松下 一之

Background: The liver secretes very-low-density lipoproteins (VLDLs) and plays a key role in lipid metabolism. Plasma total triglyceride (TG) level variations have been studied in patients with hepatitis C virus (HCV)-related chronic hepatitis (CH-C). However, the results of these studies are variable. A homogenous assay protocol was recently proposed to directly measure the TG content in VLDL (VLDL-TG) and VLDL remnants. Methodology/Principal Findings: Using the assay protocol, we determined serum VLDL-TG levels in 69 fasting patients with biopsy-proven HCV-related chronic liver disease and 50 healthy subjects. Patients were classified into stages F0-F4 using the 5-point Desmet scale. Serum total TG levels in patients with non-cirrhotic (F1-F3) CH-C did not demonstrate significant differences compared with healthy subjects, but serum VLDL-TG levels did demonstrate significant differences. Mean serum VLDL-TG levels tended to decrease with disease progression from F1 to F4 (cirrhosis). Compared with healthy subjects, serum non-VLDL-TG levels significantly increased in patients with stages F2 and F3 CH-C; however, we observed no significant difference in patients with liver cirrhosis. Furthermore, the serum VLDL-TG/non-VLDL-TG ratio, when taken, demonstrated a significant decrease in patients with CH-C from the mildest stage F1 onward. Conclusions/Significance: The decrease in serum VLDL-TG levels was attenuated by increase in non-VLDL-TG levels in patients with non-cirrhotic CH-C, resulting in comparable total TG levels. Results of previous studies though variable, were confirmed to have a logical basis. The decrease in the serum VLDL-TG/non-VLDL-TG ratio as early as stage F1 demonstrated TG metabolic alterations in early stages of CH-C for the first time. The involvement of TG metabolism in CH-C pathogenesis has been established in experimental animals, while conventional TG measurements are generally considered as poor indicators of CH-C progression in clinical practice. The serum VLDL-TG/non-VLDL-TG ratio, which focuses on TG metabolic alterations, may be an early indicator of CH-C.

We searched for novel tumor markers of pancreatic cancer by three-step serum proteome analysis. Twelve serum abundant proteins were depleted using immunoaffinity columns followed by fractionation by reverse-phase high-performance liquid chromatography. Proteins in each fraction were separated by two-dimensional gel electrophoresis. Then the gel was stained by Coomassie Brilliant Blue. Protein spots in which the expression levels were significantly different between cancer and normal control were identified by LC-MS/MS. One hundred and two spots were upregulated, and 84 spots were downregulated in serum samples obtained from patients with pancreatic cancers, and 58 proteins were identified by mass spectrometry. These candidate proteins were validated using western blot analysis and enzyme-linked immunosorbent assay (ELISA). As a result of these validation process, we could confirm that the serum levels of apolipoprotein A-IV, vitamin D-binding protein, plasma retinol-binding protein 4, and tetranectin were significantly decreased in patients with pancreatic cancer.

A 45-year-old man suffering intermittent melena for several years was referred to our hospital for detailed examinations. A capsule endoscopy and a double-balloon enteroscopy revealed a tumor with a depression on the smooth surface, at approximately 150 cm proximal to the ileocecal valve. We made a diagnosis of ileal carcinoma by biopsy, and a partial ileal resection with lymph node dissection was performed. The resected specimen showed a yellowish tumor measuring 1.2 × 0.8 cm in size with a depression on the smooth surface. Based on the histological examination and immunohistochemical staining we diagnosed ileal carcinoid tumor. The patient is doing well 2 years after the surgery without any signs of recurrence. © 2011 The Japanese Society of Gastroenterological Surgery.

Angiogenesis plays an essential role in the growth and metastasis of esophageal carcinoma. Vascular endothelial growth factor, thymidine phosphorylase, fibroblast growth factor, midkine, and hepatocyte growth factor have been reported to be vital molecules for tumor angiogenesis. Polymorphisms in gene encoding angiogenic factors or their receptors may alter protein expression and/or activity. Increased angiogenic-factor expression and increased serum levels of these molecules were found to be associated with poor treatment response and poor prognosis. We reviewed the clinicopathological significance of angiogenesis-related molecules in patients with esophageal carcinoma. Antiangiogenic molecular-treatment strategies are also discussed. (Ann Thorac Cardiovasc Surg 2010; 16: 389-393)

Background: Serum levels of novel hydroxy polyunsaturated ultra long-chain fatty acids (hPULCFAs) have been previously shown to be reduced in pre-treatment CRC patients compared to disease-free subjects, independent of disease stage. However, whether reduced levels of hPULCFAs result from the presence of cancer is currently unknown, as is the distribution of hPULCFAs in the general population. The following studies were carried out to assess whether conventional therapy would result in restoration of systemic hPULCFAs in CRC patients, and to investigate the relationship between hPULCFA levels and age. Methods: Tandem mass spectrometry was used to determine serum levels of the 28 carbon-containing hPULCFA C28H46O4 (CRC-446) in the following cohorts: two independent Japanese CRC populations following surgical tumor removal (n = 86), a North American Caucasian CRC cohort (n = 150) following post-surgery combination chemo/radiation therapy, 990 randomly selected anonymized serum samples from subjects ranging between 11 and 99 years of age, as well as longitudinally collected serum samples from healthy normals (n = 8, up to 90 weeks) and stage IV CRC subjects on combination therapy (n = 12, up to 63 weeks). Results: Serum CRC-446 levels in CRC subjects were significantly lower than controls (mean of 0.297 +/- 0.07 ug/ml in controls versus 0.092 +/- 0.03 in CRCs, p < 0.001), and were unaffected by surgical tumor removal or by chemo/radiation treatment (p > 0.05 between pre vs post surgery). CRC-446 levels showed a strong inverse association with age (p < E-11) across the randomly-selected cohort of 990 subjects, with no correlation observed in the CRC-positive subjects. Longitudinal intra-subject results, however, showed relatively stable CRC-446 levels over the short term of up to 90 weeks in both disease-free subjects and late-stage CRC patients. Conclusions: Our findings show that CRC-446 levels are not affected by conventional CRC treatment and inversely correlate with age, which suggest that reduced serum CRC-446 levels likely exist prior to the development of CRC. Extrapolation of the results to a simple screening scenario showed that, compared to fecal blood testing, pre-colonoscopy screening using serum CRC-446 levels would require 80% fewer colonoscopies, would identify risk in subjects under the age of 50, and would result in increased numbers of early cases detected. The precise role these serum metabolites play in the aetiology of cancer development remains to be determined.

Background: Serum levels of novel hydroxy polyunsaturated ultra long-chain fatty acids (hPULCFAs) have been previously shown to be reduced in pre-treatment CRC patients compared to disease-free subjects, independent of disease stage. However, whether reduced levels of hPULCFAs result from the presence of cancer is currently unknown, as is the distribution of hPULCFAs in the general population. The following studies were carried out to assess whether conventional therapy would result in restoration of systemic hPULCFAs in CRC patients, and to investigate the relationship between hPULCFA levels and age. Methods: Tandem mass spectrometry was used to determine serum levels of the 28 carbon-containing hPULCFA C28H46O4 (CRC-446) in the following cohorts: two independent Japanese CRC populations following surgical tumor removal (n = 86), a North American Caucasian CRC cohort (n = 150) following post-surgery combination chemo/radiation therapy, 990 randomly selected anonymized serum samples from subjects ranging between 11 and 99 years of age, as well as longitudinally collected serum samples from healthy normals (n = 8, up to 90 weeks) and stage IV CRC subjects on combination therapy (n = 12, up to 63 weeks). Results: Serum CRC-446 levels in CRC subjects were significantly lower than controls (mean of 0.297 +/- 0.07 ug/ml in controls versus 0.092 +/- 0.03 in CRCs, p < 0.001), and were unaffected by surgical tumor removal or by chemo/radiation treatment (p > 0.05 between pre vs post surgery). CRC-446 levels showed a strong inverse association with age (p < E-11) across the randomly-selected cohort of 990 subjects, with no correlation observed in the CRC-positive subjects. Longitudinal intra-subject results, however, showed relatively stable CRC-446 levels over the short term of up to 90 weeks in both disease-free subjects and late-stage CRC patients. Conclusions: Our findings show that CRC-446 levels are not affected by conventional CRC treatment and inversely correlate with age, which suggest that reduced serum CRC-446 levels likely exist prior to the development of CRC. Extrapolation of the results to a simple screening scenario showed that, compared to fecal blood testing, pre-colonoscopy screening using serum CRC-446 levels would require 80% fewer colonoscopies, would identify risk in subjects under the age of 50, and would result in increased numbers of early cases detected. The precise role these serum metabolites play in the aetiology of cancer development remains to be determined.

Background: Pseudohyperkalemia is uncommon, but important. Local release of potassium caused by contraction of the forearm muscles from a tightly clenched fist or repeated fist clenching during phlebotomy is a recognized cause of pseudohyperkalemia. We investigated the use of a standard protocol to avoid fist clenching during phlebotomy. Study Design: Quality improvement report. Setting & Participants: In 7 healthy volunteers, 10 blood samples were collected over 10-second intervals after 20 repeated fist clenching and unclenching movements. In 86 healthy volunteers, 3 blood samples were collected with and without prior fist clenching. Between September 1, 2006, and June 30, 2007, peripheral venous blood samples were collected from 73,846 outpatients at Chiba University Hospital without a protocol to avoid fist clenching. Between July 1, 2007, and March 31, 2009, blood samples were collected from 171,053 outpatients using the protocol. Quality Improvement Plan: After July 1, 2007, blood samples were collected from the basilic or cephalic vein without making a fist or by making a fist using minimal gripping strength. Also, when multiple specimens were obtained from 1 patient, the specimen for measuring serum electrolytes was obtained after the other specimens. Outcomes & Measurements: Pseudohyperkalemia, defined as unexplained serum potassium level >= 6.5 mmol/L. Results: In the 7 volunteers, the decrease in serum potassium levels after cessation of fist clenching ranged from 8.4%-25.9%. In the 86 volunteers, the percentage with a decrease in serum potassium level >= 0.2 mmol/L between the first and third samples was 25.6% versus 6.7% with or without prior fist clenching, respectively. In clinical practice, we observed 8 cases of pseudohyperkalemia before implementing the protocol (0.0081%) and 1 case (0.00058%) after implementing the protocol (P = 0.001). Limitations: Causes of hyperkalemia before using precautions were assessed using retrospective analyses. Conclusions: Avoiding fist clenching during phlebotomy and not using the first specimen for electrolyte measurements when obtaining multiple specimens from a single patient can reduce the occurrence of pseudohyperkalemia. Am J Kidney Dis 56: 686-692. Crown Copyright (C) 2010 Published by Elsevier Inc. on behalf of the National Kidney Foundation, Inc. All rights reserved.

Background: Pseudohyperkalemia is uncommon, but important. Local release of potassium caused by contraction of the forearm muscles from a tightly clenched fist or repeated fist clenching during phlebotomy is a recognized cause of pseudohyperkalemia. We investigated the use of a standard protocol to avoid fist clenching during phlebotomy. Study Design: Quality improvement report. Setting & Participants: In 7 healthy volunteers, 10 blood samples were collected over 10-second intervals after 20 repeated fist clenching and unclenching movements. In 86 healthy volunteers, 3 blood samples were collected with and without prior fist clenching. Between September 1, 2006, and June 30, 2007, peripheral venous blood samples were collected from 73,846 outpatients at Chiba University Hospital without a protocol to avoid fist clenching. Between July 1, 2007, and March 31, 2009, blood samples were collected from 171,053 outpatients using the protocol. Quality Improvement Plan: After July 1, 2007, blood samples were collected from the basilic or cephalic vein without making a fist or by making a fist using minimal gripping strength. Also, when multiple specimens were obtained from 1 patient, the specimen for measuring serum electrolytes was obtained after the other specimens. Outcomes & Measurements: Pseudohyperkalemia, defined as unexplained serum potassium level >= 6.5 mmol/L. Results: In the 7 volunteers, the decrease in serum potassium levels after cessation of fist clenching ranged from 8.4%-25.9%. In the 86 volunteers, the percentage with a decrease in serum potassium level >= 0.2 mmol/L between the first and third samples was 25.6% versus 6.7% with or without prior fist clenching, respectively. In clinical practice, we observed 8 cases of pseudohyperkalemia before implementing the protocol (0.0081%) and 1 case (0.00058%) after implementing the protocol (P = 0.001). Limitations: Causes of hyperkalemia before using precautions were assessed using retrospective analyses. Conclusions: Avoiding fist clenching during phlebotomy and not using the first specimen for electrolyte measurements when obtaining multiple specimens from a single patient can reduce the occurrence of pseudohyperkalemia. Am J Kidney Dis 56: 686-692. Crown Copyright (C) 2010 Published by Elsevier Inc. on behalf of the National Kidney Foundation, Inc. All rights reserved.

Lamins, major components of the nuclear lamina, undergo phosphorylation at multiple residues during cell cycle progression, but their detailed phosphorylation kinetics remain largely undetermined. Here, we examined changes in the phosphorylation of major phosphorylation residues (Thr14, Ser17, 5er385, Ser387, and Ser401) of lamin B2 and the homologous residues of lamin B1, A/C during the cell cycle using novel antibodies to the site-specific phosphorylation. The phosphorylation levels of these residues independently changed during the cell cycle. Thr14 and Ser17 were phosphorylated during G(2)/M phase to anaphase/telophase. Ser385 was persistently phosphorylated during mitosis to G(1) phase, whereas Ser387 was phosphorylated discontinuously in prophase and G(1) phase. Ser401 phosphorylation was enhanced in the G(1)/S boundary. Immunoprecipitation using the phospho-antibodies suggested that metaphase-phosphorylation at Thr14, Ser17, and Ser385 of lamins occurred simultaneously, whereas G(1)-phase phosphorylation at Ser385 and Ser387 occurred in distinct pools or with different timings. Additionally, we showed that lamin B2 phosphorylated at Ser17, but not Ser385, Ser387 and Ser401, was exclusively nonionic detergent soluble, depolymerized forms in growing cells, implicating specific involvement of Ser17 phosphorylation in lamin depolymerization and nuclear envelope breakdown. These results suggest that the phosphorylations at different residues of lamins might play specific roles throughout the cell cycle. (C) 2010 Elsevier Inc. All rights reserved.

Lamins, major components of the nuclear lamina, undergo phosphorylation at multiple residues during cell cycle progression, but their detailed phosphorylation kinetics remain largely undetermined. Here, we examined changes in the phosphorylation of major phosphorylation residues (Thr14, Ser17, 5er385, Ser387, and Ser401) of lamin B2 and the homologous residues of lamin B1, A/C during the cell cycle using novel antibodies to the site-specific phosphorylation. The phosphorylation levels of these residues independently changed during the cell cycle. Thr14 and Ser17 were phosphorylated during G(2)/M phase to anaphase/telophase. Ser385 was persistently phosphorylated during mitosis to G(1) phase, whereas Ser387 was phosphorylated discontinuously in prophase and G(1) phase. Ser401 phosphorylation was enhanced in the G(1)/S boundary. Immunoprecipitation using the phospho-antibodies suggested that metaphase-phosphorylation at Thr14, Ser17, and Ser385 of lamins occurred simultaneously, whereas G(1)-phase phosphorylation at Ser385 and Ser387 occurred in distinct pools or with different timings. Additionally, we showed that lamin B2 phosphorylated at Ser17, but not Ser385, Ser387 and Ser401, was exclusively nonionic detergent soluble, depolymerized forms in growing cells, implicating specific involvement of Ser17 phosphorylation in lamin depolymerization and nuclear envelope breakdown. These results suggest that the phosphorylations at different residues of lamins might play specific roles throughout the cell cycle. (C) 2010 Elsevier Inc. All rights reserved.

We detected adenoviral DNA fragments in excretions of 10 esophageal cancer patients by DNA-PCR after tumor injection of Ad-CMV-vector. A total of 220 samples consisting of feces, gargling saliva, urine, and blood plasma were assessed. A total of 29.7% of feces samples and 13.2% of gargling saliva samples were positive for adenoviral DNA fragments, but 89.7% of the positive feces samples and all of the positive gargling saliva samples turned negative on day 12 after tumor injection. Although adenoviral DNA fragments may be pathogen-free, patients' feces and gargling saliva contain adenoviral DNA fragments for 12 days after injection. (Cancer Sci 2010; 101: 289-291).

Objective: The antitumor mechanism of histone deacetylase (HDAC) inhibitors differs from conventional antitumor agents. HDAC inhibitors may be effective as novel therapeutic agents for esophageal squamous cell carcinoma (ESCC). This study describes the antiproliferative activity of CHAP31, a novel HDAC inhibitor. Furthermore, the molecular mechanism of CHAP31-induced apoptosis was investigated in ESCC. Methods/Results: The antitumor activity of CHAP31 was tested in esophageal cancer cell lines (T.Tn and TE2), and potent antitumor activity was observed in vitro and in vivo. In addition, CHAP31 induced apoptosis in esophageal cancer cells. Next, the mechanisms of CHAP31-induced apoptosis were examined using quantitative real-time RT-PCR and Western blotting. No processing of caspase 8 was observed, but CHAP31 induced the cleavage of caspase 9 and up-regulation of the Bax/Bcl-2 protein ratio. Conclusion: This study provides new and important information on the potent antitumor activity of CHAP31 and the apoptotic pathway induced by CHAP31 in human esophageal cancer cell lines T.Tn and TE2. In contrast to previous reports showing that apoptosis induced by HDAC inhibitors includes the extrinsic pathway, in our study, apoptosis induced by CHAP31 in the human esophageal cell lines T.Tn and TE2 involved only the intrinsic pathway. Copyright (C) 2010 S. Karger AG, Basel

We detected adenoviral DNA fragments in excretions of 10 esophageal cancer patients by DNA-PCR after tumor injection of Ad-CMV-vector. A total of 220 samples consisting of feces, gargling saliva, urine, and blood plasma were assessed. A total of 29.7% of feces samples and 13.2% of gargling saliva samples were positive for adenoviral DNA fragments, but 89.7% of the positive feces samples and all of the positive gargling saliva samples turned negative on day 12 after tumor injection. Although adenoviral DNA fragments may be pathogen-free, patients' feces and gargling saliva contain adenoviral DNA fragments for 12 days after injection. (Cancer Sci 2010; 101: 289-291).

To investigate changes in protein expression by proteomic analysis in the sera of patients with sepsis and to identify new biomarkers of sepsis. A total of 45 consecutive patients with severe sepsis or septic shock (sepsis group), 22 healthy volunteers, and 23 patients undergoing off-pump coronary artery bypass grafting (control group). Serum samples from eight patients of each group underwent proteomic analysis involving removal of 12 major proteins and subsequent reversed-phase high-performance liquid chromatography fractionation and one-dimensional electrophoresis. The intensity of 41 bands (with 12 proteins identified) increased and that of 42 bands (with 22 proteins identified) decreased in the sepsis group. Results of proteomic analysis successfully validated by Western blotting and/or enzyme-linked immunosorbent assay for three proteins (YKL-40, lipocalin 2, and S100A9) increased in the sepsis group as well as two proteins (retinol-binding protein, vitamin D-binding protein) decreased. Serum YKL-40 levels (sYKL-40) on intensive care unit (ICU) admission were assessed by enzyme-linked immunosorbent assay between the two groups; resulting YKL-40 was significantly higher in the sepsis group (P < 0.001). Furthermore, sYKL-40 on ICU admission was significantly higher in patients with positive blood culture (P < 0.005), patients with septic shock (P < 0.05), and patients requiring continuous hemodiafiltration (P < 0.05) or hydrocortisone replacement therapy (P < 0.005) during subsequent treatment. A positive correlation between sYKL-40 and blood IL-6 level on ICU admission was noted in the sepsis group (r = 0.465, P < 0.01). YKL-40 identified by proteomic analysis is considered as a biomarker of sepsis. However, further investigation is needed to clarify its roles and clinical usefulness as a biomarker.

To investigate changes in protein expression by proteomic analysis in the sera of patients with sepsis and to identify new biomarkers of sepsis. A total of 45 consecutive patients with severe sepsis or septic shock (sepsis group), 22 healthy volunteers, and 23 patients undergoing off-pump coronary artery bypass grafting (control group). Serum samples from eight patients of each group underwent proteomic analysis involving removal of 12 major proteins and subsequent reversed-phase high-performance liquid chromatography fractionation and one-dimensional electrophoresis. The intensity of 41 bands (with 12 proteins identified) increased and that of 42 bands (with 22 proteins identified) decreased in the sepsis group. Results of proteomic analysis successfully validated by Western blotting and/or enzyme-linked immunosorbent assay for three proteins (YKL-40, lipocalin 2, and S100A9) increased in the sepsis group as well as two proteins (retinol-binding protein, vitamin D-binding protein) decreased. Serum YKL-40 levels (sYKL-40) on intensive care unit (ICU) admission were assessed by enzyme-linked immunosorbent assay between the two groups; resulting YKL-40 was significantly higher in the sepsis group (P < 0.001). Furthermore, sYKL-40 on ICU admission was significantly higher in patients with positive blood culture (P < 0.005), patients with septic shock (P < 0.05), and patients requiring continuous hemodiafiltration (P < 0.05) or hydrocortisone replacement therapy (P < 0.005) during subsequent treatment. A positive correlation between sYKL-40 and blood IL-6 level on ICU admission was noted in the sepsis group (r = 0.465, P < 0.01). YKL-40 identified by proteomic analysis is considered as a biomarker of sepsis. However, further investigation is needed to clarify its roles and clinical usefulness as a biomarker.

Background Serological identification of antigens by recombinant cDNA expression cloning (SEREX) is an established method for detecting new tumor-specific antigens. Antibodies to SEREX antigens may be useful for the detection of esophageal squamous cell carcinoma (SCC). Methods A phage cDNA library of a human esophageal SCC cell line was screened using sera of patients with esophageal SCC. The presence and levels of serum antibodies to SEREX antigens were established by Western blotting and enzyme-linked immunosorbent assay (ELISA) using purified recombinant antigen proteins, respectively. Results The newly identified esophageal SCC antigen is encoded by a novel gene located on chromosome 1, here designated CUEC-23. SerumCUEC-23-antibodies (s-CUEC-23-Abs) were detected in 14 of 54 patients with esophageal SCC (26%) by Western blot analysis. Esophageal SCCs were positive for s-CUEC-23-Abs together with CEA, SCC-Ag or CYFRA21-1 in 44, 41 and 52% of cases, respectively. There was no detectable association between the presence of s-CUEC-23-Abs and clinicopathological variables. ELISA showed that the levels of s-CUEC-23-Abs were significantly higher in patients with esophageal SCC than in healthy volunteers (17% in the former using the mean + 3 SD of s-CUEC-23- Abs in healthy controls as the cutoff). Conclusion A new tumor antigen, CUEC-23, was identified by SEREX screening. s-CUEC-23-Abs might be a useful serum marker to detect esophageal SCC.

Background/Aims: Treatment strategy for T2/T3 esophageal carcinoma has become controversial because of recent improvements in chemoradiation therapy. Only a few study analyzed the prognostic impact of clinicopathological factors for surgical outcome of patients with esophageal carcinoma focused on T2/T3 tumors. Methodology: Subjects of this study were 187 patients with pathological T2 (n=46) or pathological T3 (n=141) thoracic esophageal squamous cell carcinoma who received surgical treatment without neoadjuvant therapy. The impact of clinicopathological factors on survival was evaluated by univariate and multivariate analysis. Results: Overall 5-year survival rate of all patients was 38%. Dismal 5-year overall survival rates were observed in patients with 5 or more positive nodes (11%). Multivariate analysis indicated that lymph node metastases (hazard ratio; 2.07, 95%IC, 1.20-3.56, p<0.01) and the number of metastatic nodes (hazard ratio; 1.77, 95%IC, 1.112.82, p=0.02) were independent risk factors for poor survival. However, tumor depth itself was not an independent risk factor for survival. Conclusions: Although survival of patients with pathological T2 or T3 was partly dependent on tumor depth, it was mainly dependent on lymph node status. Multiple positive nodes were independent risk factors for poor survival.

We performed SEREX (serological identification of antigens by recombinant cDNA expression cloning) using the sera of patients with esophageal squamous cell carcinoma (SCC), and examined whether some of the SEREX antigens can affect chemosensitivity against anticancer drugs. We isolated a novel gene which was designated as AISEC (antigen identified by SEREX for esophageal carcinoma). RT-PCR analysis showed that the mRNA expression levels of AISEC were higher in esophageal SCC tissues than in their normal counterparts. By transfection into activated Ha-ras-transformed NIH3T3 (ras-NIH) mouse fbroblasts, we isolated a clone, FAISEC-3, which stably expressed AISEC. FAISEC-3 cells were more resistant to anticancer drugs, such as mitomycin C, ifosfamide, vincristine, camptothecin and etoposide, than parental ras-NIH cells. Luciferase reporter assay after a transient transfection with AISEC cDNA or the control vector revealed that the transactivity of p53 was suppressed by AISEC in a dose-dependent manner. These results suggested that esophageal SCC tissues produce AISEC in increased amounts, which can reduce the chemosensitivity against anticancer drugs possibly by suppressing the p53 transactivation ability.

Background Although the presence of serum p53 antibody (s-p53-Abs) before treatment has been shown to correlate with poor prognosis and lymph node metastasis in esophageal cancer, there has been little information about postoperative s-p53-Abs titer and perioperative changes of s-p53-Abs titers in patients with esophageal carcinoma. Methods A highly specific enzyme-linked immunosorbent assay was used to analyze s-p53-Abs in 110 patients with esophageal squamous cell carcinoma before and 1 month after surgery. The cutoff level of 1.3 U/ml was used to indicate seropositive patients. Impact of postoperative s-p53-Abs titer and perioperative changes of s-p53-Abs on survival was evaluated. Results Forty (36%) of 110 patients were positive for s-p53-Abs before surgery and 35 patients (32%) were positive after surgery. s-p53-Abs titer generally decreased after surgery. Among sero-positive patients, the patients who remained sero-positive after surgery (n = 28) had a worse prognosis than patients who showed sero-conversion (P = 0.02). Among sero-positive patients, the nondecreased titer group showed significantly unfavorable survival (P < 0.01). Multivariate analysis revealed that postoperative s-p53-Abs was an independent risk factor for worse overall survival (adjusted hazard ratio = 3.05; 95% confidence interval = 1.11-8.33; P = 0.03). Conclusions Perioperative monitoring of s-p53-Abs titers was useful to identify patients with esophageal cancer with a high risk for tumor recurrence and a poor prognosis. Continuous sero-positive patients and/or nondecreased titer group, even after surgery, showed significantly unfavorable survival.

Background Although the presence of serum p53 antibody (s-p53-Abs) before treatment has been shown to correlate with poor prognosis and lymph node metastasis in esophageal cancer, there has been little information about postoperative s-p53-Abs titer and perioperative changes of s-p53-Abs titers in patients with esophageal carcinoma. Methods A highly specific enzyme-linked immunosorbent assay was used to analyze s-p53-Abs in 110 patients with esophageal squamous cell carcinoma before and 1 month after surgery. The cutoff level of 1.3 U/ml was used to indicate seropositive patients. Impact of postoperative s-p53-Abs titer and perioperative changes of s-p53-Abs on survival was evaluated. Results Forty (36%) of 110 patients were positive for s-p53-Abs before surgery and 35 patients (32%) were positive after surgery. s-p53-Abs titer generally decreased after surgery. Among sero-positive patients, the patients who remained sero-positive after surgery (n = 28) had a worse prognosis than patients who showed sero-conversion (P = 0.02). Among sero-positive patients, the nondecreased titer group showed significantly unfavorable survival (P < 0.01). Multivariate analysis revealed that postoperative s-p53-Abs was an independent risk factor for worse overall survival (adjusted hazard ratio = 3.05; 95% confidence interval = 1.11-8.33; P = 0.03). Conclusions Perioperative monitoring of s-p53-Abs titers was useful to identify patients with esophageal cancer with a high risk for tumor recurrence and a poor prognosis. Continuous sero-positive patients and/or nondecreased titer group, even after surgery, showed significantly unfavorable survival.

Based on the genetic background of cancer, we have been trying to develop novel diagnostic and therapeutic strategies against human cancers. c-myc gene activation has been detected in many human cancers, indicating a key role of c-myc in tumor development. Thus targeting c-myc gene suppression is a promising strategy for cancer treatment. Recently, an interaction between FIR (FUSE-Binding Protein-Interacting Repressor) and TFIIH/p89/XPB helicase was found to repress c-myc transcription and so might be important for suppressing tumor formation. Previously, we have shown that the expression of splicing variant of FIR is elevated in colorectal cancer tissues and promotes tumor development by disabling FIR-repression to sustain high levels of c-Myc, opposing apoptosis in cancer cells. In this study, FIR recombinant adenovirus vector induces tumor growth suppression against tumor xenografts in animal model experiment. Together, one clue to the development of cancer diagnosis and therapies directed against c-Myc may go through FIR and its splicing variant.

【背景】 本邦における原発性肝細胞癌の８割はC型肝炎ウイルスによる慢性的な肝障害に起因しており、C型肝炎ウイルスへの推定罹患者数は200万人と言われている。肝細胞癌に特異性の高い腫瘍マーカーとしてAFP・AFP-L3・PIVKA-_II_が有用とされているが、腫瘍マーカー単独での早期癌の検出率は非常に低く、確定診断には超音波検査等の画像診断や、患者への負担の大きい肝生検を必要としているのが現状である。超音波検査による小病変の拾い上げは検者の熟練度に左右されやすく、絶対的なものではない。したがって早期診断、治療方針の検討を行う上で、侵襲性が低く、簡便である血液を用いた検査として、高感度な新規腫瘍マーカーの発見が望まれる。<BR> 【方法】 血清プロテオーム解析の問題点として、「存在量の多いタンパク質の除去」・「キャリアータンパク質に結合しているタンパク質の抽出」等が挙げられる。この問題点を解決した方法として、北里大学で開発されたペプチド抽出法（K法）がある。K法はアセトン沈殿法や限外ろ過膜法に比較して低分子量タンパク質及びペプチドの抽出効率及び再現性に優れ、またアルブミン等のキャリアプロテインに結合している成分の抽出も可能にしている。 C型肝炎ウイルス感染をバックグラウンドに持つ原発性肝細胞癌患者及び肝硬変患者血清と健常者血清各5例に応用した。 血清中のペプチド成分を抽出し、逆相HPLCで分画後、各フラクションをMALDI-TOF MS（AutoFlex II, Bruker Daltonics Inc）で比較解析した。<BR> 【結果・考察】 原発性肝細胞癌患者血清中で有意に増大しているピークを３本検出し、そのうち2ピークの同定に成功した。2ピークは同一のタンパク質の断片であり、肝細胞癌患者血清での報告がなされていないペプチドである。本ペプチドは原発性肝細胞癌の診断の新たな指標になる可能性があり、残り1ピークの同定と平行して、今後多検体での検討を進めていく。

Based on the genetic background of cancer, we have been trying to develop novel diagnostic and therapeutic strategies against human cancers. c-myc gene activation has been detected in many human cancers, indicating a key role of c-myc in tumor development. Thus targeting c-myc gene suppression is a promising strategy for cancer treatment. Recently, an interaction between FIR (FUSE-Binding Protein-Interacting Repressor) and TFIIH/p89/XPB helicase was found to repress c-myc transcription and so might be important for suppressing tumor formation. Previously, we have shown that the expression of splicing variant of FIR is elevated in colorectal cancer tissues and promotes tumor development by disabling FIR-repression to sustain high levels of c-Myc, opposing apoptosis in cancer cells. In this study, FIR recombinant adenovirus vector induces tumor growth suppression against tumor xenografts in animal model experiment. Together, one clue to the development of cancer diagnosis and therapies directed against c-Myc may go through FIR and its splicing variant.

Based on the genetic background of cancer, we have been trying to develop novel diagnostic and therapeutic strategies against human cancers. c-myc gene activation has been detected in many human cancers, indicating a key role of c-myc in tumor development. Thus targeting c-myc gene suppression is a promising strategy for cancer treatment. Recently, an interaction between FIR (FUSE-Binding Protein-Interacting Repressor) and TFIIH/p89/XPB helicase was found to repress c-myc transcription and so might be important for suppressing tumor formation. Previously, we have shown that the expression of splicing variant of FIR is elevated in colorectal cancer tissues and promotes tumor development by disabling FIR-repression to sustain high levels of c-Myc, opposing apoptosis in cancer cells. In this study, FIR recombinant adenovirus vector induces tumor growth suppression against tumor xenografts in animal model experiment. Together, one clue to the development of cancer diagnosis and therapies directed against c-Myc may go through FIR and its splicing variant.

Early diagnosis of hepatocellular carcinoma (HCC) greatly improves its prognosis. However, the distinction between benign and malignant tumors is often difficult, and novel immunohistochemical markers are necessary. Using agarose two-dimensional fluorescence difference gel electrophoresis, we analyzed HCC tissues from 10 patients. The fluorescence volumes of 48 spots increased and 79 spots decreased in tumor tissues compared with adjacent nontumor tissue, and 83 proteins were identified by mass spectrometry. Immunoblot confirmed that the expression of clathrin heavy chain (CHC) and Ku86 significantly increased, whereas formiminotransferase cyclodeaminase (FTCD), rhodanese, and vinculin decreased in tumor. The protein expression in tumor and nontumor tissues was further evaluated by immunostaining. Interestingly, CHC and FTCD expression was strikingly different between tumor and nontumor tissues. The sensitivity and specificity of individual markers or a combination for the detection of HCC were 51.8% and 95.6% for CHC, 61.4% and 98.5% for FTCD, and 80.7% and 94.1% for CHC+FTCD, respectively. Strikingly, the sensitivity and specificity increased to 86.7% and 95.6% when glypican-3, another potential biomarker for HCC, was used with FTCD. Moreover, CHC and FTCD were useful to distinguish early HCC from benign tumors such as regenerative nodule or focal nodular hyperplasia, because the sensitivity and specificity of the markers are 41.2% and 77.8% for CHC, 44.4% and 80.0% for FTCD, which is comparable with those of glypican-3 (33.3% and 100%). The sensitivity significantly increased by combination of these markers, 72.2% for CHC+FTCD, and 61.1% for CHC+glypican-3 and FTCD+glypican-3, as 44.4% of glypican-3 negative early HCC were able to be detected by either CHC or FTCD staining. Conclusion: Immunostaining of CHC and FTCD could make substantial contributions to the early diagnosis of HCC.

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. OCMV-beta gal. 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.

Extensive lymphadenectomy, including upper mediastinum, for thoracic esophageal carcinoma was introduced at the beginning of 1980s. However, the efficacy has not been analyzed in large series at a single institute. We evaluated factors potentially related to improved surgical results in patients with thoracic esophageal squamous cell carcinoma (SCC). From 1959 to 1998, a total of 792 patients with thoracic esophageal SCC underwent R0 surgery. A variety of clinicopathological factors were compared among patients treated from 1990 to 1998 (recent group, n=164) and 1959 to 1989 (former group, n=628). The recent group showed significantly better survival than the former group (5-year survival rates: 51 versus 17%, P=0.01), partly because earlier stage disease was included in the recent group than in the former group. Multivariable analysis, using the Cox regression analysis, indicated the time period of surgery, age, tumor location, the number of positive nodes (&gt;5), venous invasion, and tumor-node-metastasis stage. Upper mediastinum lymphadenectomy was also an independent factor to improve survival of patients with thoracic esophageal SCC.

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. OCMV-beta gal. 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.

Background Although thoracic lymph node metastasis in patients with thoracic esophageal squamous cell carcinoma (SCC) has been reported to be a negative risk factor for long-term survival, only a few studies have evaluated the clinicopathologic difference between the impact of metastasis to the paraesophageal lymph nodes and to the nonparaesophageal lymph nodes. The purpose of this study was to evaluate surgical outcome after the clearance of metastatic thoracic lymph nodes. Methods Retrospectively reviewed were 164 consecutive patients with thoracic esophageal SCC who had not had preoperative treatment and underwent surgery from 1980 to 2005 and were found to have thoracic lymph node metastases. Of these patients, 83 underwent surgery from 1980 to 1994 and 81 from 1995 to 2005. Univariate and multivariate analyses were performed to evaluate the impact of nonparaesophageal lymph node metastasis on survival. Results Univariate analysis revealed that T3/T4 tumors and the presence of nonparaesophageal node metastases were associated with only a 20% overall five-year survival rate. The overall five-year survival for the most recent period was significantly better than for the former period (42% vs. 13%, p &lt; 0.01). Based on a multivariate analysis of prognostic impact of each nonparaesophageal node, the presence of metastatic subcarinal and/or posterior mediastinal nodes was an independent risk factor for reduced survival. Conclusion Surgical outcome for patients with thoracic esophageal cancer and metastatic thoracic lymph nodes has improved during the last 25 years. Although postoperative chemotherapy might improve survival, the presence of T3/T4 tumors and/or metastatic nonparaesophageal nodes were unfavorable factors for survival.

In recent years, the development of gene therapy systems as new treatment or prevention strategies for various malignant diseases has been explored. Based on the genetic background of esophageal cancer, several molecular therapies have been developed. In this article, we review p53 genetic alterations and angiogenesis in esophageal cancer. Our recent clinical results from a phase I/II study of adenoviral-mediated p53 gene therapy for patients with un-resectable esophageal cancer are reviewed. Lastly we review the available experimental therapeutic models from the literature and our experimental work over the past few years are reviewed. (Ann Thorac Cardiovasc Surg 2008; 14: 3-8)

Background Although thoracic lymph node metastasis in patients with thoracic esophageal squamous cell carcinoma (SCC) has been reported to be a negative risk factor for long-term survival, only a few studies have evaluated the clinicopathologic difference between the impact of metastasis to the paraesophageal lymph nodes and to the nonparaesophageal lymph nodes. The purpose of this study was to evaluate surgical outcome after the clearance of metastatic thoracic lymph nodes. Methods Retrospectively reviewed were 164 consecutive patients with thoracic esophageal SCC who had not had preoperative treatment and underwent surgery from 1980 to 2005 and were found to have thoracic lymph node metastases. Of these patients, 83 underwent surgery from 1980 to 1994 and 81 from 1995 to 2005. Univariate and multivariate analyses were performed to evaluate the impact of nonparaesophageal lymph node metastasis on survival. Results Univariate analysis revealed that T3/T4 tumors and the presence of nonparaesophageal node metastases were associated with only a 20% overall five-year survival rate. The overall five-year survival for the most recent period was significantly better than for the former period (42% vs. 13%, p &lt; 0.01). Based on a multivariate analysis of prognostic impact of each nonparaesophageal node, the presence of metastatic subcarinal and/or posterior mediastinal nodes was an independent risk factor for reduced survival. Conclusion Surgical outcome for patients with thoracic esophageal cancer and metastatic thoracic lymph nodes has improved during the last 25 years. Although postoperative chemotherapy might improve survival, the presence of T3/T4 tumors and/or metastatic nonparaesophageal nodes were unfavorable factors for survival.

Background: p53 gene therapy has been examined in several clinical trials, however, the results of those trials have mostly been unsatisfactory due to the low efficacy of this therapy. Leptomycin B (LMB) is an antibiotic originally isolated from Streptomyces that has the ability to inhibit the export of proteins containing a nuclear export signal from the nucleus to the cytoplasm. Currently, it has been shown that p53 protein has a nuclear export signal. In this study, we assessed whether LMB augments the transduced p53 gene effect. Methods: Antiproliferative effect of LMB was assessed in human esophageal squamous cancer cell lines. Accumulation of p53 protein into the nucleus by LMB was observed by fluorescence microscopy. The combined effect of p53 and LMB was evaluated in in vitro experiments. Results: LMB induced cell death in a dose-dependent manner and p53 drastically accumulated in the nucleus after LMB treatment. The combinatory treatment of p53 gene and LMB significantly increases the efficiency compared to either agent alone. Conclusions: Our findings suggest that LMB has a potent ability to augment the effect of the tumor suppressor p53 in esophageal squamous cancer cell lines and that it is a promising component in p53 gene therapy. Copyright (C) 2008 S. Karger AG, Basel.

Peroxiredoxins (Prdxs) are a family of antioxidant enzymes that are also known as scavengers of peroxide in mammalian cells. Some reports have shown that the overexpression of Prdx1, which is one of the peroxiredoxins that is a ubiquitously expressed protein, was related to a poor prognosis in several types of human cancers. In this study, we investigated the expression levels of Prdx1 in esophageal squamous cell carcinoma by immunohistochemistry, and the correlation between the Prdx1 expression and the clinical status was elucidated. Immunohistochemical staining was performed in 114 samples which were collected from surgical esophageal cancer specimens. Cytoplasmic staining of Prdx1 was evaluated based on the following scoring criteria: Grade I, negative or weak staining; Grade II, moderate staining; and Grade III, strong staining. The percentage of patients with a Grade I expression of Prx1 was 20% (23 of 114), 44% had Grade II (50 of 114), and 36% had Grade 111 (41 of 114). The Prdx1 immunoreactivity showed an inverse significant correlation with T-category (P &lt; 0.0001), lymph node metastasis (P=0.048), and stage (P=0.001). In addition, the patients with tumors exhibiting a reduced Prdx1 expression had shorter overall survival (P=0.022) in comparison to the patients with tumors which had a higher Prdx1 expression. Currently, Prdx1 has been shown to act as a tumor suppressor. Our results provide strong evidence that the reduced Prdx1 expression is an important factor in esophageal squamous cancer progression and could serve as a useful prognostic marker.

Peroxiredoxins (Prdxs) are a family of antioxidant enzymes that are also known as scavengers of peroxide in mammalian cells. Some reports have shown that the overexpression of Prdx1, which is one of the peroxiredoxins that is a ubiquitously expressed protein, was related to a poor prognosis in several types of human cancers. In this study, we investigated the expression levels of Prdx1 in esophageal squamous cell carcinoma by immunohistochemistry, and the correlation between the Prdx1 expression and the clinical status was elucidated. Immunohistochemical staining was performed in 114 samples which were collected from surgical esophageal cancer specimens. Cytoplasmic staining of Prdx1 was evaluated based on the following scoring criteria: Grade I, negative or weak staining; Grade II, moderate staining; and Grade III, strong staining. The percentage of patients with a Grade I expression of Prx1 was 20% (23 of 114), 44% had Grade II (50 of 114), and 36% had Grade 111 (41 of 114). The Prdx1 immunoreactivity showed an inverse significant correlation with T-category (P &lt; 0.0001), lymph node metastasis (P=0.048), and stage (P=0.001). In addition, the patients with tumors exhibiting a reduced Prdx1 expression had shorter overall survival (P=0.022) in comparison to the patients with tumors which had a higher Prdx1 expression. Currently, Prdx1 has been shown to act as a tumor suppressor. Our results provide strong evidence that the reduced Prdx1 expression is an important factor in esophageal squamous cancer progression and could serve as a useful prognostic marker.

Histone deacetylase inhibitors (HDACIs) are promising therapeutic agents with the potential for regulating cell cycle, differentiation and apoptosis in cancer cells. HDACI activity is associated with selective transcriptional regulation and altering gene expression. However, the exact mechanisms leading to the antitumor effect of HDACIs are not fully understood. FK228, one of the powerful HDACIs, strongly inhibited cell growth of T.Tn and TE2 cells and induced apoptosis. Therefore, comprehensive analysis of the changes in gene expression in human esophageal cancer cell lines by the HDACI FK228 was carried out by microarray analysis. This analysis was used to clarify the expression profiles of genes after exposure to FK228. Of the 4,608 genes analyzed, 93 genes in T.Tn and 65 genes in TE2 were up- or down-reaulated 2-fold or more at least at one time point during FK228 exposure and they were classified into four clusters based on their expression patterns. Among them, 15 genes were contained in both cell lines and their expression patterns were similar. Except p21, Prdx1 (reported by us) and IGFBP3, the behaviour/expression of 12 highly responsive genes has still not been reported in esophageal cancer cells. These observations of the expression patterns of functionally classified genes provided insights into the mechanism of the antitumor effect of FK228 in esophageal squamous carcinoma.

Histone deacetylase inhibitors (HDACIs) are promising therapeutic agents with the potential for regulating cell cycle, differentiation and apoptosis in cancer cells. HDACI activity is associated with selective transcriptional regulation and altering gene expression. However, the exact mechanisms leading to the antitumor effect of HDACIs are not fully understood. FK228, one of the powerful HDACIs, strongly inhibited cell growth of T.Tn and TE2 cells and induced apoptosis. Therefore, comprehensive analysis of the changes in gene expression in human esophageal cancer cell lines by the HDACI FK228 was carried out by microarray analysis. This analysis was used to clarify the expression profiles of genes after exposure to FK228. Of the 4,608 genes analyzed, 93 genes in T.Tn and 65 genes in TE2 were up- or down-reaulated 2-fold or more at least at one time point during FK228 exposure and they were classified into four clusters based on their expression patterns. Among them, 15 genes were contained in both cell lines and their expression patterns were similar. Except p21, Prdx1 (reported by us) and IGFBP3, the behaviour/expression of 12 highly responsive genes has still not been reported in esophageal cancer cells. These observations of the expression patterns of functionally classified genes provided insights into the mechanism of the antitumor effect of FK228 in esophageal squamous carcinoma.

Background: This study aimed to evaluate the surgical outcome of esophagectomy in patients 75 years of age and older with thoracic esophageal carcinoma. Patients and Methods: Between 1980 and 2002, 55 (46%) of 120 patients 75 years of age and older with thoracic esophageal carcinoma underwent an esophagectomy. The risk factors that resulted in decreased survival were analyzed by both univariate and multivariate analyses. Differences in surgical outcome and long-term survival between the earlier time period (1980-1989) and later time period (1990-2002) were analyzed separately. Results: Overall resection rate in elderly patients in both periods was similar (44%, earlier period; 46%, later period). Postoperative complications significantly reduced long-term survival (adjusted hazard ratio for death, 4.05; 95% confidence interval (CI), 1.70-9.62; P &lt; 0.01). Surgical blood loss greater than 1,000 ml was less frequently observed in the later period than in the earlier period (19% vs. 54%, P = 0.01). The postoperative morbidity rate was lower in the later than in the earlier period (29% vs. 63%, P = 0.02). Overall 5-year survival rate was significantly higher in the later period than in the earlier period (57% vs. 18%, P &lt; 0.01). Conclusions: Elderly patients who underwent an esophagectomy in the later period appeared to manifest less neoadjuvant treatment, less surgical stress, fewer postoperative complications, and a better long-term survival than those treated in the earlier period.

Background: The location and clinical impact of solitary lymph node metastasis from thoracic esophageal carcinoma have not been evaluated sufficiently. Methods: A consecutive series of 91 patients with a solitary positive lymph node who underwent curative surgery for thoracic esophageal carcinoma was investigated. The prognostic impact was evaluated by univariate analysis and multivariate analysis using Cox's proportional hazards model. Results: A total of 52 (57%) of the 91 patients showed a solitary positive node beyond the thorax. While 29% of the patients with an upper thoracic tumor showed a cervical node, 13% of the patients with a middle tumor and none of the patients with a lower tumor showed a cervical node. Tumor depth and venous invasion were found to be independent risk factors for poor survival. Conclusions: The solitary positive lymph nodes were broadly distributed depending on the tumor location and tumor depth. Tumor depth and venous invasion were risk factors for poor survival in these patients. (c) 2006 Excerpta Medica Inc. All rights reserved.

Purpose. Clinical trials of carbon ion therapy have been performed due to the advantages of high-dose energy delivery with precise localization control to targeted organs and strong cell-killing activities to cancers. Perforated intestines as a late morbidity after carbon ion radiotherapy for uterine cancers were examined to reveal the biological characteristics of carbon ion for future applications for the treatment of gastrointestinal cancers. Methods. Between June 1995 and December 2004, 94 patients with carcinoma of the uterine cervix or corpus were treated with carbon ion therapy. Among them, 9 patients (9.6%) developed major late gastrointestinal (GI) complications. Four out of 9 patients had intestinal perforations excised operatively at our institute. The postoperative clinical courses and histopathological findings of the excised intestine were investigated. Results. Carbon ion irradiation severely damaged smooth muscle layers by coagulation necrosis as well as atrophy of the intestinal epithelium and middle-sized arterial thromboses of the intestines. After evaluating late complications, the dose constraints on the GI tracts were set under 60 GyE to prevent major complications. Thereafter, the incidence of major GI complications markedly decreased. Conclusion. Our findings demonstrated the characteristic histopathological effects of carbon ion radiotherapy and thus are expected to facilitate future additional applications of carbon ion radiotherapy for the treatment of gastrointestinal cancers.