Tomoyoshi Uetake

DICKKOPF-4 and -2 genes are upregulated in human colorectal cancer.

To comprehensively screen for genetic events underlying colorectal cancer, we performed suppression subtraction hybridization analysis on an advanced colon cancer. Because Dickkopf‐4, a member of the Dickkopf family acting as a Wnt‐signaling modulator, was identified as one of the upregulated genes in this specimen, we investigated expression profiles of all the Dickkopf family members in 55 colorectal tumors (21 cancers and 34 adenomas). We also investigated mechanisms regulating the expression of Dickkopf‐4 in these cancers in vitro and in vivo. Compared with normal adjacent mucosae, Dickkopf‐4 (median 27.4, P < 0.01) and ‐2 (median 51.4, P < 0.01) were strongly expressed in colorectal cancers. The level of Dickkopf‐4 was positively correlated with fibroblast growth factor‐20 (rs = 0.61, P = 0.00017), a representative β‐catenin transcriptional target gene, and with the degree of nuclear accumulation of β‐catenin in colorectal tumors. Dickkopf‐4 was induced by activated β‐catenin in vitro. Reciprocally, recombinant Dickkopf‐4 significantly inhibited T‐cell factor/lymphocyte enhancer factor reporter activity stimulated by recombinant Wnt3a in human embryonic kidney 293 cells. We conclude that Dickkopf‐4 and ‐2 are significantly upregulated in most colorectal tumors, and that Dickkopf‐4 upregulation reflects activation of the Wnt/canonical pathway. (Cancer Sci 2009; 100: 1923–1930)

Hepatocellular carcinoma risk assessment using gadoxetic acid-enhanced hepatocyte phase magnetic resonance imaging.

To investigate whether the patients with hypovascular liver nodules determined on the arterial phase and hypointensity on the hepatocyte phase gadoxetic acid‐enhanced magnetic resonance imaging (hypovascular hypointense nodules) are at increased risk of hepatocarcinogenesis, we assessed subsequent typical hepatocellular carcinoma (HCC) development at any sites of the liver with and without such nodules.

Deep Sequencing and Phylogenetic Analysis of Variants Resistant to Interferon-Based Protease Inhibitor Therapy in Chronic Hepatitis Induced by Genotype 1b Hepatitis C Virus

ABSTRACT Because of recent advances in deep sequencing technology, detailed analysis of hepatitis C virus (HCV) quasispecies and their dynamic changes in response to direct antiviral agents (DAAs) became possible, although the role of quasispecies is not fully understood. In this study, to clarify the evolution of viral quasispecies and the origin of drug-resistant mutations induced by interferon (IFN)-based protease inhibitor therapy, the nonstructural-3 (NS3) region of genotype 1b HCV in 34 chronic hepatitis patients treated with telaprevir (TVR)/pegylated interferon (PEG-IFN)/ribavirin (RBV) was subjected to a deep sequencing study coupled with phylogenetic analysis. Twenty-six patients (76.5%) achieved a sustained viral response (SVR), while 8 patients did not (non-SVR; 23.5%). When the complexity of the quasispecies was expressed as the mutation frequency or Shannon entropy value, a significant decrease in the IFNL3 (rs8099917) TT group and a marginal decrease in the SVR group were found soon (12 h) after the introduction of treatment, whereas there was no decrease in the non-SVR group and no significant decrease in mutation frequency in the IFNL3 TG/GG group. In the analysis of viral quasispecies composition in non-SVR patients, major populations greatly changed, accompanied by the appearance of resistance, and the compositions were unlikely to return to the pretreatment composition even after the end of therapy. Clinically TVR-resistant variants were observed in 5 non-SVR patients (5/8, 62.5%), all of which were suspected to have acquired resistance by mutations through phylogenetic analysis. In conclusion, results of the study have important implications for treatment response and outcome in interferon-based protease inhibitor therapy. IMPORTANCE In the host, hepatitis C virus (HCV) consists of a variety of populations (quasispecies), and it is supposed that dynamic changes in quasispecies are closely related to pathogenesis, although this is poorly understood. In this study, recently developed deep sequencing technology was introduced, and changes in quasispecies associated with telaprevir (TVR)/pegylated interferon (PEG-IFN)/ribavirin (RBV) triple therapy and their clinical significance were investigated extensively by phylogenetic tree analysis. Through this study, the associations among treatment response, changes in viral quasispecies complexity in the early stage of treatment, changes in the quasispecies composition, and origin of TVR-resistant variant HCV were elucidated.

Stratifying the risk of lymph node metastasis in undifferentiated-type early gastric cancer

AIM To study how lymph node metastasis (LNM) risk is stratified in undifferentiated-type early gastric cancer (undiff-EGC) dependent on combinations of risk factors. METHODS Five hundred and sixty-seven cases with undiff-EGC undergoing gastrectomy with lymphadenectomy were examined retrospectively. Using clinicopathological factors of patient age, location, size, an endoscopic macroscopic tumor form, ulceration, depth, histology, lymphatic involvement (LI) and venous involvement (VI), LNM risk was examined and stratified by conventional statistical analysis and data-mining analysis. RESULTS LNM was positive in 44 of 567 cases (7.8%). Univariate analysis revealed > 2 cm, protrusion, submucosal (sm), mixed type, LI and VI as significant prognostic factors and > 2 cm and LI-positive were independent factors by multivariate analysis. In preoperatively evaluable factors excluding LVI, sm and > 2 cm were independent factors. According to the depth and size, cases were categorized into the low-risk group [m and ≤ 2 cm, 0% (LNM incidence)], the moderate-risk group (m and > 2 cm, 5.6%; and sm and ≤ 2 cm, 6.0%), and the high-risk group (sm and > 2 cm, 19.3%). On the other hand, LNM occurred in 1.4% in all LI-negative cases, greatly lower than 28.2% in all LI-positive cases, and LNM incidence was low in LI-negative cases even in the moderate- and high-risk groups. CONCLUSION LNM-related factors in undiff-EGC were depth and size preoperatively while those were LI and size postoperatively. Among these factors, LI was the most significantly correlated factor.

Analysis of the complete open reading frame of genotype 2b hepatitis C virus in association with the response to peginterferon and ribavirin therapy.

Background and Aims Patients infected with genotype 2b hepatitis C virus (HCV) generally can achieve favorable responses to pegylated-interferon plus ribavirin therapy (PEG-IFN/RBV). However, a proportion of patients show poorer responses and the correlation between viral sequence variation and treatment outcome remains unclear. Methods The pretreatment complete open reading frame (ORF) sequences of genotype 2b HCV determined by direct sequencing were investigated for correlation with the final outcome in a total of 60 patients. Results In this study group, 87.5% (14/16) of non-sustained virological response (non-SVR) patients (n = 16) were relapsers. Compared to sustained virological response (SVR) patients (n = 44), non-SVR patients were older and could not achieve prompt viral clearance after the therapy induction. Comparing each viral protein between the two groups, viral sequences were more diverse in SVR patients and that diversity was found primarily in the E1, p7, and NS5A proteins. In searching for specific viral regions associated with the final outcome, several regions in E2, p7, NS2, NS5A, and NS5B were extracted. Among these regions, part of the interferon sensitivity determining region (ISDR) was included. In these regions, amino acid substitutions were associated with the final outcome in an incremental manner, depending upon the number of substitutions. Conclusions Viral sequences are more diverse in SVR patients than non-SVR patients receiving PEG-IFN/RBV therapy for genotype-2b HCV infection. Through systematic comparison of viral sequences, several specific regions, including part of the ISDR, were extracted as having significant correlation with the final outcome.

Correlation between pretreatment viral sequences and the emergence of lamivudine resistance in hepatitis B virus infection

The emergence of amino acid or nucleotide substitutions leads to lamivudine resistance in hepatitis B virus (HBV) infected patients. The aim of this study was to investigate whether viral sequences help predict the emergence of lamivudine resistance. The study subjects comprised 59 consecutive patients infected with HBV treated with daily therapy of 100 mg lamivudine. Among those, 32 patients with adequate pretreatment serum preservation were investigated for the correlation between viral amino acid substitutions and the appearance of lamivudine resistance with consideration of clinical background by determining dominant HBV full open reading frames. Viral resistance to lamivudine emerged in 28 of 59 patients (47%) in a median period of 2.45 years. Sequence comparisons of HBV genomes between patients who later developed lamivudine resistance and patients who did not revealed the existence of significant differences between the two groups in the pre‐S1 84 (P = 0.042), pre‐S2 1 (P = 0.017) and 22 (P = 0.015), and polymerase tp 95 (P = 0.046), judged by a log‐rank test. Viral sequence analyses revealed the presence of amino acid substitutions in HBV pre‐S1 and pre‐S2 that may be associated with the emergence of lamivudine resistance during chronic HBV infection. J. Med. Virol. 84:1360–1368, 2012.

Interleukin 12(IL-12) production and its relations to other cytokines in patients with chronic hepatitis C

Abstract Recent reports emphasize the importance of the T helper (Th)1/Th2 immunological balance in viral infections. We investigated this immune response in patients infected with hepatitis C virus (HCV). Using enzyme-linked immunosorbent assays (ELISA), we measured the Staphylococcus aureus-stimulated interleukin (IL)-12, IL-10, and IL-6 production by peripheral blood mononuclear cells (PBMC), and plasma levels of transforming growth factor (TGF)-β1. Ninety HCV-seropositive patients [43 with chronic hepatitis (CH), 27 with liver cirrhosis (LC), and 20 with hepatocellular carcinoma (HCC)], were enrolled in this study. IL-12 levels produced by PBMC were significantly lower in patients infected with HCV than in normal controls. Moreover, IL-12 production decreased with the progression of liver disease in patients with chronic hepatitis C. There was no relationship between IL-12 and IL-6 production. Decrease in IL-12 production was accompanied by decrease in IL-10 production and increase in plasma TGF-β1 levels. When IL-12 production was compared with blood chemistry and laboratory parameters, there were no correlations with blood HCV RNA levels and serum immunoglobulin concentrations, but a correlation existed between serum transaminase (ALT) level and IL-12 production. These results suggest that HCV infection may suppress IL-12 production and stimulate TGF-β1 production, and these changes may be related to the progression of liver disease in patients with chronic hepatitis C.

Semiannual Imaging Surveillance Is Associated with Better Survivalin Patients with Non-B, Non-C Hepatocellular Carcinoma

Since it remains elusive whether and how the imaging surveillance affects the survival in patients with non-B, non-C hepatocellular carcinoma (NBNC-HCC), we conducted this retrospective study which investigated the association between the semiannual surveillance prior to HCC diagnosis and the survival in patients with the initial diagnosis of HCC induced by hepatitis B virus (HBV) and/or hepatitis C virus (HCV) infections (N = 141) and non-B, non-C etiology (N = 30). It was demonstrated that surveillance was less frequently performed in the NBNC-HCC patients compared to that in HCC patients with HBV and/or HCV infections (B/C-HCC patients), and the survival was unfavorable in NBNC-HCC patients. On the other hand, the survival of NBNC-HCC patients with semiannual surveillance was significantly favorable than those patients without semiannual surveillance, and the survival was similar between B/C-HCCs and NBNC-HCCs with semiannual surveillance. In conclusion, though NBNC-HCC patients compared to B/C-HCC patients had poorer prognosis overall, these NBNC-HCC patients with semiannual surveillance had a better survival almost equivalent to the survival of B/C-HCC patients with semiannual surveillance, demonstrating the clinical utility of the semiannual imaging surveillance program for NBNC-HCCs.