Bazarragchaa Damdinsuren

Treatment of hepatocellular carcinoma with major portal vein thrombosis by combined therapy with subcutaneous interferon-alpha and intra-arterial 5-fluorouracil; role of type 1 interferon receptor expression.

We previously reported the beneficial effects of combination therapy of interferon (IFN)-α/5-fluorouracil (FU) for advanced hepatocellular carcinoma (HCC) with tumour thrombi in the major portal branches. This report describes the results of longer follow-up and includes more than double the number of patients relative to the original report, and evaluates the role of IFN-α/type 2 interferon receptor (IFNAR2) expression on the response to the combination therapy. The study subjects were 55 patients with advanced HCC and tumour thrombi in the major branches of the portal vein (Vp3 or 4). They were treated with at least two courses of IFN-α/5-FU without major complication. In the 55 patients, 24 (43.6%) showed objective response (eight (14.5%) showed complete response, 16 (29.1%) partial response), four (7.3%) showed no response, and 27 (49.1%) showed progressive disease. Immunohistochemically, IFNAR2 expression was detected in nine out of 13 (69.2%) patients. There was significant difference in the time-to-progression survival (P=0.0002) and the overall survival (P<0.0001) between IFNAR2-positive and -negative cases. There was a significant correlation between IFNAR2 expression and response to IFN-α/5-FU combination therapy in univariate analysis (P=0.0070). IFN-α/5-FU combination therapy is a promising modality for advanced HCC with tumour thrombi in the major portal branches and could significantly depend on IFNAR2 expression.

JTE-522, a cyclooxygenase-2 inhibitor, is an effective chemopreventive agent against rat experimental liver fibrosis1 ☆

BACKGROUND & AIMS The aim of this study was to assess the effects of cyclooxygenase (COX)-2 inhibition on rat experimental liver fibrogenesis. METHODS We investigated the inhibitory effects of a selective COX-2 inhibitor, JTE-522, on liver fibrosis induced by a choline-deficient, l-amino acid-defined diet (CDAA). Inhibitory effect was also tested in a second model of thioacetamide (TAA)-induced liver fibrosis. RESULTS CDAA induced liver fibrosis and preneoplastic foci at 12 weeks and cirrhosis at 36 weeks. Hepatocellular carcinoma was noted in 13 of 15 rats (87%). JTE-522 significantly inhibited fibrosis and development of preneoplastic lesions in a dose-dependent manner and completely inhibited generation of cirrhosis and hepatocellular carcinoma at both low and high doses (10 and 30 mg/kg body wt/day, respectively). JTE-522 administrated only from 12 weeks to 36 weeks also prevented cirrhosis and formation of hepatocellular carcinoma. JTE-522 itself did not cause local or systemic gross or histopathologic changes at 36 weeks. Mechanistic studies indicated that the CDAA model displayed up-regulation of several biomarkers, including COX-2, arachidonate metabolite (prostaglandin E(2)), serum aspartate aminotransferase, and c-myc expression. The model also showed an increased proportion of activated hepatic stellate cells, proliferating cell nuclear antigen index, and CD45-positive inflammatory cells in the liver. JTE-522 effectively diminished these changes. JTE-522 exhibited similar antifibrosis effects in the TAA model. CONCLUSIONS Our results suggest that COX-2 is involved in CDAA- and TAA-induced liver fibrosis. Our data also indicate that JTE-522 is a potent chemopreventive agent of rat liver fibrosis with low toxicity.

Expression pattern of angiogenic factors and prognosis after hepatic resection in hepatocellular carcinoma: importance of angiopoietin‐2 and hypoxia‐induced factor‐1a

Abstract: Background: Hepatocellular carcinoma (HCC) is a hypervascular tumor and angiogenesis plays an important role in its progression. Angiogenesis is regulated by a balance between pro and antiangiogenic molecules. The aim of this study was to investigate the expressions of angiogenic factors and elucidate their roles in angiogenesis in HCC.

Partial Contribution of Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)/TRAIL Receptor Pathway to Antitumor Effects of Interferon-α/5-Fluorouracil against Hepatocellular Carcinoma

Purpose: Our purpose was to explore the contribution of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/TRAIL receptor pathway to antitumor effects of IFNα and 5-fluorouracil (5-FU) combination therapy for hepatocellular carcinoma (HCC). Experimental Design: Susceptibility of HCC cell lines to TRAIL and/or 5-FU was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The effects of 5-FU, IFNα, or both on the expression of TRAIL receptors (R1, R2, R3, and R4) on HCC cells or TRAIL in peripheral blood mononuclear cells (PBMC) were examined by flow cytometry. IFNα-induced cytotoxic effects of PBMC on HCC cell lines were examined by 51Cr release assay. TRAIL expression in peripheral blood mononuclear cells and liver tissue from patients was examined by real-time reverse transcription-PCR or immunohistochemistry. Results: HLE and HepG2 were sensitive to TRAIL, but HuH7, PLC/PRF/5, and HLF were resistant. 5-FU had synergistic effect on TRAIL in HLF and additive effect in four other HCC cell lines. TRAIL receptors on HCC cells were up-regulated by 5-FU, and IFNα induced TRAIL on CD4+ T cells, CD14+ monocytes, and CD56+ NK cells. Treatment of effector cells by IFNα and target HCC cells by 5-FU enhanced the cytotoxicity of CD14+ monocytes and CD56+ NK cells against HCC cells via a TRAIL-mediated pathway. TRAIL mRNA overexpression was noted in PBMC of HCC patients who clinically responded to IFNα/5-FU combination therapy, and TRAIL+ mononuclear cells were found in cancer tissue of a responder. Conclusion: Our results suggest that modulation of TRAIL/TRAIL receptor-mediated cytotoxic pathway might partially contribute to the anti-HCC effect of IFNα and 5-FU combination therapy.

Interferon-β Is More Potent Than Interferon-α in Inhibition of Human Hepatocellular Carcinoma Cell Growth When Used Alone and in Combination With Anticancer Drugs

AbstractBackground:The prognosis of advanced hepatocellular carcinoma (HCC) is extremely poor, but promising effects of chemotherapies combined with interferon (IFN) have been reported. Methods:To develop more effective combination therapies for HCC, we compared the antiproliferative effects of IFN-α and IFN-β in combination with various cytotoxic drugs on hepatoma cell lines using MTT assay and isobologram analysis. Results:IFN-β was more potent than IFN-α in inhibiting the cell growth of all cell lines (P < .05, two-way ANOVA). PLC/PRF/5 was more sensitive to either IFN, than HLE and HuH7. Cell growth of all cell lines was inhibited in a dose-dependent manner by 5-fluorouracil (5-FU), cisplatin (CDDP), and doxorubicin (DOX), but the sensitivities of these cells were considerably different. As for IFN-α, synergistic effects were observed when combined with 5-FU and DOX on PLC/PRF/5 cells only, whereas IFN-β showed synergistic effects with 5-FU and CDDP on HuH7 and PLC/PRF/5 cell lines. Conclusion:The spectra of the antiproliferative activity and synergistic effect of IFN-β when combined with anticancer drugs are more potent than those of IFN-α. Combinations of IFN-β and anticancer drugs may provide a better treatment of HCC when combinations with IFN-α are ineffective.

Antisense to Cyclin D1 Inhibits Vascular Endothelial Growth Factor–Stimulated Growth of Vascular Endothelial Cells: Implication of Tumor Vascularization

Purpose: Our aim was to determine the effects of cyclin D1 inhibition on tumor-associated neovascularization and endothelial cell growth. Experimental Design: We have generated adenovirus system for antisense to cyclin D1 (AS CyD1) and evaluated in vitro and in vivo effects. Small interfering RNA against cyclin D1 was also used to analyze cyclin D1 inhibition-associated vascular endothelial growth factor (VEGF) regulation. Results: The xenografts treated with adenoviral AS CyD1 showed less vessel density and displayed smaller tumor size in colon cancer cell lines HCT116 and DLD1. In vitro studies indicated that AS CyD1 decreased VEGF protein expression in DLD1 but not in HCT116. Cyclin D1 small interfering RNA caused a decrease in VEGF expression at protein and RNA levels in DLD1. A modest decrease was noted in the VEGF promoter activity, with inactivation of the STAT3 transcription factor through dephosphorylation. On the hand, the cyclin D1 inhibition plus STAT3 inhibitor markedly decreased VEGF expression in HCT116, although VEGF did not change by the STAT3 inhibitor alone. In cultures of human umbilical vein endothelial cells (HUVEC), VEGF augmented cyclin D1 expression and cell growth. AS CyD1 significantly inhibited HUVEC growth even in the presence of VEGF. AS CyD1 also significantly suppressed in vitro tube formation in VEGF-treated HUVEC and in vivo macroaneurysm formation in VEGF-treated Matrigel plug. Conclusions: Our results suggest that cyclin D1 may play a role in the maintenance of VEGF expression and that AS CyD1 could be potentially useful for targeting both cancer cells and their microenvironment of tumor vessels.

Expression of PPARδ in multistage carcinogenesis of the colorectum: implications of malignant cancer morphology

Whether peroxisome proliferator-activated receptor (PPAR) δ is a good target for the chemoprevention and/or treatment of colorectal cancer (CRC) remains controversial. Our goal was to examine PPARδ expression in multistage carcinogenesis of the colorectum and to assess the relevance of PPARδ in CRC. Immunohistochemical analysis indicated that PPARδ expression increased from normal mucosa to adenomatous polyps to CRC. In cancer tissues, the PPARδ protein was accumulated only in those cancer cells with highly malignant morphology, as represented by a large-sized nucleus, round-shaped nucleus, and presence of clear nucleoli. Interestingly, the cancer tissue often contained both PPARδ-positive and -negative areas, each retaining their respective specific morphological features. Moreover, this pattern persisted even when PPARδ-positive and -negative cells were aligned next to each other within a single cancer nest or gland and was present in the majority of CRC cases. Immunohistochemistry for Ki-67 proliferation marker showed no significant correlation between Ki-67 and PPARδ in CRC samples. Based on Western blot analysis and quantitative RT–PCR, high PPARδ protein expression correlated with high PPARδ mRNA levels. Peroxisome proliferator-activated receptor δ may have a supporting role in tumorigenesis, and the close association between PPARδ expression and malignant morphology of CRC cells suggests a pivotal role in cancer tissue.

Aberrant Expression of Connexin 26 Is Associated with Lung Metastasis of Colorectal Cancer

Purpose: Connexin 26 (Cx26) is one of the gap junction–forming family members classically considered to be tumor suppressors. However, recent studies show association of elevated expression of Cx26 with poor prognosis in several human malignancies. Furthermore, Cx26 has been observed to be indispensable to spontaneous metastasis of melanoma cells. Here, we assessed Cx26 expression in primary colorectal cancer (CRC) and the metastatic lesions to elucidate its role in metastasis. Experimental Design: Cx26 expression was assessed in 25 adenomas, 167 CRCs, and normal mucosa, together with the metastatic lesions. Results: Normal mucosa and adenomatous tissue expressed Cx26 mainly in the plasma membrane, whereas cancer cells mostly contained Cx26 in the cytoplasm. The incidence of aberrant Cx26 expression varied widely in CRC (mean, 49.5 ± 35.5%), and the expression levels were confirmed by Western blot and quantitative reverse transcription–PCR. Clinicopathologic survey revealed association of high expression with less differentiated histology and venous invasion (P = 0.0053 and P = 0.0084, respectively). Notably, high Cx26 expression was associated with shorter disease-free survival and shorter lung metastasis–free survival in 154 curatively resected CRC sets (P = 0.041 and P = 0.028, respectively). Survey of metastatic lesions revealed that lung metastasis, but not liver and lymph nodes metastases, expressed higher Cx26 than the CRC series or corresponding primary CRCs (P < 0.0001 and P = 0.0001, respectively). Conclusions: These findings suggest that aberrant expression of Cx26 plays an essential role in lung metastasis. Thus, Cx26 is a promising therapeutic target, particularly for CRC patients who develop lung metastasis.

Hepatic expression of ANG2 RNA in metastatic colorectal cancer.

We examined the RNA content of the gene encoding angiopoietin (Ang)‐2, a modifier of angiogenesis, in hepatic metastases of colorectal cancer (CRC) to explore the role of this protein in neovascularization of metastatic foci. Metastatic CRC exhibited notable blood flow and tumor vessel formation at tumor frontiers. Reverse‐transcription polymerase chain reaction assays indicated that the ANG2 RNA content was greater in metastatic CRC than in primary CRC. Investigation of metastatic foci using laser capture microdissection revealed that the RNA content of ANG2, but not ANG1, increased from the bordering liver region to the periphery of the metastatic disease, and also from the periphery to the intermediate portion of the metastatic lesion; immunohistochemical analysis confirmed that there was a corresponding gradual increase in Ang‐2 protein expression. Tie‐2, a receptor for angiopoietins, was preferentially expressed in the bordering liver region rather than in metastatic CRC. Vascular endothelial growth factor (VEGF) also exhibited an expression pattern similar to that of Ang‐2, and there was a significant correlation between the RNA content of ANG2 and that of VEGF in dissected samples (P = .002). Western blot analysis suggested that expression of Ang‐1, Ang‐2, Tie‐2, and VEGF may be regulated at a transcriptional level. The increase in ANG2 RNA content from the peripheral portion of the tumor to the intermediate portion, coinciding with the decrease in recruitment of periendothelial supporting cells around the vascular endothelial cells, suggests that Ang‐2 may play a role in the immaturity of tumor vessels. In conclusion, the current study suggests that Ang‐2 and VEGF may cooperate to enhance the formation of new blood vessels in metastases of CRC to the liver. (HEPATOLOGY 2004;39:528–539.)

Association Between Recurrence of Hepatocellular Carcinoma and α-Fetoprotein Messenger RNA Levels in Peripheral Blood

PurposeIntra- and extrahepatic recurrence is common, even after curative resection for hepatocellular carcinoma (HCC), suggesting preoperative or intraoperative tumor cell dissemination. Reverse transcription — polymerase chain reaction (RT-PCR) for α-fetoprotein (AFP) is used to detect circulating liver cancer cells. We previously developed a quantitative method that allows estimation of the AFP mRNA level by real-time PCR. In the present study, we used this method to measure the AFP mRNA level before and after resection of HCC, then correlated the findings with various clinicopathological characteristics and prognosis.MethodsWe prospectively examined peripheral blood samples from 38 patients with HCC, and bone marrow aspirate from 25 of these patients. As a control, we examined bone marrow from 20 patients with benign diseases. The follow-up period ranged from 32 to 66 months. Real-time RT-PCR was used to detect AFP mRNA levels in the samples.ResultsAFP was expressed in 9 (23.7%) of the 38 peripheral blood samples. The detection of AFP mRNA was significantly correlated with extrahepatic metastasis after primary surgery, and a shorter disease-free survival time (P = 0.0245 each). Bone marrow samples were defined as positive if they expressed AFP mRNA at levels higher than the maximum expressed level in the controls, because only 1 (5%) of the 20 control bone marrow samples had low AFP mRNA expression. Using this cutoff level, 12 (48%) of the 25 patients with HCC had positivity for AFP mRNA. The results of bone marrow RT-PCR did not correlate with the clinocopathological characteristics of prognosis.ConclusionsUsing real-time PCR to measure the AFP mRNA level in blood, but not bone marrow, could be useful for predicting postoperative tumor recurrence.