 Proteomic and Metabolic Signaling Pathways in Colorectal Adenoma Miao cui, Liang Peng, Zhiqing Wang, Yuan Hu, Xinying Wang, Yu Zhang, Juan Wang, David Zhang, Bo Jiang, Huabao Xiong, Ming-Ming Zhou, Fei Ye Background Colorectal cancer (CRC) is the leading cause of cancer deaths worldwide. One million new cases are diagnosed each year around which lead to 500,000 deaths too. Colorectal adenoma is the precancerous lesions of colorectal cancers. An adenoma takes many years to progress to carcinoma and changes in both proteomic and metabolic pathways were observed during the progression. Detection of those adenomas progressing to in situ carcinoma provides a window of opportunity for removing localized lesions, thus cure. The aim of this study is to identify proteomic and metabolic signaling pathways activated in the colorectal adenoma to facilitate the understanding of underlying mechanism. Methods The expression of 27 metabolic proteins and 172 proteins of signaling pathway in 37 paired colorectal adenoma samples were assessed using Protein Pathway Array (PPA). The value of expression of phosphorylation was normalized by Z-score. Data were analyzed by Significance Analysis of Microarrays (SAM) to detect differentially expressed proteins. Both of proteins and samples are clustered in two-dimensional hierarchical cluster by MultiExperiment Viewer v4.8 (MeV). Principal Component Analysis (PCA) was performed using Partek Genomics
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AGA Abstracts
cases (8.7%) and was significantly correlated with MSI (p ,0.01). PTEN inactivation was present in 53 cases (24%) and was significantly correlated with deeper invasion (p=0.02). IGF1R was positive in 62 cases (29%) and was significantly correlated with intestinal histological type (p=0.03) and MSI (p=0.01). Although pAkt expression was only correlated with HER2 overexpression (p=0.005), pAkt expression was more frequent in cases with PIK3CA exon 20 mutations (100%) than in those with other PIK3CA mutations (50%). In multivariable analysis of prognostic factors, male, Stage 3 or 4, PTEN inactivation and pAkt expression were significantly correlated with poor prognosis. Among the molecular alterations analyzed in this study, 90 cases (43%) showed a single alteration and 32 cases (15%) showed two or more alterations. With increase in the number of alterations, frequency of pAkt expression tended to increase and prognosis tended to be worse. p95HER2 expression was positive in 13 (34%) of 38 HER2-positive cases. The 3-year survival rate of patients with p95HER2 positive was shorter than that of other patients (48% vs 68%). Conclusions: The number of alterations in each patient was significantly correlated with poor prognosis and PI3K-Akt pathway activation. Our findings indicate that there are some interactions among alterations in the PI3K-Akt pathway and that they could affect prognosis. We characterized p95HER2 expression in GC for the first time. This result supports the use of p95HER2 expression as a biomarker in a future clinical trial of Tmab for GC.
AGA Abstracts
Suite 6.0. The signaling pathways, metabolic pathways and network constituted by these proteins were identified by using Ingenuity Pathway Analysis (IPA). Results Of 27 metabolic proteins and 172 proteins of signaling pathway screened, 82 were detected in 37 paired colorectal adenoma samples and among which 19 were identified as significant proteins (Fig 1) by comparing between adenoma and normal tissues (Q-value ,0.05). Of 19 proteins, 18 were up-regulated in adenoma including MetRS, beta-catenin, p-cdc2 (Tyr15), p-PDK1 (Ser241), et al, while one phosphoprotein was down-regulated, i.e. p-ERK (Thr202/ Tyr204)(P44/42). Sixty four matched samples (37 adenoma and their matched normal mucosa) were used for hierarchical clustering analysis and these 19 proteins correctly grouped adenoma by PCA with 89% accuracy. The top signaling pathways (Fig 2 a) affected in colorectal adenoma were IL-3 pathway (-log(p-value)= 7.54), NF- κB pathway (-log(p-value)= 5.64), et al. The top metabolic pathways (Fig 2 b) affected in colorectal adenoma were Aldosterone pathway (-log(p-value)= 3.12), Docosahexaenoic Acid (DHA) pathway (-log(pvalue)= 2.94), et al. Conclusion Our current study demonstrated that many signaling pathways and metabolic pathways were affected in colorectal adenoma which may be involved in development of adenoma. Inflammatory pathways (IL-3, NF- κB and PI3K/AKT) and metabolic pathways (docosahexaenoic acid and leptin) are altered in colorectal adenoma, suggesting the critical role of these pathways.
of activated STAT3 (PIAS3) is the target of miR-181b-1, and further show that miR-181b1 promotes glycolysis by STAT3 activated in colon cancer cells, through down-regulating PIAS3. As STAT3 activation has been reported could promote the transcription and expression of miR-181b-1. Therefore, our results support a positive feedback loop as a mechanism for persistent STAT3 activation, in which miR-181b-1 represses PIAS3 to elevate STAT3 activity, which in turn promotes miR-181b-1 transcription. PIAS3 was found down-regulated and miR-181b-1 up-regulated in human colon tumors, and inverse correlation of miR-181b-1 and PIAS3 expression was also found in colon tumors. Moreover, miR-181b-1 inhibition or PIAS3 up-regulation in colon cancer cells led to reduce STAT3 activation, glycolysis and attenuated engraft tumour growth, indicating that miR-181b-1 overexpression contributes to STAT3 activation. Revealing this novel mechanism of STAT3 activation by a miRNA offers new avenues for therapeutic interventions against colon cancer.
The scatter of 19 proteins and phosphoproteins with Q-value ,0.05.
Sa1692 Characterization of the Helicobacter pylori-Induced STAT3 Activation and the Associated Signaling Network in Gastric Cancer Junhong Zhao, Wei Kang, Yu Juan Dong, Minnie Y. Go, Joanna H. Tong, Ka Fai To, Alfred S. Cheng, Joseph J. Y. Sung, Jun Yu Background and Aims: Helicobacter pylori (H. pylori) is the most important gastric carcinogen. In this study, we sought to evaluate the effect of H. pylori infection on STAT3 activation and dissect the signaling network of STAT3 in the H. pylori-infected gastric cancer. Methods and results: The expression of the active form of STAT3, phospho-STAT3 (pSTAT3), was significantly higher in H. pylori-positive gastritis (73.5%, 61/83) than in H. pylori-negative gastritis (50%, 35/70) (P = 0.003) as shown by immunohistochemistry. On the other hand, therapy-based eradication of H. pylori significantly decreased pSTAT3 expression in 44 H. pylori-positive patients (P , 0.001). The association between H. pylori infection and pSTAT3 expression was further evaluated in mouse model with or without H. pylori infection (SS1 strain). Importantly, pSTAT3 was only detected in the H. pylori-infected gastric tissues of mice, but not in those of the control mice without H. pylori-infection, providing direct evidence that H. pylori infection stimulated the activation of STAT3 in the stomach. pSTAT3 expression was further detected in 147 human gastric cancers. pSTAT3 was marginally associated with the intestinal type than the diffuse type of gastric cancer ( P = 0.07). To elucidate the signaling network of STAT3 in the H. pylori-infected gastric cancer, the gastric cancer cell line AGS was co-cultured with two CagA+ve H. pylori strains (SS1 and ATCC43504) and one CagA-ve H. pylori strain, respectively. pSTAT3 expression was markedly induced in the ATCC43504 H. pyloriy-infected AGS as shown by Western blot. The signaling network of STAT3 induced by H. pylori was dissected using gene expression microarray. By comparing the gene expression profiles of H. pylori-infected and non-infected AGS cells by expression microarray, we identified a total of 849 genes with expression changes. KEGG pathway analysis showed that the de-regulated genes were enriched in 11 different cancer pathways including MAPK signaling pathway. Gene ontology analysis revealed significant up-regulation of genes with prominent roles in multiple cellular processes including signal transduction. Conclusions: H. pylori infection triggers the activation of STAT3 to de-regulate a multitude of tumorigenic genes which may contribute to the initiation and transformation of gastric carcinogenesis at early stage. Acknowledgment: The project was supported by a research fund (RFCID, 10090942).
The top signaling pathways (a) and metabolic pathways (b) altered in colorectal adenoma as determined by Ingenuity Pathway Analysis (IPA). Protein signaling network altered in colorectal adenoma (c). The network is displayed graphically as nodes (protein) and edges (the biological relationship between the nodes). The nodes are represented using various shapes that represent the functional class of the protein products. The up- and downregulated proteins in colorectal adenoma shaded in red and green, respectively. The nodes without color were not assessed in this study but identified by IPA as important nodes involved in the network. Sa1691 MiR-181b-1 Controls Glycolysis As STAT3 Activator in Colon Cancer Cells Xiaolin Pan, Jin Feng, Guoxin Zhang Cancer cells preferentially metabolize glucose through aerobic glycolysis. This phenomenon, known as the Warburg effect, is an anomalous characteristic of glucose metabolism in cancer cells. Signal transducer and activator of transcription factor 3 (STAT3) is constitutively activated in human malignancies, and served a crucial in cell survival, angiogenesis, immune evasion, inflammation and aerobic glycolysis. In this work, we show that protein inhibitor
AGA Abstracts
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Sa1687 SPARC Modulates Autophagy via Caspase 8 in the Presence of 5-Fluorouracil in Colorectal Cancer Mahbuba Rahman, Sharon Gorski, Isabella T. Tai Background and aims: Autophagy and apoptosis are two evolutionarily conserved processes regulating cellular fate in response to cytotoxic stress. SPARC, a matricellular protein, acts as a chemosensitizer in the presence of 5-fluorouracil (5FU) by activating the extrinsic apoptotic pathway protein caspase-8 in colorectal cancer (CRC). However, the functional relationship between autophagy and apoptosis in the presence of SPARC is largely unknown. Here we demonstrate that SPARC activation of caspase-8 in the presence of 5FU leads to a reduction in autophagic flux. Methods: The SPARC over-expressing colorectal cancer cell line MIP/SP and its empty vector control MIP/ZEO were treated with 5FU (5μM) for 72 hours. A MTT cell viability assay was performed to verify the effect of 5FU in the two cell lines. Apoptosis was assessed using TUNEL, the caspase 3/7 activity assay and western blot analysis of caspase-8 cleavage. Autophagosome accumulation was evaluated using LC3B immunofluorescence, and autophagic flux was assessed by western blot analysis of LC3BII in the presence and absence of Bafilomycin A1 (Baf). To investigate the role of SPARC or caspase-8 in autophagy modulation, the respective proteins were knocked down using SPARC-siRNA or caspase 8-siRNA. The effects on autophagy were monitored by the LC3BII flux assay and western blot analysis of autophagy proteins. Results: Treatment with 5FU resulted in reduced cell viability (MTT) in both cell lines, though was reduced significantly further in MIP/SP compared to MIP/ZEO. 5FU also led to increased TUNEL staining and reduced LC3B IF in both cell lines. However, the level of apoptosis (as assessed by casapse 8 and caspase 3/7 assay) was higher, and the reduction in autophagic flux appeared greater in the MIP/SP cell line compared to its control cell line MIP/ZEO. To investigate the potential role of SPARC in autophagy modulation, siRNA was used to knockdown the SPARC protein. Autophagic flux increased in both cell lines following SPARC knockdown. Concomitantly, the level of active caspase 8 and caspase 3/7 activity was reduced in MIP/SP even in the presence of 5FU. Knockdown of caspase 8 similarly resulted in a higher level of autophagy flux and also increased levels of ATG7 in the presence of 5FU in the MIP/SP cell line, but not in MIP/ZEO. Conclusion: These observations demonstrate that knockdown of caspase 8 in MIP/SP leads to elevated Atg7 and can partially rescue autophagic flux, suggesting that SPARC inhibits autophagy via a caspase 8-mediated mechanism. High levels of SPARC upregulate caspase 8, which not only affects the extrinsic pathway of apoptosis but also reduces autophagic flux. Our findings indicate that SPARC can augment chemosensitivity by affecting both the apoptosis and the autophagy pathways. Sa1688 Polyamine and Methionine Adenosyltransferase 2A Crosstalk in Human Colon Cancer Maria Lauda Tomasi, Minjung Ryoo, Anna Skay, Ivan Tomasi, Pasquale Giordano, Jose M. Mato, Shelly C. Lu ACKGROUND & AIMS: Colon cancer is one of the most common forms of cancer and a leading cause of cancer associated deaths in the United States. Methionine adenosyltransferase 2A (MAT2A) encodes for the enzyme that catalyzes the formation of S-adenosylmethionine (SAMe), the principal biological methyl donor and precursor of polyamines. We reported that MAT2A expression is increased in human colon cancer and in colon cancer cells treated with mitogens. Induction of MAT2A was required in order for several growth factors to exert the mitogenic effect. We also observed that silencing MAT2A resulted in apoptosis. The aim of the current work was to examine the mechanism responsible for MAT2Adependent growth and apoptosis. METHODS: Studies were conducted using human colon cancer cell line RKO and colon cancer specimens. Real-time PCR and Western blotting measured gene and protein expression, respectively. Expression of MAT2A and ornithine decarboxylase (ODC) was varied using siRNA or overexpression vector. RESULTS: In RKO cells, MAT2A siRNA treatment for 24 hours reduced MAT2A mRNA level by 75% and resulted in depletion of cellular SAMe and putrescine levels by 70 and 75%, respectively, increased apoptosis and growth suppression. Spermidine and spermine levels were unchanged at this time point but fell by 48 to 72 hours afterwards. Putrescine supplementation (100pmol/L) blunted significantly MAT2A siRNA-induced apoptosis and growth suppression. Putrescine treatment raised MAT2A mRNA level by 4.3-fold. In addition, putrescine treatment also increased the expression of AP-1 family members c-Jun and c-Fos, which are known to trans-activate the human MAT2A promoter and increase its expression. In human colon cancer specimens, the mRNA and protein levels of MAT2A, ODC, c-Jun and c-Fos are all elevated as compared to adjacent non-tumorous tissues. Overexpression of ODC (the ratelimiting enzyme in polyamine biosynthesis that catalyzes the formation of putrescine) in RKO cells also raised MAT2A mRNA level by 50%. CONCLUSIONS: There is a cross-talk
Sa1690 Proteomic and Metabolic Signaling Pathways in Colorectal Adenoma Miao cui, Liang Peng, Zhiqing Wang, Yuan Hu, Xinying Wang, Yu Zhang, Juan Wang, David Zhang, Bo Jiang, Huabao Xiong, Ming-Ming Zhou, Fei Ye Background Colorectal cancer (CRC) is the leading cause of cancer deaths worldwide. One million new cases are diagnosed each year around which lead to 500,000 deaths too. Colorectal adenoma is the precancerous lesions of colorectal cancers. An adenoma takes many years to progress to carcinoma and changes in both proteomic and metabolic pathways were observed during the progression. Detection of those adenomas progressing to in situ carcinoma provides a window of opportunity for removing localized lesions, thus cure. The aim of this study is to identify proteomic and metabolic signaling pathways activated in the colorectal adenoma to facilitate the understanding of underlying mechanism. Methods The expression of 27 metabolic proteins and 172 proteins of signaling pathway in 37 paired colorectal adenoma samples were assessed using Protein Pathway Array (PPA). The value of expression of phosphorylation was normalized by Z-score. Data were analyzed by Significance Analysis of Microarrays (SAM) to detect differentially expressed proteins. Both of proteins and samples are clustered in two-dimensional hierarchical cluster by MultiExperiment Viewer v4.8 (MeV). Principal Component Analysis (PCA) was performed using Partek Genomics
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AGA Abstracts
cases (8.7%) and was significantly correlated with MSI (p ,0.01). PTEN inactivation was present in 53 cases (24%) and was significantly correlated with deeper invasion (p=0.02). IGF1R was positive in 62 cases (29%) and was significantly correlated with intestinal histological type (p=0.03) and MSI (p=0.01). Although pAkt expression was only correlated with HER2 overexpression (p=0.005), pAkt expression was more frequent in cases with PIK3CA exon 20 mutations (100%) than in those with other PIK3CA mutations (50%). In multivariable analysis of prognostic factors, male, Stage 3 or 4, PTEN inactivation and pAkt expression were significantly correlated with poor prognosis. Among the molecular alterations analyzed in this study, 90 cases (43%) showed a single alteration and 32 cases (15%) showed two or more alterations. With increase in the number of alterations, frequency of pAkt expression tended to increase and prognosis tended to be worse. p95HER2 expression was positive in 13 (34%) of 38 HER2-positive cases. The 3-year survival rate of patients with p95HER2 positive was shorter than that of other patients (48% vs 68%). Conclusions: The number of alterations in each patient was significantly correlated with poor prognosis and PI3K-Akt pathway activation. Our findings indicate that there are some interactions among alterations in the PI3K-Akt pathway and that they could affect prognosis. We characterized p95HER2 expression in GC for the first time. This result supports the use of p95HER2 expression as a biomarker in a future clinical trial of Tmab for GC.
AGA Abstracts
Suite 6.0. The signaling pathways, metabolic pathways and network constituted by these proteins were identified by using Ingenuity Pathway Analysis (IPA). Results Of 27 metabolic proteins and 172 proteins of signaling pathway screened, 82 were detected in 37 paired colorectal adenoma samples and among which 19 were identified as significant proteins (Fig 1) by comparing between adenoma and normal tissues (Q-value ,0.05). Of 19 proteins, 18 were up-regulated in adenoma including MetRS, beta-catenin, p-cdc2 (Tyr15), p-PDK1 (Ser241), et al, while one phosphoprotein was down-regulated, i.e. p-ERK (Thr202/ Tyr204)(P44/42). Sixty four matched samples (37 adenoma and their matched normal mucosa) were used for hierarchical clustering analysis and these 19 proteins correctly grouped adenoma by PCA with 89% accuracy. The top signaling pathways (Fig 2 a) affected in colorectal adenoma were IL-3 pathway (-log(p-value)= 7.54), NF- κB pathway (-log(p-value)= 5.64), et al. The top metabolic pathways (Fig 2 b) affected in colorectal adenoma were Aldosterone pathway (-log(p-value)= 3.12), Docosahexaenoic Acid (DHA) pathway (-log(pvalue)= 2.94), et al. Conclusion Our current study demonstrated that many signaling pathways and metabolic pathways were affected in colorectal adenoma which may be involved in development of adenoma. Inflammatory pathways (IL-3, NF- κB and PI3K/AKT) and metabolic pathways (docosahexaenoic acid and leptin) are altered in colorectal adenoma, suggesting the critical role of these pathways.
of activated STAT3 (PIAS3) is the target of miR-181b-1, and further show that miR-181b1 promotes glycolysis by STAT3 activated in colon cancer cells, through down-regulating PIAS3. As STAT3 activation has been reported could promote the transcription and expression of miR-181b-1. Therefore, our results support a positive feedback loop as a mechanism for persistent STAT3 activation, in which miR-181b-1 represses PIAS3 to elevate STAT3 activity, which in turn promotes miR-181b-1 transcription. PIAS3 was found down-regulated and miR-181b-1 up-regulated in human colon tumors, and inverse correlation of miR-181b-1 and PIAS3 expression was also found in colon tumors. Moreover, miR-181b-1 inhibition or PIAS3 up-regulation in colon cancer cells led to reduce STAT3 activation, glycolysis and attenuated engraft tumour growth, indicating that miR-181b-1 overexpression contributes to STAT3 activation. Revealing this novel mechanism of STAT3 activation by a miRNA offers new avenues for therapeutic interventions against colon cancer.
The scatter of 19 proteins and phosphoproteins with Q-value ,0.05.
Sa1692 Characterization of the Helicobacter pylori-Induced STAT3 Activation and the Associated Signaling Network in Gastric Cancer Junhong Zhao, Wei Kang, Yu Juan Dong, Minnie Y. Go, Joanna H. Tong, Ka Fai To, Alfred S. Cheng, Joseph J. Y. Sung, Jun Yu Background and Aims: Helicobacter pylori (H. pylori) is the most important gastric carcinogen. In this study, we sought to evaluate the effect of H. pylori infection on STAT3 activation and dissect the signaling network of STAT3 in the H. pylori-infected gastric cancer. Methods and results: The expression of the active form of STAT3, phospho-STAT3 (pSTAT3), was significantly higher in H. pylori-positive gastritis (73.5%, 61/83) than in H. pylori-negative gastritis (50%, 35/70) (P = 0.003) as shown by immunohistochemistry. On the other hand, therapy-based eradication of H. pylori significantly decreased pSTAT3 expression in 44 H. pylori-positive patients (P , 0.001). The association between H. pylori infection and pSTAT3 expression was further evaluated in mouse model with or without H. pylori infection (SS1 strain). Importantly, pSTAT3 was only detected in the H. pylori-infected gastric tissues of mice, but not in those of the control mice without H. pylori-infection, providing direct evidence that H. pylori infection stimulated the activation of STAT3 in the stomach. pSTAT3 expression was further detected in 147 human gastric cancers. pSTAT3 was marginally associated with the intestinal type than the diffuse type of gastric cancer ( P = 0.07). To elucidate the signaling network of STAT3 in the H. pylori-infected gastric cancer, the gastric cancer cell line AGS was co-cultured with two CagA+ve H. pylori strains (SS1 and ATCC43504) and one CagA-ve H. pylori strain, respectively. pSTAT3 expression was markedly induced in the ATCC43504 H. pyloriy-infected AGS as shown by Western blot. The signaling network of STAT3 induced by H. pylori was dissected using gene expression microarray. By comparing the gene expression profiles of H. pylori-infected and non-infected AGS cells by expression microarray, we identified a total of 849 genes with expression changes. KEGG pathway analysis showed that the de-regulated genes were enriched in 11 different cancer pathways including MAPK signaling pathway. Gene ontology analysis revealed significant up-regulation of genes with prominent roles in multiple cellular processes including signal transduction. Conclusions: H. pylori infection triggers the activation of STAT3 to de-regulate a multitude of tumorigenic genes which may contribute to the initiation and transformation of gastric carcinogenesis at early stage. Acknowledgment: The project was supported by a research fund (RFCID, 10090942).
The top signaling pathways (a) and metabolic pathways (b) altered in colorectal adenoma as determined by Ingenuity Pathway Analysis (IPA). Protein signaling network altered in colorectal adenoma (c). The network is displayed graphically as nodes (protein) and edges (the biological relationship between the nodes). The nodes are represented using various shapes that represent the functional class of the protein products. The up- and downregulated proteins in colorectal adenoma shaded in red and green, respectively. The nodes without color were not assessed in this study but identified by IPA as important nodes involved in the network. Sa1691 MiR-181b-1 Controls Glycolysis As STAT3 Activator in Colon Cancer Cells Xiaolin Pan, Jin Feng, Guoxin Zhang Cancer cells preferentially metabolize glucose through aerobic glycolysis. This phenomenon, known as the Warburg effect, is an anomalous characteristic of glucose metabolism in cancer cells. Signal transducer and activator of transcription factor 3 (STAT3) is constitutively activated in human malignancies, and served a crucial in cell survival, angiogenesis, immune evasion, inflammation and aerobic glycolysis. In this work, we show that protein inhibitor
AGA Abstracts
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