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M2086 Comparative Proteomics Analysis to Identify Biomarkers of Hepatocellular Carcinoma
AGA Abstracts
and prognostic improvements for these patients. In this regard, γ2 chain of laminin-5 (LNγ2) and matrilysin (MMP7) are engaging targets for research. Preferential expression of the LNγ2 and matrilysin, both of which are targets of Wnt/β-catenin, in invading carcinoma cells has been implicated in the progression of other types of tumors. The aim of this study was to clarify the clinicopathological and functional significance of LNγ2 and matrilysin expression in biliary tract cancer. Expression of LNγ2 and matrilysin was analyzed by real-time RTPCR and immunostaining in cancer cell lines and carcinomas of the gallbladder (n = 19), extrahepatic bile duct (n = 29), and ampulla (n = 14). Zymography, in situ zymography, migration and invasion assays, and siRNA experiments were done. LNγ2 positivity in cancer cells at the invasive front was seen in 47% of gallbladder cancers, 69% of bile duct cancers, and 50% of ampullary cancers. LNγ2 positivity was significantly correlated with depth of invasion, histologic type, and advanced pTNM stage. Compared with matrilysin, LNγ2 was overexpressed more predominantly at the invasive front. Matrilysin positivity in cancer cells at the invasive front was seen in 50% of gallbladder cancers, 80% of bile duct cancers, and 71% of ampullary cancers. Expression of active matrilysin detected by zymography was significantly correlated with depth of invasion and advanced pTNM stage. There was a tendency that matrilysin expression was correlated with LNγ2 expression. Functional significance of these interactions was shown by various In Vitro analyses, including invasion assay. Our results suggest that LNγ2 and matrilysin play a key role in the progression of biliary tract cancer.
reported. It is unclear whether GPC3 needs to be processed by convertases for its activity. We analyzed whether GPC3 expression is correlated with expressions of specific matrix metalloproteinases (MMPs) and/or other proteases and Wnt signaling molecules. Expressions of GPC3, MMPs, and Wnt signaling molecules were analyzed in HCC cell lines and tissues by real-time RT-PCR and immunostaining. Activities of MMPs were analyzed by zymography. Serum GPC3 levels were analyzed by ELISA. In Vitro invasion assay was done for GPC3 siRNA-transfected HCC cell lines. GPC3 was overexpressed in most HCCs and its serum levels were significantly increased in a large proportion of patients with HCC. Overexpressions of various MMPs, such as MMP14 and MMP7, and Wnt signaling molecules, such as Wnt5A, were observed in HCCs. Some of these expressions were significantly correlated with GPC3 expression. Moreover, shedding of GPC3 by activated protease was found in HCC tissues. Functional significance of these interactions was shown by various In Vitro analyses, including invasion assay. These results suggest that GPC3, in conjunction with MMPs and Wnt signaling molecules, plays an important role in the progression of HCC. M2089 Difference in Presentation in Hepatocellular Carcinoma in Egyptian and US Cohorts Mohamed Amer, Ed Barber, Fatma Barakat, A. Ibrahim, M. Saleh, M. Salama, E. Rewish, Elliot Alpert, Khaled Selim, Yuko Kono, Tarek Hassanein
M2086
Background: The incidence of Hepatocellular Carcinoma (HCC) is increasing around the world particularly in areas where HCV and HBV infections are endemic. Early detection is crucial for effective intervention. The aim of the study is to explore if HCC presents differently in different parts of the world. Methods: We conducted a cross-sectional study of 2 cohorts of patients presenting with HCC in Egypt and the United States (US). 499 consecutive patients seen in the National Liver Institute in Egypt (Group A) and 128 patients seen at a tertiary Liver Center in the United States (Group B) were the subjects of the study. Results: Mean age of the Egyptian and US patients was 55±9 and 57±9 years respectively (p=0.018). Most cases occurred in males (81.5% in Group A and 75.7% in Group B). HCV infection was reported in 88.5% of patients in Group A and 75.8% in Group B (p=0.004). There was no difference between the 2 cohorts in severity of liver disease at presentation using Child-Turcotte-Pugh classification. However, the characteristics of the tumors were different between the 2 groups in: a) size of lesions (7.2±3.5 cm vs. 4.1±3.3 cm respectively, p<0.001); b) number of lesions (>= 3 lesions occurred in 28.4% vs. 12.5% respectively p<0.001); and c) presence of vascular invasion (20.6% vs. 7% p<0.001 respectively). Summary: In summary, the main cause of HCC in both cohorts was HCV infection. HCC presented in Egyptian patients a) at a younger age and b) in more advanced stage. These findings underscore the importance of preventing spread of HCV infection at a younger age, treating HCV infection and establishing universal screening programs for HCC.
Comparative Proteomics Analysis to Identify Biomarkers of Hepatocellular Carcinoma Marie Yeo, Young Mi Na, Dong-Kyu Kim, Hee Jin Park, Kwang Jae Lee, Sung Won Cho To facilitate the identification of candidate molecular biomarkers that are linked to the pathogenesis of hepatocellular carcinoma, we investigated protein expression profiles of 4 pairs of tumors and the adjacent non-tumor resected from HCC patients using by 2DE. 197 protein spots were found to be significantly different between the two groups. Four candidate biomarkers including methionin adenosyltransferase 1 (MAT1), sulfotransferase 1 (SULT1), apolipoprotein E (ApoE) and heat shock protein 27 (HSP27) showed a distinct expression pattern in hepatocellular carcinoma compared to the surrounding non-tumor tissues. We further verified the signature of the 4 key biomarkers in 21 pairs of HCC tissues by Western blotting and immunohistochemistry. Methionin adenosyltransferase 1 (MAT1) and sulfotransferase 1 (SULT1) were significantly down-regulated, whereas apolipoprotein E (ApoE) and heat shock protein 27 (HSP27) were up-regulated in HCC. The sensitivity and specificity of the individual biomarkers for the diagnosis of HCC were 76.2% and 95.2% for ApoE, 66.7% and 71.4% for HSP27, 95.2% and 71.4% for SULT1, and 57.1% and 85.7% for MAT1. When at least 2 of them were positive, the sensitivity and specificity were significantly improved to 100% and 90.5%. The present HCC proteome approach may be useful for identification of individual proteins which may offer a novel way to diagnose HCC.
M2090 Study of Ultrasonographic Image Using the New Contrast Agent Perflubutane in Patients with Metastatic Liver Tumors Hideki Hayashi, Youichi Nishigaki, Tomohiro Kato, Akihiko Sugiyama, Eiichi Tomita
M2087 The Expression Patterns of Growth Factors Are Different in Gallbladder Cancers According to the Gross Type Se-Joon Lee, Yonghan Paik, DongKi Lee, KwanSik Lee
(Background) The new contrast agent perflubutane shows excellent ultrasonographic pictures in real time and for a longer time than the older agent, galactose-palmitic acid. It has been reported that the image of metastatic tumors in liver have revealed hypovascularity. In this study, we investigated the blood flow of metastatic tumors in liver with ultrasonography using perflubutane. (Patients and methods) We performed ultrasonographies of 147 patients using perflubutane from January to July 2007 for diagnosis of liver tumors. Sixteen of these cases were diagnosed as metastatic liver tumors by enhanced CT or MRI. Of these, the primary tumors were colon cancer in 10 patients, gastric cancer in 3, lung cancer in 2, and urinary tract cancer in 1. The average tumor diameter was 21.7 mm (range: 10-63 mm), and the average depth from the skin was 44 mm (range: 23-75 mm). After bolus injection of perflubutane (0.015 ml/kg) into the forearm, the early vascular phase and late vascular phase were evaluated during the next 90 seconds using the pulse subtraction method with Aplio (Toshiba, Japan), the wideband pulse inversion method with EUB-8500 (Hitachi, Japan), or the pulse inversion and power modulation method with iU22 (Philips, USA). The post-vascular phase was evaluated 10 minutes after injection. MI was 0.2-0.3 and FR was 12-15 Hz. Additionally, we checked undyed central area of the tumors. (Results) In the early vascular phase, all 16 cases were dyed as hypervascular tumors. Of these, 9 were dyed in the peripheral area and the other 7 cases showed a dyed mosaic pattern. Although 12 cases that were shown to be hypovascular by CT or MRI were dyed hypervascular, they were found to be hypervascular by enhanced ultrasonography using perflubutane. In the post-vascular phase, all 16 cases showed clear avascular areas an average of 80 seconds after the injection. (Conclusion) Metastatic liver tumors viewed ultrasonographically using perflubutane are shown to be hypervascular. This new finding using the new contrast agent perflubutane may be helpful for more correct diagnoses of metastatic liver tumors. Further studies are needed.
Backgrounds: The expression of oncogene products and growth factors are involved in the carcinogenic process. The gallbladder (GB) cancer shows very poor prognosis and is more invasive and metastatic in infiltrative type than fungating type of GB cancer. However, there have been few reports addressing the relation of oncogenes and growth factors to the gross type of GB cancer. Aims: This study is conducted to evaluate the immunohistochemical expression of oncogenic products and growth factors according to the gross type of GB cancer. Methods: Seventy-five resected GB cancer specimens were used to immunohistochemical assay. The expression of tumor suppression gene products (p53) and growth factors (epidermal growth factor [EGF] and transforming growth factors-β [TGF-β]) was determined by immunohistochemical staining on paraffin embedded serial sections. Results: The mean age was 63.4 (range 39-94) years and 35 were males and 40 females. 62 cases were confirmed and could be classified as fungating type (30 cases) and infiltrating type (32 cases) by radiological, gross and pathologic findings. The other 13 cases were undetermined in gross type. The expression patterns of growth factors were 4 different types, all negative for growth factors were 18, EGF dominant 21, TGF-β dominant 19, equally expressed cases were 17. Expression of p53 protein was found in 57.3% (43/75) in GB cancer, but there was no difference between the expression of p53 protein and the growth factors. Proportions of positive stain for EGF and TGF-β in relation to the gross types of GB cancer were studied. EGF dominant cases were 21 (fungating type 17 vs. infiltrative type 4). TGF-β dominant cases were 19 cases (fungating type 3 vs. infiltrative type 16). There was a significant difference between the gross type and expression patterns of growth factors type (p<0.05). Conclusions: The difference between the gross type of the GB cancer and expression patterns of the growth factors may suspect that the different carcinogenic processes are involved according to the gross type of GB cancer.
T1023 M2088 Epidemiology of Opioid Bowel Dysfunction and Narcotic Bowel Syndrome in the Community Rok Seon Choung, G. Richard Locke, Cathy Schleck, Alan R. Zinsmeister, Nicholas J. Talley
Glypican-3, in Conjunction with MMPs and Wnt Signaling Molecules, Plays An Important Role in the Progression of Hepatocellular Carcinoma Chie Miyamoto, Hiroyuki Yamamoto, Shigeru Sasaki, Noriyuki Akutsu, Hiroaki Taniguchi, Nobuki Miyamoto, Katsuhiko Nosho, Tadateru Maehata, Kentaro Yamashita, Yasushi Adachi, Yoshiaki Arimura, Fumio Itoh, Kohzoh Imai, Yasuhisa Shinomura
Background: Opioids have long been recognized to affect gastrointestinal motility. Narcotic bowel syndrome (NBS) is defined as chronic abdominal pain with chronic narcotic use. Due to increasing narcotic use, NBS may be under-recognized and perhaps more prevalent. Aim: To assess whether bowel dysfunction is associated with chronic narcotic use and estimate the prevalence of narcotic bowel syndrome in the community. Methods: Validated selfreport GI symptom questionnaires were mailed to 4898 randomly selected people in the
Hepatocellular carcinoma (HCC) is one of the most prevalent cancers worldwide, and its incidence is still increasing. Glypican-3 (GPC3) is a member of the glypican family of GPI-anchored cell-surface heparan sulfate proteoglycans. Expression of GPC3 is frequently upregulated in HCCs. GPC3 reportedly promotes growth, migration, and invasiveness of HCC cells by stimulating canonical Wnt signaling, although conflicting results were also
AGA Abstracts
A-466
and prognostic improvements for these patients. In this regard, γ2 chain of laminin-5 (LNγ2) and matrilysin (MMP7) are engaging targets for research. Preferential expression of the LNγ2 and matrilysin, both of which are targets of Wnt/β-catenin, in invading carcinoma cells has been implicated in the progression of other types of tumors. The aim of this study was to clarify the clinicopathological and functional significance of LNγ2 and matrilysin expression in biliary tract cancer. Expression of LNγ2 and matrilysin was analyzed by real-time RTPCR and immunostaining in cancer cell lines and carcinomas of the gallbladder (n = 19), extrahepatic bile duct (n = 29), and ampulla (n = 14). Zymography, in situ zymography, migration and invasion assays, and siRNA experiments were done. LNγ2 positivity in cancer cells at the invasive front was seen in 47% of gallbladder cancers, 69% of bile duct cancers, and 50% of ampullary cancers. LNγ2 positivity was significantly correlated with depth of invasion, histologic type, and advanced pTNM stage. Compared with matrilysin, LNγ2 was overexpressed more predominantly at the invasive front. Matrilysin positivity in cancer cells at the invasive front was seen in 50% of gallbladder cancers, 80% of bile duct cancers, and 71% of ampullary cancers. Expression of active matrilysin detected by zymography was significantly correlated with depth of invasion and advanced pTNM stage. There was a tendency that matrilysin expression was correlated with LNγ2 expression. Functional significance of these interactions was shown by various In Vitro analyses, including invasion assay. Our results suggest that LNγ2 and matrilysin play a key role in the progression of biliary tract cancer.
reported. It is unclear whether GPC3 needs to be processed by convertases for its activity. We analyzed whether GPC3 expression is correlated with expressions of specific matrix metalloproteinases (MMPs) and/or other proteases and Wnt signaling molecules. Expressions of GPC3, MMPs, and Wnt signaling molecules were analyzed in HCC cell lines and tissues by real-time RT-PCR and immunostaining. Activities of MMPs were analyzed by zymography. Serum GPC3 levels were analyzed by ELISA. In Vitro invasion assay was done for GPC3 siRNA-transfected HCC cell lines. GPC3 was overexpressed in most HCCs and its serum levels were significantly increased in a large proportion of patients with HCC. Overexpressions of various MMPs, such as MMP14 and MMP7, and Wnt signaling molecules, such as Wnt5A, were observed in HCCs. Some of these expressions were significantly correlated with GPC3 expression. Moreover, shedding of GPC3 by activated protease was found in HCC tissues. Functional significance of these interactions was shown by various In Vitro analyses, including invasion assay. These results suggest that GPC3, in conjunction with MMPs and Wnt signaling molecules, plays an important role in the progression of HCC. M2089 Difference in Presentation in Hepatocellular Carcinoma in Egyptian and US Cohorts Mohamed Amer, Ed Barber, Fatma Barakat, A. Ibrahim, M. Saleh, M. Salama, E. Rewish, Elliot Alpert, Khaled Selim, Yuko Kono, Tarek Hassanein
M2086
Background: The incidence of Hepatocellular Carcinoma (HCC) is increasing around the world particularly in areas where HCV and HBV infections are endemic. Early detection is crucial for effective intervention. The aim of the study is to explore if HCC presents differently in different parts of the world. Methods: We conducted a cross-sectional study of 2 cohorts of patients presenting with HCC in Egypt and the United States (US). 499 consecutive patients seen in the National Liver Institute in Egypt (Group A) and 128 patients seen at a tertiary Liver Center in the United States (Group B) were the subjects of the study. Results: Mean age of the Egyptian and US patients was 55±9 and 57±9 years respectively (p=0.018). Most cases occurred in males (81.5% in Group A and 75.7% in Group B). HCV infection was reported in 88.5% of patients in Group A and 75.8% in Group B (p=0.004). There was no difference between the 2 cohorts in severity of liver disease at presentation using Child-Turcotte-Pugh classification. However, the characteristics of the tumors were different between the 2 groups in: a) size of lesions (7.2±3.5 cm vs. 4.1±3.3 cm respectively, p<0.001); b) number of lesions (>= 3 lesions occurred in 28.4% vs. 12.5% respectively p<0.001); and c) presence of vascular invasion (20.6% vs. 7% p<0.001 respectively). Summary: In summary, the main cause of HCC in both cohorts was HCV infection. HCC presented in Egyptian patients a) at a younger age and b) in more advanced stage. These findings underscore the importance of preventing spread of HCV infection at a younger age, treating HCV infection and establishing universal screening programs for HCC.
Comparative Proteomics Analysis to Identify Biomarkers of Hepatocellular Carcinoma Marie Yeo, Young Mi Na, Dong-Kyu Kim, Hee Jin Park, Kwang Jae Lee, Sung Won Cho To facilitate the identification of candidate molecular biomarkers that are linked to the pathogenesis of hepatocellular carcinoma, we investigated protein expression profiles of 4 pairs of tumors and the adjacent non-tumor resected from HCC patients using by 2DE. 197 protein spots were found to be significantly different between the two groups. Four candidate biomarkers including methionin adenosyltransferase 1 (MAT1), sulfotransferase 1 (SULT1), apolipoprotein E (ApoE) and heat shock protein 27 (HSP27) showed a distinct expression pattern in hepatocellular carcinoma compared to the surrounding non-tumor tissues. We further verified the signature of the 4 key biomarkers in 21 pairs of HCC tissues by Western blotting and immunohistochemistry. Methionin adenosyltransferase 1 (MAT1) and sulfotransferase 1 (SULT1) were significantly down-regulated, whereas apolipoprotein E (ApoE) and heat shock protein 27 (HSP27) were up-regulated in HCC. The sensitivity and specificity of the individual biomarkers for the diagnosis of HCC were 76.2% and 95.2% for ApoE, 66.7% and 71.4% for HSP27, 95.2% and 71.4% for SULT1, and 57.1% and 85.7% for MAT1. When at least 2 of them were positive, the sensitivity and specificity were significantly improved to 100% and 90.5%. The present HCC proteome approach may be useful for identification of individual proteins which may offer a novel way to diagnose HCC.
M2090 Study of Ultrasonographic Image Using the New Contrast Agent Perflubutane in Patients with Metastatic Liver Tumors Hideki Hayashi, Youichi Nishigaki, Tomohiro Kato, Akihiko Sugiyama, Eiichi Tomita
M2087 The Expression Patterns of Growth Factors Are Different in Gallbladder Cancers According to the Gross Type Se-Joon Lee, Yonghan Paik, DongKi Lee, KwanSik Lee
(Background) The new contrast agent perflubutane shows excellent ultrasonographic pictures in real time and for a longer time than the older agent, galactose-palmitic acid. It has been reported that the image of metastatic tumors in liver have revealed hypovascularity. In this study, we investigated the blood flow of metastatic tumors in liver with ultrasonography using perflubutane. (Patients and methods) We performed ultrasonographies of 147 patients using perflubutane from January to July 2007 for diagnosis of liver tumors. Sixteen of these cases were diagnosed as metastatic liver tumors by enhanced CT or MRI. Of these, the primary tumors were colon cancer in 10 patients, gastric cancer in 3, lung cancer in 2, and urinary tract cancer in 1. The average tumor diameter was 21.7 mm (range: 10-63 mm), and the average depth from the skin was 44 mm (range: 23-75 mm). After bolus injection of perflubutane (0.015 ml/kg) into the forearm, the early vascular phase and late vascular phase were evaluated during the next 90 seconds using the pulse subtraction method with Aplio (Toshiba, Japan), the wideband pulse inversion method with EUB-8500 (Hitachi, Japan), or the pulse inversion and power modulation method with iU22 (Philips, USA). The post-vascular phase was evaluated 10 minutes after injection. MI was 0.2-0.3 and FR was 12-15 Hz. Additionally, we checked undyed central area of the tumors. (Results) In the early vascular phase, all 16 cases were dyed as hypervascular tumors. Of these, 9 were dyed in the peripheral area and the other 7 cases showed a dyed mosaic pattern. Although 12 cases that were shown to be hypovascular by CT or MRI were dyed hypervascular, they were found to be hypervascular by enhanced ultrasonography using perflubutane. In the post-vascular phase, all 16 cases showed clear avascular areas an average of 80 seconds after the injection. (Conclusion) Metastatic liver tumors viewed ultrasonographically using perflubutane are shown to be hypervascular. This new finding using the new contrast agent perflubutane may be helpful for more correct diagnoses of metastatic liver tumors. Further studies are needed.
Backgrounds: The expression of oncogene products and growth factors are involved in the carcinogenic process. The gallbladder (GB) cancer shows very poor prognosis and is more invasive and metastatic in infiltrative type than fungating type of GB cancer. However, there have been few reports addressing the relation of oncogenes and growth factors to the gross type of GB cancer. Aims: This study is conducted to evaluate the immunohistochemical expression of oncogenic products and growth factors according to the gross type of GB cancer. Methods: Seventy-five resected GB cancer specimens were used to immunohistochemical assay. The expression of tumor suppression gene products (p53) and growth factors (epidermal growth factor [EGF] and transforming growth factors-β [TGF-β]) was determined by immunohistochemical staining on paraffin embedded serial sections. Results: The mean age was 63.4 (range 39-94) years and 35 were males and 40 females. 62 cases were confirmed and could be classified as fungating type (30 cases) and infiltrating type (32 cases) by radiological, gross and pathologic findings. The other 13 cases were undetermined in gross type. The expression patterns of growth factors were 4 different types, all negative for growth factors were 18, EGF dominant 21, TGF-β dominant 19, equally expressed cases were 17. Expression of p53 protein was found in 57.3% (43/75) in GB cancer, but there was no difference between the expression of p53 protein and the growth factors. Proportions of positive stain for EGF and TGF-β in relation to the gross types of GB cancer were studied. EGF dominant cases were 21 (fungating type 17 vs. infiltrative type 4). TGF-β dominant cases were 19 cases (fungating type 3 vs. infiltrative type 16). There was a significant difference between the gross type and expression patterns of growth factors type (p<0.05). Conclusions: The difference between the gross type of the GB cancer and expression patterns of the growth factors may suspect that the different carcinogenic processes are involved according to the gross type of GB cancer.
T1023 M2088 Epidemiology of Opioid Bowel Dysfunction and Narcotic Bowel Syndrome in the Community Rok Seon Choung, G. Richard Locke, Cathy Schleck, Alan R. Zinsmeister, Nicholas J. Talley
Glypican-3, in Conjunction with MMPs and Wnt Signaling Molecules, Plays An Important Role in the Progression of Hepatocellular Carcinoma Chie Miyamoto, Hiroyuki Yamamoto, Shigeru Sasaki, Noriyuki Akutsu, Hiroaki Taniguchi, Nobuki Miyamoto, Katsuhiko Nosho, Tadateru Maehata, Kentaro Yamashita, Yasushi Adachi, Yoshiaki Arimura, Fumio Itoh, Kohzoh Imai, Yasuhisa Shinomura
Background: Opioids have long been recognized to affect gastrointestinal motility. Narcotic bowel syndrome (NBS) is defined as chronic abdominal pain with chronic narcotic use. Due to increasing narcotic use, NBS may be under-recognized and perhaps more prevalent. Aim: To assess whether bowel dysfunction is associated with chronic narcotic use and estimate the prevalence of narcotic bowel syndrome in the community. Methods: Validated selfreport GI symptom questionnaires were mailed to 4898 randomly selected people in the
Hepatocellular carcinoma (HCC) is one of the most prevalent cancers worldwide, and its incidence is still increasing. Glypican-3 (GPC3) is a member of the glypican family of GPI-anchored cell-surface heparan sulfate proteoglycans. Expression of GPC3 is frequently upregulated in HCCs. GPC3 reportedly promotes growth, migration, and invasiveness of HCC cells by stimulating canonical Wnt signaling, although conflicting results were also
AGA Abstracts
A-466