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Background/aim: Direct-acting antiviral (DAA) therapies for patients with hepatitis C virus (HCV) infection deliver higher cure rates and lower frequencies of adverse events than existing therapies, though DAA treatment costs $45,000-64,000 in Japan. The prognosis of patients who require new long-term care insurance (LTCI) certification is inferior to that of patients who do not. Here, we clarify the factors associated with new LTCI certification in elderly patients with HCV infection who undergo DAA therapy., Patients and Methods: We retrospectively surveyed 53 patients aged ≥70 years who were treated with DAAs, and evaluated the factors associated with new LTCI certification., Results: Of 53 patients, 10 required new LTCI certification. Age ≥85 years and a modified Japanese Cardiovascular Health Study index ≥2 were independently associated with new LTCI certification., Conclusion: In elderly HCV patients, poor frailty status strongly predicted new LTCI certification after DAA therapy., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background/aim: Microsomal prostaglandin (PG) E synthase-1 (mPGES-1) is a terminal enzyme in PGE 2 synthesis and highly expressed in several cancers. In this study, to reveal the involvement of mPGES-1 in skin carcinogenesis, the effect of mPGES-1 deficiency on two-stage skin carcinogenesis in mice was investigated., Materials and Methods: A two-stage skin carcinogenesis model using 7,12-dimethylbenz[a]anthracene (DMBA) as an initiator and 12-O-tetradecanoylphorbol-13-acetate (TPA) as a promoter was applied on mPGES-1 knockout (KO) mice and littermate wild-type mice of a Balb/c genetic background., Results: DMBA/TPA-induced skin carcinogenesis was suppressed in mPGES-1 KO mice. The induction of IL-17 and other inflammatory cytokines by TPA was also suppressed by mPGES-1 deficiency, although DMBA-induced apoptosis was not affected., Conclusion: mPGES-1 promotes chemically induced skin carcinogenesis and might play an important role in the TPA-induced promotion phase of the two-stage skin carcinogenesis model. mPGES-1 inhibition may be a therapeutic target for skin cancer., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: It is difficult to treat the American Joint Committee on Cancer (AJCC)/International Union against Cancer (UICC) T3 hepatocellular carcinoma (HCC), curatively., Patients and Methods: We compared the clinicopathology of T3 group (n=44: T3a 25, T3b 19) with that of the T1 (n=257) or T2 group (n=120) and evaluated favorable conditions of hepatectomy for T3 HCC patients., Results: The T3 group had significantly higher hepatitis B surface antigen (HBsAg)-positive rates and better liver function. Infiltrative large tumors located beyond one sub-segment with intrahepatic metastasis were significantly more common. Significantly, more non-curative large hepatectomies with transarterial embolization were performed. There was no significant difference between T3 and 2 groups in 5-year disease-free survival (DFS) and survival (S). Tumor size more than 55 mm and serum albumin less than 3.5 g/dl were risk factors of hepatectomy for T3 HCC patients by multivariate analysis., Conclusion: Surgeons should resect AJCC/UICC T3 HCC lesions if the patient is able to tolerate surgery., (Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Unlabelled: BACK GROUND/AIM: The purpose of the present study was to clarify the clinicopathological features of non-hepatitis B and -C (NBNC) hepatocellular carcinoma (HCC), the incidence of which has been increasing., Patients and Methods: Two hundred and eighty-four patients with HCC were classified into three groups according to viral hepatitis status, namely NBNC, hepatitis B, and hepatitis C. We compared the three groups and studied related risk factors., Results: Patients without cirrhosis who had increased number of platelets and diabetes mellitus, and a serum alpha-feto-protein (AFP) level <100 ng/dl were more common in the NBNC group. The cumulative survival and disease-free survival were better in the NBNC group than in the other groups. The tumor size and hepatitis B or C viral status were found to be independent risk factors of disease-free survival and the presence of multiple lesions was the only independent risk factor of survival., Conclusion: Close follow-up of NBNC liver cirrhosis and early detection of NBNC-HCC can improve the prognosis after surgery., (Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Background: The prognostic value of lymph node (LN) status in patients who underwent resection for pancreatic cancer (PC) was examined in the present study., Patients and Methods: Fifty-six patients who underwent macroscopic curative resection for PC were analyzed. Twelve factors, including the number of LN metastases, LN ratio, and N category according to the Japanese Pancreatic Society classification, were analyzed using univariate and multivariate analysis., Results: The optimal cut-off value was 0.2 for the LN ratio. Positive surgical margins (p=0.022) and LN ratio ≥0.2 (p=0.017) were identified as independent prognostic factors. Among the 33 patients with regional LN metastasis, patients with LN ratio ≥0.2 had significantly worse prognosis than those with LN ratio <0.2 (median survival time 14 vs. 26 months, p=0.048), however, the differences in survival between those with N1 and those with N2 by Japanese Pancreatic Society classification were not statistically significant (p=0.85)., Conclusion: The LN ratio might be more useful than other parameters as a predictor for survival after resection of PC.

Background: It is of great clinical concern to preoperatively predict the need for blood transfusions during hepatectomy for hepatocellular carcinoma (HCC)., Patients and Methods: A total of 168 consecutive patients undergoing elective hepatectomy for HCC were retrospectively reviewed. We investigated preoperative factors potentially influencing intraoperative blood transfusion and established a predictive scoring system for intraoperative blood transfusion., Results: Thirty-eight patients (22.6%) received red cell blood transfusion during surgery. A preoperative predicting scoring system for blood transfusion was constructed using the following four factors: platelet count <10×10(4)/mm(3) (2 points), α-fetoprotein ≥80 ng/ml (1 point), tumor size ≥4.0 cm (1 point), and major hepatectomy (1 point). The nomogram showed an area under the curve (AUC) of 0.760. This scoring system was highly predictive for blood transfusion (AUC=0.758). When the score was 0 points, the incidence of intraoperative blood transfusion was 3%. The rate increased to 10% and 38% when the score was 1 and 2 points, respectively, and reached 45% when the score was 3 points or more., Conclusion: This predictive scoring system would be useful for preoperatively assessing the need for intraoperative blood transfusions during hepatectomy for HCC.

Background: The aim of this study was to characterize changes in outcome following hepatectomy for hepatocellular carcinoma (HCC) over a 30-year period., Patients and Methods: The clinical records of 372 patients who underwent a macroscopic curative hepatectomy for treatment of HCC between 1980 and 2009 were retrospectively examined. Patients were divided into two groups: an early group (1980-2000) and a late group (2001-2009). The relationship between the chronological treatment period and the surgical outcomes was investigated., Results: The disease-free survival (DFS) was comparable between the early and late groups (at 5 years: 30.3% vs. 31.2%, p=0.526), however, the overall survival (OS) of the late group was significantly better than the one of the early group (at 5 years: 80.1% vs. 50.4%, p<0.001), with this being an independent prognostic factor. Among the 148 patients who underwent transarterial chemoembolization (TACE) for initial hepatic recurrence, the OS after initial recurrence of the late group was significantly better than that of the early group (p=0.002). The OS after initial recurrence was significantly better in patients who underwent repeat hepatectomy than in those who underwent TACE (p=0.044)., Conclusion: The results of hepatectomy for HCC have improved over time by use of various combination therapies after initial HCC recurrence. A repeat hepatectomy may be an acceptable treatment option for HCC recurrence in selected patients.

Background: Endoscopic resection (ER) causes inflammation, edema, fibrosis of the stomach, and severe adhesions around the surrounding tissue. However, little is known about the clinical influence of ER on subsequent laparoscopic gastrectomy (LG) for gastric cancer., Patients and Methods: Between October 2007 and April 2011, 202 consecutive patients underwent curative LG for gastric cancer. Out of these, 30 (15%) LG cases had previously undergone ER and further LG to achieve a cure. The remaining 172 patients (85%) had no history of ER. We reviewed their hospital records retrospectively., Results: Intraperitoneal adhesions were detected in all patients after ER. However, there were no significant differences in clinical outcome, such as surgical duration, blood loss, number of dissected lymph nodes, complications, or hospital stay, between LG cases with and those without previous ER. Of the 30 cases of LG with previous ER, 15 treated within two months after ER had greater blood loss (p<0.005) and a longer surgical duration (p=0.06). LG cases with major or minor perforation during ER also had slightly greater blood loss (p=0.07) than those without. However, the number of dissected lymph nodes associated with surgical curability and postoperative complications were not significantly different in patients with and those without these clinical factors., Conclusion: LG can be performed curatively and safely even after ER. Perforation during ER and shorter time to LG after ER may be potential risks of surgical difficulty in subsequent LG.

Elevated serum interleukin-6 (IL-6) levels have been associated with tumor progression and poor prognosis in patients with esophageal carcinoma. The purpose of the present study was to clarify such a relationship in patients with esophageal squamous cell carcinoma with a focus on the possible influence of chemoradiotherapy (CRT) on tumor IL-6 expression. Data regarding 41 patients with clinical T3-T4 tumors who underwent induction chemoradiotherapy followed by surgery (CRT group) and 60 patients with clinical T1-T4 tumors who underwent surgery alone (Surgery group) between 2001 and 2010, were retrospectively analyzed. Tumor IL-6 expression in resected specimens was evaluated by immunohistochemistry. Tumor IL-6 expression was detected in patients with advanced tumors in the Surgery group (21.1% in p(stage) III-IV vs. 0.2% in p(stage) I-II; 27.8% in pT3-4 vs. 0% in pT1-2), and also correlated with primary tumor progression and surgical curability in the Surgery group. In addition, patients with IL-6-positive tumors had significantly shorter overall survival than those with IL-6-negative tumors in the CRT group, and tumor IL-6 expression had an independent prognostic value in multivariate analysis, whereas no significant difference in overall survival was observed between patients with IL-6-positive and those with IL-6-negative tumors in the Surgery group. These results indicate that pre-treatment tumor IL-6 expression correlates with primary tumor progression, and CRT-induced tumor IL-6 expression predicts poor prognosis.

Interleukin-6 (IL-6) has been associated with disease progression and poor prognosis in esophageal carcinoma. The aim of this study was to investigate the possible influence of IL-6 on the biological activities of esophageal carcinoma cells in terms of invasiveness. The human esophageal carcinoma cell line, KYSE170, was transfected with a plasmid vector expressing IL-6, and a stable transfectant overexpressing IL-6 was established. Invasiveness was evaluated by an invasion assay and compared between IL-6 and control transfectants. The invasiveness of the IL-6 transfectant was significantly higher than that of the control transfectant, and was significantly reduced by IL-6-specific siRNA. In reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, IL-8 expression was significantly higher in the IL-6 transfectant than in the control transfectant, whereas the expression of Hepatocyte growth factor (HGF) and Vascular endothelial growth factor (VEGF) was not different. IL-8 expression in the IL-6 transfectant was significantly inhibited by IL-8-specific siRNA, whereas IL-6 expression was not. In addition, the invasiveness of the IL-6 transfectant was significantly reduced by IL-8-specific siRNA. These results indicate that the overexpression of IL-6 increases the invasiveness of KYSE170 esophageal carcinoma cells and IL-6-induced IL-8 plays a predominant role in increasing invasiveness.

Background/aim: This study aimed to clarify the efficacy of anatomical hepatectomy in patients with preserved liver function., Patients and Methods: We compared the clinicopathology of the anatomical hepatectomy (AH) group (n=264) with that of the non-anatomical hepatectomy (NAH) group (n=85) and evaluated favorable conditions of anatomical hepatectomy for patients with HCC with an indocyanine green 15-minute retention rate of less than 30%., Results: There was no significant difference between the two groups in five-year disease-free survival. However, disease-free survival of the AH group was significantly better than that of the NAH group when patients had T1 tumors, tumors without intrahepatic metastasis, tumors located within one subsegment, or serum alpha feto-protein less than 100 ng/dl (p=0.015, p=0.009, p=0.046 and p=0.036, respectively). Anatomical hepatectomy was an independent favorable prognostic factor by multivariate analysis taking into consideration clinical factors, which could be clarified pre- or intraoperatively (p=0.003)., Conclusion: Anatomical hepatectomy should be performed for HCC patients with HCC with preserved liver function.

Previous clinicopathological studies demonstrated that overexpression of cyclooxygenase-2 (COX-2) is associated with a poor treatment response of esophageal carcinoma. The aim of this study was to elucidate the role of COX-2 overexpression in the chemosensitivity of esophageal carcinoma cells. TE13 human esophageal squamous cell carcinoma cells were transfected with a COX-2 constitutive expression vector, and stable transfectants overexpressing COX-2 were established. COX-2 overexpression in COX-2 transfectants was confirmed with western blotting and prostaglandin-E(2) (PGE(2)) assay. Chemosensitivity testing revealed that sensitivity of COX-2 transfectants to 5-fluorouracil and cisplatin was significantly lower than in control vector-only transfectants, and that sensitivity of COX-2 transfectants was restored by the transfection of COX-2-specific siRNA. In addition, expression of antiapoptotic B-cell lymphoma-extra large (BCL-xL) and myeloid cell leukaemia-1 (MCL-1) was increased in COX-2 transfectants. These results indicate that COX-2 overexpression may reduce the chemosensitivity of esophageal carcinoma cells through up-regulation of the expression of antiapoptotic BCL-2 family proteins.

Background: Precise staging of gastric cancer is essential when selecting for the appropriate treatment approach. 5-Aminolevulinic acid (5-ALA) is metabolized and accumulated as protoporphyrin IX (PpIX), which is a photosensitizer. In this study, we evaluated the diagnostic usefulness of 5-ALA for demonstrating peritoneal dissemination in patients with gastric cancer., Materials and Methods: Fluorescence laparoscopy (FL) was performed in 13 patients with gastric cancer. All patients received preoperative oral administration of 5-ALA. Fluorescence-light laparoscopies were sequentially performed intraoperatively., Results: In four out of the 13 patients, primary tumors were detected by FL and demonstrated serosal invasion on histological examination. Five out of the 13 patients demonstrated peritoneal metastases, and one patient demonstrated superficial liver micrometastases, by FL. All of these lesions were diagnosed as metastatic lesions by hematoxylin and eosin staining., Conclusion: These findings show that FL diagnosis with 5-ALA is accurate and suitable for the detection of peritoneal metastases and of superficial liver micrometastases in patients with gastric cancer.

Background/aim: Hepatectomy without recurrence for more than five years may be considered as being a sufficient resection of primary hepatocellular carcinoma (HCC). The purpose of the present study was to clarify the predictive factors for sufficient resection for HCC patients., Patients and Methods: In our Department, 77 patients with HCC survived without recurrence for more than five years following hepatectomy (curative group) and 187 patients underwent hepatectomy but developed recurrence within five years (recurrence group). We compared the clinicopathology between these two groups and evaluated the favorable factors for HCC without recurrence using logistic regression analysis., Results: The curative group was associated with three independent factors by multivariate logistic regression analyses: tumor size, serum indocyanine green retention rate (ICG15) and anatomic wide hepatectomy (all p<0.05)., Conclusion: Sufficient resection is closely related to patient, tumor, and treatment factors. Anatomic wide hepatectomy for small (<29 mm) HCC in patients with sufficient liver function (ICG15 <16.5%) can achieve sufficient resection.

Background: The hepatocyte growth factor (HGF)-Met pathway plays a role in progression to malignant characteristics in tumors, including that of resistance to anticancer drugs. The purpose of this study was to elucidate the possibility of the combination therapy of NK4, a competitive antagonist for HGF., Materials and Methods: We established a genetically modified murine colon cancer cell, CT26, to produce abundant NK4 (CT26/NK4). Cell proliferation, apoptosis, cell cycle, intracellular signaling, and 5-fluorouracil (5-FU) metabolism of this cell line were examined., Results: There was no difference in thymidylate synthase mRNA level between mock-transfected control CT26 cells and CT26/NK4 cells, suggesting that NK4 expression does not change 5-FU metabolism. NK4 gene expression enhanced 5-FU-induced caspase-3 and -7 activation of CT26. Cell cycle analysis showed that NK4 gene expression and 5-FU treatment caused an increase in the proportion of sub-G(1) cells. On 5-FU treatment, phosphorylation of Akt and Erk1/2 was suppressed in CT26/NK4 less than in mock-transfected cells. 5-FU showed a stronger cytotoxic activity towards CT26/NK4 cells than control CT26 cells., Conclusion: 5-FU exerts an additional effect on apoptosis of NK4-expressing CT26 cells by down-regulating intracellular signaling of the HGF/c-Met pathway.

Background: Recent findings suggest that cells with surface CD markers include cancer stem cells (CSCs) which can produce a cancer cluster, and that the presence of CSCs may be linked with prognosis. CD133 and CD44 are among the most useful markers for identification of colorectal CSCs., Materials and Methods: An immunohistological analysis of CD133 and CD44 was performed using tissue from cases shown to be locoregionally recurrent or non-recurrent clinico-pathologically., Results: The CD133-positive rates were 38.7% and 59.23% in non-recurrent and recurrent cases, respectively, and the CD44-positive rates were 35.5% and 44.4%, respectively. Expression of the CD markers had no correlation with other clinicopathological factors. The prognosis of patients who were positive for both markers was significantly worse than that of other patients., Conclusion: These results suggest that detection of CD133 and CD44 can provide useful information for selection of treatment and performance of intensive follow-up of colorectal cancer.

Background/aim: RUNX3 is a novel gastric cancer tumor suppressor. RUNX3 promoter hypermethylation is associated with many types of cancer, including colorectal cancer. Furthermore, the RUNX3 promotor is one of the CpG island methylator phenotype (CIMP)-specific promotors. CIMP is a distinct phenotype associated with microsatellite instability (MSI) in colorectal cancer. In this study, the suitability of the quantitative analysis of RUNX3 promoter hypermethylation as a novel serum tumor marker was investigated. Moreover, we investigated the relationship between RUNX3 promoter methylation and MSI in colorectal cancer., Patients and Methods: A RUNX3 real-time quantitative methylation-specific PCR (RTQ-MSP) technique we developed was used to analyze the CpG sites in the RUNX3 promoter of 119 colorectal tumors and 344 sera from colorectal cancer patients. MSI analysis of 119 colorectal tumors was performed with five microsatellite markers (BAT25, BAT26, D5S346, D2S123, and D17S250)., Results: Proximal colon tumors exhibited significantly higher RUNX3 methylation than their paired normal tissues (p=0.0438). Analysis of the clinicopathological parameters revealed that a proximal location (p=0.0054), lymphatic invasion (p<0.0001), and an advanced pathological stage (p=0.0018) were associated with significantly higher RUNX3 methylation. Assessment of the relationship between RUNX3 methylation and tumor MSI revealed 11 out of 13 tumors with high-frequency MSI (85%) were positive for RUNX3 hypermethylation, significantly more than the tumors with low-frequency MSI or which were microsatellite stable (34%, p=0.0070). In preoperative sera from 344 colorectal cancer patients, significantly higher RUNX3 methylation was associated with lymphatic invasion (p=0.0487) and an advanced pathological stage (p=0.0466). Post-operative follow-up data revealed that recurrence cases exhibited significantly higher preoperative serum RUNX3 methylation than non-recurrence cases (p=0.0003). Concomitant analysis of carcinoembryonic antigen (CEA) levels in the preoperative sera showed that 17.7% (61/344) were CEA-negative but RUNX3 methylation-positive, which means assessing both serum RUNX3 methylation and CEA should improve diagnosis of colorectal carcinoma., Conclusion: RTQ-MSP-based quantification of serum RUNX3 methylation is useful for the detection and monitoring of colorectal cancer.

Peritoneal dissemination of gastric cancer is a refractory disease. This paper focuses on the efficacy of actin-binding marine macrolide latrunculin A, which quickly inhibits actin polymerization and disrupts the function of the actin cytoskeleton. The effects of latrunculin A on cell viability in vitro were evaluated by treatment of MKN45 or NUGC-4 cell cultures. An in vitro viability assay demonstrated an anticancer effect of latrunculin A in a dose-dependent manner. Latrunculin A induced acute cell injury and programmed cell death through activating the caspase-3/7 pathway. In vivo, MKN45 or NUGC-4 cells were intraperitoneally inoculated into nude mice, as a model of peritoneal dissemination. Intraperitoneal (i.p.) injection of latrunculin A significantly improved survival rate in mice without any major side-effects. Data indicated that latrunculin A has strong anticancer effects, and it may be a new candidate i.p. drug against peritoneal dissemination of gastric cancer in humans.

Background: Recently, the use of histone deacetylase inhibitors (HDACI) with gene therapy has been shown to improve the effect of this therapy. The effectiveness of one of the novel HDACIs, FK228, was examined in adenovirus-mediated p53 gene therapy of esophageal squamous cell carcinoma (ESCC)., Materials and Methods: The expression levels of coxsackie and adenovirus receptor (CAR) in ESCC patients were examined immunohistochemically. CAR induction by FK228 in ESCC cells was analyzed by real-time PCR and Western blotting. The efficiencies of adenoviral transduction treated with FK228 were determined using AV1.0CMV-betagal. The acetylation of p53 protein was detected by Western blotting., Results: CAR expression was reduced in some tumor specimens compared to that in normal specimens. CAR expression was increased by FK228 in both in vitro and in vivo experiments. FK228 improved the efficiency of adenovirus infection. Acetylated p53 protein was increased in a dose- and a time-dependent manner., Conclusion: Our findings suggest that FK228 has a potent ability to augment the effect of adenovirus-mediated p53 gene therapy in ESCC cells.

We previously performed SEREX (serological identification of antigens by recombinant expression cloning) using the sera of patients with esophageal squamous cell carcinoma (SCC), and isolated a variant clone (AK093616) of ubiquitin-conjugating enzyme E21 (UBE2I). This clone was tentatively designated as UBE2I-v5 and analyzed for biological function by transient transfection of the cDNA into activated Ha-ras-transformed NIH3T3 (ras-NIH) mouse fibroblasts. Chemosensitivity to 92 cytotoxic drugs was compared between UBE2I-v5-transfected cells and the parental ras-NIH cells. The UBE2I-v5-transfected cells were more sensitive than the parental cells to anticancer drugs such as vincristine (VCR), mitoxantrone (MIT) and etoposide (VP16). The regression analysis of the total chemosensitivity pattern of UBE2I-vS-transfected cells revealed that the function of UBE2I-v5 was positively related to RPA2 (replication protein A2), Rho-GDI (Rho guanine nucleotide dissociation inhibitor a), FUS (putative tumor suppressor) and TKT (transketolase) but negatively related to Per-1 (period-I), Ran (nuclear Ras-related protein), PTEN (phosphatase and tensin homolog), C/EBPalpha (CCAAT/enhancer binding protein a) and the tumor suppressor p53. Thus, it is possible that UBE21-v5 plays a role in carcinogenesis by suppressing the function of CIEBPa and/or p53 via RPA2-like activity.

Unlabelled: We examined the effect of the combination of herpes simplex virus type1-thymidine kinase (HSV-TK)-suicide gene therapy and interleukin-21 (IL-21) immune gene therapy., Materials and Methods: To improve tumor specificity and the safety of gene therapy, we designed the suicide gene (HSV-TK) to be driven by midkine (MK) minimal promoter (MKp-TK). Plasmid DNA containing HSV-TK-suicide gene or IL-21 gene was injected into TE2 and Colon26 tumors developed in nude mice and electric pulses were then delivered., Results: Tumors transduced with the MKp-TK gene demonstrated increased sensitivity to ganciclovir (GCV) in vitro and in vivo. MK minimal promoter conferred efficient transcriptional activity to the HSV-TK-suicide gene and in vivo electroporation was an effective method for transducing the MKp-TK gene. IL-21-transduced tumors disappeared completely in syngeneic BALB/c mice. However, the tumors were not suppressed completely in T-cell-depleted nude mice, and antitumor effects were absent in NK-cell-depleted mice. These data suggest that IL-21 induces T- and NK-cell-dependent antitumor effects. Furthermore, the growth of IL-21-producing tumors subsequently transduced with MKp-TK by electroporation was significantly retarded compared with control groups., Conclusion: Using the minimal promoter region of MK to drive the HSV-TK gene and in vivo electroporation to transduce IL-21 DNA into the tumors produced an efficient gene therapy with improved safety. To our knowledge, this is the first report of a combination gene therapy using HSV-TK/GCVand IL-21.

Unlabelled: The quantity and quality of circulating DNA fragments was analyzed by quantitative real-time polymerase chain reactions (qPCR) in plasma from patients with esophageal carcinomas, in order to assess their diagnostic value., Patients and Methods: Plasma was collected preoperatively from 24 patients with esophageal cancer and 21 healthy controls. qPCR was performed using two primer sets for the beta-actin gene, amplifying short and long segments., Results: The DNA concentrations in both the short and long assays of esophageal cancer patients were significantly higher than the controls (p<0.001). The area under the receiver-operating characteristic curve was 0.83 (short) and 0.91 (long) for esophageal cancer patients versus the controls. There was also a significant difference in DNA integrity (short/long) between esophageal cancer patients and the control group (p= 0.001)., Conclusion: qPCR assays for plasma DNA concentrations and their integrity can serve as new diagnostic markers for screening and monitoring patients with esophageal cancer.

Background: The diagnostic value of circulating mRNA for the early detection of primary and recurrent gastric cancer was evaluated., Patients and Methods: Circulating hTERT and MUC1 mRNA were amplified in the plasma from 52 gastric cancer patients (40 preoperative and 12 postoperative patients) and 20 healthy controls. The results were compared with those of a circulating cancer cell assay and methylation-specific polymerase chain reaction assay., Results: Cell-free mRNA of the analyzed genes was detected in 6 (15%) preoperative gastric cancer patients (hTERT: 3 and MUC1: 4 patients) and 2 follow-up patients. These mRNAs were not detected in the plasma from healthy volunteers. There was no correlation between the results of the cell-free mRNA and the other assays., Conclusion: Detection of circulating cell-free mRNA might serve as a new complementary tumor marker for gastric cancer.

Background: Gastric carcinogenesis is thought to involve multiple genetic and epigenetic changes. The relationships between the promoter methylation status of relevant genes in the serum and outcomes in patients undergoing curative gastrectomy for cancer were investigated., Materials and Methods: Pre-operative serum samples obtained from 97 gastric cancer patients, who underwent radical gastrectomy, were subjected to methylation-specific polymerase chain reaction (MSP) assays for the p16, E-cadherin and retinoic acid receptor beta (RARbeta) genes., Results: Promoter hypermethylation of p16, E-cadherin and the RARbeta gene was detected in sera from 18 (19%), 24 (25%) and 24 patients (25%), respectively. Altogether, 47 patients (48%) showed hypermethylation of at least one gene analyzed. Survival curves differed significantly between groups defined by the methylation status of E-cadherin (p<0.05), but not those defined by p16 or RARbeta (p =0. 77 and 0. 19, respectively)., Conclusion: Serum MSP assays can provide not only diagnostic, but also prognostic information in gastric cancer.

Background: To evaluate the efficacy and toxicity of concurrent chemoradiation in patients with esophageal cancer aged 75 and older., Patients and Methods: Twenty-two elderly patients were treated with concurrent chemoradiation. Seventeen received a combination of cisplatin/carboplatin and 5-fluorouracil, 5 received daily 5-fluorouracil. Total doses of radiotherapy ranged from 50 to 65 Gy., Results: Nineteen out of 22 patients completed the planned chemoradiation. Five in 8 patients (63%) with T1-2 lesion and 1 in 14 patients (7o%) with T3-4 lesion achieved complete response (p=0.011). Only 2 patients experienced grade 3 toxicities. Grade 2 or worse hematological toxicities were less frequently observed in patients who received local-field radiotherapy (p=0.006). There was no isolated lymph node recurrence. The median survival time was 9 months., Conclusion: This study showed the potential for chemoradiation in elderly patients. A planning target volume of radiotherapy should include only clinically involved lesions in this setting.

Background: Radiofrequency ablation (RFA) is one of the standard percutaneous therapies for hepatocellular carcinoma (HCC), but RFA has not been applied to treat bone metastasis from HCC., Case: A 65-year-old male patient, who underwent hepatectomy for HCC two and a half years previously, complained of pain in his right thigh. Imaging modalities and a needle biopsy revealed metastatic HCC in his right acetabulum. The first RFA therapy was attempted under computed tomography (CT) guidance with 42 Gy radiation therapy. The second RFA therapy was performed for tumor recurrence in his pelvis at 4 years after the first RFA., Conclusion: RFA is a safe, easy, repeatable and effective therapy and should be one of the most important therapeutic modalities for bone metastasis from HCC.

Background: The objectives of this study were to evaluate the efficacy and toxicity of concurrent chemoradiation in patients with esophageal cancer aged 80 and older., Patients and Methods: Seven patients with esophageal cancer, aged 80 or more were treated with chemoradiation. Five received a systemic combination of cisplatin and 5-fluorouracil while 2 received daily 5-fluorouracil, concurrent with radiotherapy. The total doses of radiotherapy ranged from 50 to 65 Gy., Results: Complete response was obtained in 3 patients, and partial response and no change in 2 cases each. Esophageal passage improved in 4 patients. The treatment was well-tolerated. There was no death attributable to any adverse treatment effects. None of the patients experienced grade 3 or worse acute toxicities., Conclusion: This study demonstrated that advanced age per se is not a sufficient reason to exclude elderly patients from aggressive treatment.

Electroporation facilitates transfer of chemicals or plasmid DNA from extracellular milieu into cells by increasing the permeability of the cell membrane. Delivery of electric pulses to established tumors thereby can improve the susceptibility of tumors to an anti-cancer agent administered. We examined whether electroporation-mediated transfer of cytokine genes into solid tumors could produce anti-tumor effects in the tumor-bearing mice. Plasmid DNA containing cytokine genes were injected into human esophageal T.Tn tumors developed in nude mice and electric pulses were then delivered. Administration of murine GM-CSF or human IL-2 gene followed by electroporation significantly suppressed the subsequent growth of T.Tn tumors and prolonged the survival of the inoculated mice. In contrast, electroporation-mediated introduction of a control gene, human GM-CSF gene, whose products do not bind to murine GM-CSF receptors, did not achieve any anti-tumor effects. In vivo transfection of cytokine genes with electroporation could be a possible therapeutic strategy for established solid tumors.

Peritoneal dissemination is one of the major patterns of postoperative recurrence and a major cause of death in pancreatic ductal adenocarcinoma. To predict the development of postoperative peritoneal dissemination (PPD), correlations between the occurrence of PPD and clinicopathological factors were analyzed in 23 patients with pancreatic ductal carcinoma who underwent tumor resection and were carefully monitored after the operation. Fifteen cases (65.2%) developed PPD, but the other 8 cases (34.8%) did not meet the criteria for a diagnosis of PPD. Location of the tumor in the pancreatic tail or body (p = 0.0041, HR = 10.827, 95% CI = 2.126-55.127) and the presence of extra-tumoral perineural invasion (ETNI) (p = 0.0490, HR = 0.114, 95% CI = 0.013-0.990) were found to have predictive value for the development of PPD. These results suggested that intensive therapy selectively focused on the peritoneum in cases with ETNI, might prevent the development of PPD and reduce the side effects of treatment in cases without ETNI.

Induction of apoptosis with chemotherapeutic agents or radiation in tumours is frequently related to the status of those p53 gene of the tumours. To examine whether forced expression of the wild-type p53 gene in tumour cells can modulate their susceptibility to radiation and anti-cancer agents, we retrovirally transduced two types of human breast cancer cell lines, which respectively harboured a mutated p53 gene (OCUB-M) or wild-type p53 gene (YMB-1), with the wild-type p53 gene. Transduced cells which consistently expressed the wild-type p53 gene (OCUB-M/p53, YMB-1/p53) proliferated at the same rate as control cells which were transduced with the beta-galactosidase gene (OCUB-M/lacz, YMB-1/lacz). However, sensitivity to radiation was increased in OCUB-M/p53 cells but not in YMB-1/p53 cells. In vitro chemosensitivity to DNA-damaging anticancer agents such as cyclophosphamide and 5-fluorouracil was not influenced by the transduction of the wild-type p53 gene in either cells. Expression of the wild-type p53 gene in p53-mutated human breast cancer cells can therefore increase their sensitivity to radiation but not their chemosensitivity. Therapeutic effects following by the transduction of the wild-type p53 gene were not observed in breast cancer cells already bearing the wild-type p53 gene.

We examined the effect of herpes simplex virus-thymidine kinase gene (HSV-TK)-mediated suicide gene therapy on human esophageal cancer. Two human lines, T.Tn cells which bears truncated p53 and TE2 cells with wild-type p53, were transduced with the HSV-TK gene and tested for their sensitivities to a prodrug, ganciclovir (GCV). The transduced cells, T.Tn/TK and TE2/TK, increased in vitro sensitivity to GCV compared with that of respective wild-type cells. However, the growth suppression of T.Tn/TK tumors induced by GCV was marginal in nude mice and the tumors regrew thereafter. In contrast, the growth of TE2/TK tumors was significantly inhibited by GCV and all the tumors disappeared. The status of the p53 gene of tumor cells thereby may influence the efficacy of the HSV-TK/GCV system.

The inoculation of cytokine-producing tumor cells into syngeneic animals can produce antitumor effects. We previously reported that injection of a mixed population of murine colon carcinoma (Colon 26) cells transduced with interleukin-4 (IL-4) gene with granulocyte macrophage-colony stimulating factor (GM-CSF) gene developed subcutaneous tumors in syngeneic mice, but these tumors thereafter regressed spontaneously (Oncology 1997;54:69-73). In this study, we found the generation of tumor-specific acquired immunity in the mice which had rejected the tumors of a mixed population of IL-4 and GM-CSF producer cells, and that the immunity was mediated by mature T cells. Moreover, local secretion of both IL-4 and GM-CSF in the vicinity of tumors was found to be crucial for this rejection. These data suggest that enhanced host defense brought about by the combined expression of IL-4 and GM-CSF genes in tumor cells is a potential therapeutic strategy.

We examined the antitumor effect of low and high interleukin-2 (IL-2) producers of murine colon carcinoma cells (Colon 26) which were generated by transduction with IL-2 gene in an experimental lung metastasis model using syngeneic mice. Intravenous injection of the low IL-2 producer cells formed multiple lung metastatic foci and the survival of the mice was not different from that of the mice injected with wild-type cells. However, the mice administrated with the high producer cells survived significantly longer. Subcutaneous inoculation of the low producers, although it caused the development of local tumors at the inoculation sites in some of the mice tested, inhibited lung metastasis of wild-type cells subsequently inoculated and prolonged the survival of the mice rechallenged with Meth A cells, syngeneic fibrosarcoma cells. In contrast, inoculation of the high producers did not cause the development of subcutaneous tumors and inhibited the experimental metastasis of parental but not Meth A cells inoculated thereafter. Thus, the amount of secreted IL-2 from tumor cells differentially influences antitumor effects by inducing tumor specific and nonspecific immunity.

We examined the correlation between the amount of exogenous protein secreted and that of its viral transcript from PA317 packaging cells transduced with a cytokine gene. Amphotropic packaging cells for the retrovirus expressing murine interleukin-15 (IL-15) gene were cloned, and we examined 22 clones for both the amount of IL-15 secreted with enzyme-linked immunosorbent assay and the IL-15 transcript released as infectious virions by slot blot-hybridization analysis. The study revealed that there was no statistical correlation between them, and suggests that the integrated transcript is used discordantly. Thus, the examination of secreted protein from packaging cells cannot be substituted for laborious assays to determine the amount of virion transcripts when isolating high titer packaging cells.

We have examined the effects of irradiation on the cytokine secretion from genetically modified human esophageal and gastric carcinomas. Both cell lines were transduced retrovirally to secrete interleukin-2, interleukin-6 and granulocyte-macrophage colony-stimulating factor, respectively. The metabolism of all the transduced cells was partially inhibited by 6 Gy, and greatly inhibited by 20 Gy irradiation. Cytokine productions, however, was not affected by 6 Gy in many cases, and continued to be detected even after 60 Gy irradiation. The analysis of the dose and time of irradiation in each cytokine producer is useful for the designing of tumor vaccine using cytokine gene transfer.