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Tumor doubling time and vascularity predict prognosis of patients with small hepatocellular carcinoma in Japan
Category 4: Hepatocellular carcinoma, liver regeneration, apoptosis - ~ H G F MODIFIES BUT DOES NOT INHIBIT THE HEPATOCARCINOGENESIS INDUCED BY DIETHYLNITROSAMINE IN TRANSGENIC MOUSE MODELS Andras Kiss l, Valentina Factor 2, Pal Kaposi Novak I , Elisabeth Conner2, Zsuzsa SchaffI , Snorri Thorgeirsson 2. 12nd Inst. of Pathology,
Semmelweis Medical University, Budapest, Hungary; 2Nat. Cancer Inst. NIH, Bethesda, MD, USA Background: Experimental data suggest that HGF plays a role as a homeostatic modulator in liver carcinogenesis. Sustained overexpression of HGF prevented malignant conversion of preneoplastic foci in HGF/c-myc double transgenic mouse model. Moreover, HGF completely inhibited the tumor promotion by phenobarbital. Aim: The objective of the present study was to characterize the effect of HGF on diethylnitrosoamine (DEN) induced hepatocarcinogenesis in HGE HGF/c-myc and c-myc transgenic animal models. Methods: Homozygous HGE c-myc and HGF/c-myc double transgenic mice were treated with DEN and followed for 6 months after treatment. The expression of transgenes and their receptors were identified by Northern blotting and PCR. Pathological alterations were diagnosed by histology. Results: HGF and c-myc transgenes were expressed continuously. The liver/body weight ratio after DEN treatment was significantly lower in HGF homozygous mice compared to c-myc mice at four month. Tissue expression of TGF-beta-RII and c-met checked by immunohistocbemistry did not show major alteration in the foci and tumors of different transgenic lines compared to the surrounding tissue. Similarly, the expression of bog did not reveal major differences among transgenic lines. Interestingly, human HCC samples showed downregulation of c-met in one third of cases and downregulation of TGF-beta RII in fifty percent of cases. Large dysplastic cells, however, showed lack of TGF-beta RII expression. Characteristicly, only the tumors of the DEN treated c-myc mice contained areas of oval cell proliferation. Conclusion: Taken together, our data suggest that simultaneous overexpression of HGF modifies, however, does not inhibit the DEN induced hepatocarcinogenesis driven by c-myc.
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TUMOR DOUBLING TIME AND VASCULARITY PREDICT PROGNOSIS OF PATIENTS WITH SMALL HEPATOCELLULAR CARCINOMA IN JAPAN
Ryoko Kuromatsu, Masatoshi Tanaka, Eiji Ando, Michio Sata. Dept. of
Medicine II, Kurume University School of Medicine, Japan Aim: It is important for the selection of the treatment of hepatoceUular carcinoma (HCC) to know the natural history of HCC. We report the analysis of the history of small HCC in Japan. Patients and Methods: Forty-nine patients with HCC less than 3 cm in diameter and less than 3 nodules (mean age; 70 years old, men/women 34/25, tumor stage T1/T2/T3; 19/27/3, Child-Pugh grade A/B/C; 16/18/15) were enrolled and followed regularly. The survival rates and the prognostic factors (tumor size, tumor number, tumor vascularity (hyper/hypo), ultrasonic morphology (low/high), tumor doubling time, tumor stage, Child-Pugh grade) were analyzed. Results: 1) Child-Pugh grade was the most important factor for the prognosis in 49 patients. Almost all patients with Child-Pugh grade C (15/16) died within 2 years. Then, we analyzed the prognosis factors in 34 patients with Child-Pugh grade A and B. 2) Three- and 5-year survival rates in 34 patients with Child-Pugh grade A and B were 58% and 29%, respectively. The prognostic factors using Cox's proportional hazard model were tumor doubling time (p = 0.0016), tumor vascularity (p = 0.0316), and ultrasonic morphology (p = 0.0309). Tumor size (p -- 0.3439) and tumor number (p -0.5117) were not significant for the prognosis. Conclusions: From these results, we suggested the following; 1) a treatment of HCC is not necessary in patients with Child-Pugh grade C. 2) the factors showing the malignant potentials, such as doubling time and vascularity, were more important to the prognosis than tumor size and number. In the analysis of the prognosis of small HCC treated, tumor vascularity and doubling time should be considered as the prognostic factors.
[-~THE ANTI-APOPTOTIC AND PRO-PROLIFERATIVE p53-RELATED DN-p73 PROTEIN IS EXPRESSED IN HUMAN HCC
MASSIVE INDUCED BY INTERLEUKIN-6 • 2 - OVEREXPRESSION - • LIVER GROWTH IN MICE
Emanuele Palescandolo l , Antonio Costanzo l , Stefania Vossio I , Natalia Pediconi 1, Paola Merlo 1, Cristiana Almerighil, Man:ella Fulco 1, Elisabetta Cariani 2, Clara Balsano 1,3, Massimo Levrero 1,4. l Fondazione
Teresa Zimmers, Iain McKillop, Robert Pierce, J.Y. Yoo, Patricia Murtha-Riel, Leonidas Koniaris. Department of Surgery,
Andrea Cesalpino, University of Rome 'La Sapienza', Rome; 21II Laboratory, Spedali Civili, Brescia; 3Dept. Internal Medicine, University of L'Aquila, L'Aquila; 4Dept. Internal Medicine, University of Cagliari, Cagliari, Italy
University of Rochester School of Medicine and Dentistry, Rochester, NY, USA Mice deficient in the multifunctional cytokine interleukin-6 (IL-6) display abnormal and delayed liver regeneration and repair, suggesting a role for IL-6 in promoting liver growth. Hyperstimulation with IL-6 slows liver regeneration, however, and elevated IL-6 is observed in chronic liver injury, suggesting that IL-6 may actually inhibit liver growth. We investigated the growth regulatory properties of IL-6 in vivo by growing CHO tumors overexpessing human IL-6 in nude mice. CHO-IL-6 tumor-bearing mice developed dramatic hepatomegaly and hepatocyte hyperplasia in the absence of hepatic injury. Increased liver mass was noted at six days and by days 12 to 15 livers were two to three times larger than those from mice bearing control tumors. Liver growth was accompanied by high levels of serum human IL-6 and activation of the IL-6 signaling pathway, including increased expression of IL-6 receptor alpha/gp80, activation of the STAT-3 and MAP kinase signaling pathways and expression of downstream target genes. Furthermore, hepatocyte growth factor and epidermal growth factor receptor signaling were decreased in hypertrophied livers, suggesting that IL-6 induced liver growth may be independent of these known hepatocyte mitogens. Taken together, these results suggest that IL-6 can function as a complete hepatic mitogen in vivo.
Several genetic and epigenetic alterations affecting the pRb, p53, TGF-beta and beta-catenin pathways have been described in human HCCs. Searching for new candidate HCC genes, we have characterizez the expression and function of the p53-related P73 gene. p73 is expressed as multiple COOH-terminus splice variants and a dominant negative (DN) variant, acting as transrepressors of p53 and p73 dependent transcription, has been described. P73, differently from P53, is rarely mutated in cancer suggesting that p73 deregulated expression rather than loss of function may contribute to cell transformation, p73 is indeed overexpressed in many HCCs. We have studied p73 expression at the RNA and protein level in HCC cell lines and in human HBV- and HCV-related HCCs. The pattern of p73 COOHsplicing isoforms expressed is variable and complex, with detectable levels of the shorter isoforms, as already described in myeloid leukemias. In addition, both DN-p73 transcripts (RT-PCR) and proteins (immunoblotting and immunohistochemistry) are largely expressed. We also cloned DN-p73 alternative promoter (P2p73). Differently from the Plp73 promoter that directs the expression of TA-p73 proteins, the P2p73 promoter is activated by TNF-a and downregulated by retinoids. Functional studies showed that the tumor derived DN-p73 blocks p53 and TA-p73 dependent transcription, counteract both p53 and TA-p73 growth suppression and apoptosis and protects cells from DNA damage induced apoptosis. In conclusion we have identified a p73 variant that trans-represses wild type p53 and TA-p73
Semmelweis Medical University, Budapest, Hungary; 2Nat. Cancer Inst. NIH, Bethesda, MD, USA Background: Experimental data suggest that HGF plays a role as a homeostatic modulator in liver carcinogenesis. Sustained overexpression of HGF prevented malignant conversion of preneoplastic foci in HGF/c-myc double transgenic mouse model. Moreover, HGF completely inhibited the tumor promotion by phenobarbital. Aim: The objective of the present study was to characterize the effect of HGF on diethylnitrosoamine (DEN) induced hepatocarcinogenesis in HGE HGF/c-myc and c-myc transgenic animal models. Methods: Homozygous HGE c-myc and HGF/c-myc double transgenic mice were treated with DEN and followed for 6 months after treatment. The expression of transgenes and their receptors were identified by Northern blotting and PCR. Pathological alterations were diagnosed by histology. Results: HGF and c-myc transgenes were expressed continuously. The liver/body weight ratio after DEN treatment was significantly lower in HGF homozygous mice compared to c-myc mice at four month. Tissue expression of TGF-beta-RII and c-met checked by immunohistocbemistry did not show major alteration in the foci and tumors of different transgenic lines compared to the surrounding tissue. Similarly, the expression of bog did not reveal major differences among transgenic lines. Interestingly, human HCC samples showed downregulation of c-met in one third of cases and downregulation of TGF-beta RII in fifty percent of cases. Large dysplastic cells, however, showed lack of TGF-beta RII expression. Characteristicly, only the tumors of the DEN treated c-myc mice contained areas of oval cell proliferation. Conclusion: Taken together, our data suggest that simultaneous overexpression of HGF modifies, however, does not inhibit the DEN induced hepatocarcinogenesis driven by c-myc.
~-7
81
TUMOR DOUBLING TIME AND VASCULARITY PREDICT PROGNOSIS OF PATIENTS WITH SMALL HEPATOCELLULAR CARCINOMA IN JAPAN
Ryoko Kuromatsu, Masatoshi Tanaka, Eiji Ando, Michio Sata. Dept. of
Medicine II, Kurume University School of Medicine, Japan Aim: It is important for the selection of the treatment of hepatoceUular carcinoma (HCC) to know the natural history of HCC. We report the analysis of the history of small HCC in Japan. Patients and Methods: Forty-nine patients with HCC less than 3 cm in diameter and less than 3 nodules (mean age; 70 years old, men/women 34/25, tumor stage T1/T2/T3; 19/27/3, Child-Pugh grade A/B/C; 16/18/15) were enrolled and followed regularly. The survival rates and the prognostic factors (tumor size, tumor number, tumor vascularity (hyper/hypo), ultrasonic morphology (low/high), tumor doubling time, tumor stage, Child-Pugh grade) were analyzed. Results: 1) Child-Pugh grade was the most important factor for the prognosis in 49 patients. Almost all patients with Child-Pugh grade C (15/16) died within 2 years. Then, we analyzed the prognosis factors in 34 patients with Child-Pugh grade A and B. 2) Three- and 5-year survival rates in 34 patients with Child-Pugh grade A and B were 58% and 29%, respectively. The prognostic factors using Cox's proportional hazard model were tumor doubling time (p = 0.0016), tumor vascularity (p = 0.0316), and ultrasonic morphology (p = 0.0309). Tumor size (p -- 0.3439) and tumor number (p -0.5117) were not significant for the prognosis. Conclusions: From these results, we suggested the following; 1) a treatment of HCC is not necessary in patients with Child-Pugh grade C. 2) the factors showing the malignant potentials, such as doubling time and vascularity, were more important to the prognosis than tumor size and number. In the analysis of the prognosis of small HCC treated, tumor vascularity and doubling time should be considered as the prognostic factors.
[-~THE ANTI-APOPTOTIC AND PRO-PROLIFERATIVE p53-RELATED DN-p73 PROTEIN IS EXPRESSED IN HUMAN HCC
MASSIVE INDUCED BY INTERLEUKIN-6 • 2 - OVEREXPRESSION - • LIVER GROWTH IN MICE
Emanuele Palescandolo l , Antonio Costanzo l , Stefania Vossio I , Natalia Pediconi 1, Paola Merlo 1, Cristiana Almerighil, Man:ella Fulco 1, Elisabetta Cariani 2, Clara Balsano 1,3, Massimo Levrero 1,4. l Fondazione
Teresa Zimmers, Iain McKillop, Robert Pierce, J.Y. Yoo, Patricia Murtha-Riel, Leonidas Koniaris. Department of Surgery,
Andrea Cesalpino, University of Rome 'La Sapienza', Rome; 21II Laboratory, Spedali Civili, Brescia; 3Dept. Internal Medicine, University of L'Aquila, L'Aquila; 4Dept. Internal Medicine, University of Cagliari, Cagliari, Italy
University of Rochester School of Medicine and Dentistry, Rochester, NY, USA Mice deficient in the multifunctional cytokine interleukin-6 (IL-6) display abnormal and delayed liver regeneration and repair, suggesting a role for IL-6 in promoting liver growth. Hyperstimulation with IL-6 slows liver regeneration, however, and elevated IL-6 is observed in chronic liver injury, suggesting that IL-6 may actually inhibit liver growth. We investigated the growth regulatory properties of IL-6 in vivo by growing CHO tumors overexpessing human IL-6 in nude mice. CHO-IL-6 tumor-bearing mice developed dramatic hepatomegaly and hepatocyte hyperplasia in the absence of hepatic injury. Increased liver mass was noted at six days and by days 12 to 15 livers were two to three times larger than those from mice bearing control tumors. Liver growth was accompanied by high levels of serum human IL-6 and activation of the IL-6 signaling pathway, including increased expression of IL-6 receptor alpha/gp80, activation of the STAT-3 and MAP kinase signaling pathways and expression of downstream target genes. Furthermore, hepatocyte growth factor and epidermal growth factor receptor signaling were decreased in hypertrophied livers, suggesting that IL-6 induced liver growth may be independent of these known hepatocyte mitogens. Taken together, these results suggest that IL-6 can function as a complete hepatic mitogen in vivo.
Several genetic and epigenetic alterations affecting the pRb, p53, TGF-beta and beta-catenin pathways have been described in human HCCs. Searching for new candidate HCC genes, we have characterizez the expression and function of the p53-related P73 gene. p73 is expressed as multiple COOH-terminus splice variants and a dominant negative (DN) variant, acting as transrepressors of p53 and p73 dependent transcription, has been described. P73, differently from P53, is rarely mutated in cancer suggesting that p73 deregulated expression rather than loss of function may contribute to cell transformation, p73 is indeed overexpressed in many HCCs. We have studied p73 expression at the RNA and protein level in HCC cell lines and in human HBV- and HCV-related HCCs. The pattern of p73 COOHsplicing isoforms expressed is variable and complex, with detectable levels of the shorter isoforms, as already described in myeloid leukemias. In addition, both DN-p73 transcripts (RT-PCR) and proteins (immunoblotting and immunohistochemistry) are largely expressed. We also cloned DN-p73 alternative promoter (P2p73). Differently from the Plp73 promoter that directs the expression of TA-p73 proteins, the P2p73 promoter is activated by TNF-a and downregulated by retinoids. Functional studies showed that the tumor derived DN-p73 blocks p53 and TA-p73 dependent transcription, counteract both p53 and TA-p73 growth suppression and apoptosis and protects cells from DNA damage induced apoptosis. In conclusion we have identified a p73 variant that trans-represses wild type p53 and TA-p73