bluuebunny/crossref_metadata_2025 huggingface dataset:
Reduced Expression of Laminin α3 and α5 Chains in Non‐small Cell Lung Cancers
Authors: Akashi Takumi , Ito Eisaku , Eishi Yoshinobu , Koike Morio , Nakamura Kyoichi , Burgeson Robert E.
Published: August 2005

The basement membrane is considered to act as a barrier which hinders cancer cells from invading the surrounding stroma. In order to assess changes in essential components during neoplasia in the lung, we immunohistochemically studied distribution patterns of laminins α3 and α5 in 40 adeno‐carcinomas and 8 squamous cell carcinomas. The α5 chain was generally preserved at the periphery, frequently disrupted in foci with alveolar collapse and absent in foci of fibroblastic proliferation within adenocarcinomas. Fragmentation and absence of laminin α3 chain were more prominent than for α5 chain. Laminin α3 chain was partially fragmented or absent in peripheral areas of adenocarcinomas, being significantly different from α5 chain. Non‐small cell lung cancers with reduced α5 chain showed a tendency for greater lymph node metastasis. In cultured normal air way epithelial cells, both laminin α3 and α5 chains were found to be expressed by northern analysis. Eleven of the twelve cultured lung cancer cell lines did not express a3 chain and expression of α5 chain was reduced in three. Quantitative RT‐PCR analysis also demonstrated expression of laminin α3 chain in adenocarcinoma tissues to be significantly lower than in normal lung tissues. These results suggest that expression of laminin a chains is often reduced in lung cancer cells and this might contribute to basement membrane fragmentation and subsequent proliferation of stromal elements, as well as play some role in the process of cancer cell invasion.
Efficient Gene Transduction by RGB‐fiber Modified Recombinant Adenovirus into Dendritic Cells
Authors: Asada‐Mikami Rumiko , Heike Yuji , Kanai Sachiyo , Azuma Masato , Shirakawa Kazuo , Takaue Yoichi , Krasnykh Victor , Curiel David T. , Terada Masaaki , Abe Tatsuo , Wakasugi Hiro
Published: August 2005

Dendritic cells (DC) are important antigen‐presenting cells in the development of an anti‐tumor T cell response. To extend the range of current immuno/gene therapies, we tested luciferase‐expressing RGD‐adenovirus (Ad) (Ad5lucRGD)‐mediated transduction into DC. Phenotypically characterized DC were generated from peripheral blood CD14+ cells by incubation with granulocyte‐macrophage colony‐stimulating factor, interleukin‐4 and tumor necrosis factor a. On the 7th day of culture, the cells became mature DC with a CD1a+, CD11c+, CD80+, CD83+, CD86+, human leukocyte antigen (HLA)‐DR+, CD14− phenotype. The expression of (α,β3 integrin was enhanced on day 3 and returned to the basal level on day 7. We then compared the transduction efficiency of an AdSlucRGD system to that using conventional Ad, in cells harvested on days 1, 3 and 7 of culture. Luciferase activity was negligible in AdCMVLuc, but remarkable in cells processed with Ad5lucRGD. Activity was maximal in cells that had been cultured for 3 days. Recombinant Ad5 fiber knob protein blocked AdCMVLuc‐ and Ad5lucRGD‐mediated gene transduction by 90% and 20%, respectively. Surface markers and cytokine production were not affected by Ad5lucRGD‐mediated transduction.
Antitumor Activity and Pharmacokinetics of TAS‐106, l‐(3‐C‐Ethynyl‐β‐D‐ribo‐pentofuranosyl)cytosine
Authors: Shimamoto Yuji , Fujioka Akio , Kazuno Hiromi , Murakami Yuko , Ohshimo Hideyuki , Kato Toshiyuki , Matsuda Akira , Sasaki Takuma , Fukushima Masakazu
Published: August 2005

We examined the effects of dosage schedule on antitumor activity in vitro and in vivo to determine the optimal administration schedule for a new nucleoside antimetabolite l‐(3‐C‐ethynyl‐β‐D‐ribo‐pentofuranosyl)cytosine (ECyd, TAS‐106). The cytotoxicity of TAS‐106 in vitro against human tumors was evaluated at three drug exposure periods. TAS‐106 exhibited fairly potent cytotoxicity even with 4 h exposure, and nearly equivalent and sufficiently potent cytotoxicity with 24 and 72 h exposures. These results suggest that long‐term exposure to TAS‐106 will not be required to achieve maximal cytotoxicity. The antitumor activity of TAS‐106 in vivo was compared in nude rat models bearing human tumors on three administration schedules, once weekly, 3 tunes weekly, and 5 tunes weekly for 2 or 4 consecutive weeks. TAS‐106 showed strong antitumor activity without serious toxicity on all three schedules, but the antitumor activity showed no obvious schedule‐dependency in these models. When tumor‐bearing nude rats were given a single i.v. dose of [3H]TAS‐106, tumor tissue radioactivity tended to remain high for longer periods of time as compared to the radioactivity in various normal tissues. Furthermore, when the metabolism of TAS‐106 in the tumor was examined, it was found that TAS‐106 nucleotides (including the active metabolite, the triphosphate of TAS‐106) were retained at high concentrations for prolonged periods. These pharmacodynamic features of TAS‐106 may explain the strong antitumor activity without serious toxicity, observed on intermittent administration schedules, in nude rat models with human tumors. We therefore consider TAS‐106 to be a promising compound which merits further investigation in patients with solid tumors.
Expression of gp34 (OX40 Ligand) and OX40 on Human T Cell Clones
Authors: Takasawa Naruhiko , Ishii Naoto , Higashimura Norikazu , Murata Kazuko , Tanaka Yuetsu , Nakamura Masataka , Sasaki Takeshi , Sugamura Kazuo
Published: August 2005

gp34, which we previously cloned, is a ligand of OX40 (CD 134), a costimulatory molecule involved in T cell activation. To elucidate the role of human OX40/OX40L interaction, we examined the expression of gp34 (OX40L) and OX40 in normal human hematopoietic cells by using flow cytometry. OX40 expression is observed on activated T cells, while OX40L is expressed in antigen‐presenting cells. However, cytotoxic T lymphocyte (CTL) clones specific for Epstein‐Barr virus (EBV)‐transformed autologous lymphoblastic cell lines (LCLs) induced both OX40 and OX40L expression after antigen or T cell receptor (TCR) stimulation. This study suggests a possible function of OX40L/OX40, through T cell‐T cell interaction, in the reactivation of memory T cells in an auto‐crine manner, with implications for the pathogenesis of viral infections and neoplasms.
Down‐regulation in Multiple Human Cancers of a Novel Gene, DMHC, from 17q25.1 That Encodes an Integral Membrane Protein
Authors: Mikami Iwao , Harada Haruhito , Nagai Hisaki , Tsuneizumi Michiko , Nobe Yukiko , Koizumi Kiyoshi , Sugano Sumio , Tanaka Shigeo , Emi Mitsuru
Published: August 2005

Frequent observations of allelic loss in chromosomal band 17q25.1 in a variety of human cancers have suggested that one or more tumor suppressor genes are present in that region. Moreover, a genetic locus for hereditary focal non‐epidermolytic palmoplantar keratoderma, a condition associated with cancer of the esophagus (TOC; Tylosis with Oesophageal Cancer), lies in the same region. We screened cell lines derived from a variety of human cancers by reverse transcription‐polymerase chain reaction (RT‐PCR) to detect alterations in expression of genes within the region in question, by examining expressed sequence tags located there. These experiments identified an 1834‐bp full‐length cDNA encoding a novel, 441‐amino acid integral membrane protein with seven putative transmembrane domains. This gene showed loss or extreme decrease of expression in 6 of 10 uterine cancer‐cell lines, 2 of 11 hepatic cell carcinoma‐cell lines, 2 of 7 lung cancer‐cell lines, 1 of 6 gastric cancer‐cell lines, and 1 of 10 breast cancer‐cell lines. (We named it DMHC (“down‐regulated in multiple human cancers”).) Our results suggest that loss of expression of DMHC at 17q25.1 may play an important role in development of variety of human cancers.
Co‐transduction of Apaf‐1 and Caspase‐9 Augments Etoposide‐induced Apoptosis in U‐373MG Glioma Cells
Authors: Shinoura Nobusada , Sakurai Shinya , Asai Akio , Kirino Takaaki , Hamada Hirofumi
Published: August 2005

Several apoptosis‐related genes have been reported to be involved in chemotherapy‐induced apoptosis in cancers. An assessment of the relationship between expression of those genes and the degree of chemotherapy‐induced apoptosis may be useful in improving the efficacy of cancer therapy. We transduced Apaf‐1 (apoptotic protease‐activating factor‐1) and caspase‐9 into U‐373MG glioma cells using adenovirus (Adv) vectors in the presence of etoposide and evaluated the degree of apoptosis. The degree of apoptosis in etoposide‐treated U‐373MG cells infected with Adv for Apaf‐1 (Adv‐APAFl) was higher (27%) than that in cells infected with control Adv (14%), that in cells infected with Adv for caspase‐9 (Adv‐Casp9) was higher (34%) than that in cells infected with Adv‐APAFl, and that in cells infected with both Adv‐APAFl and Adv‐Casp9 was the highest (41%). Treatment with etoposide increased expression of p53 and decreased expression of Bcl‐XL in U‐373MG cells which harbored mutant p53. These results indicate that the expression of Apaf‐1 and caspase‐9 may be important determinants in predicting the sensitivity of cancers to chemotherapy. Adv‐mediated co‐transduction of Apaf‐1 and caspase‐9 should render cancer cells highly sensitive to chemotherapy.
Contribution of Caspase‐3 Differs by p53 Status in Apoptosis Induced by X‐Irradiation
Authors: Kobayashi Daisuke , Tokino Takashi , Watanabe Naoki
Published: August 2005

We investigated the effect of p53 status on involvement of caspase‐3 activation in cell death induced by X‐irradiation, using rat embryonic fibroblasts (REFs) transduced with a temperature‐sensitive mutant (mt) p53 gene. Cells with wild‐type (wt) p53 showed greater resistance to X‐irradiation than cells with mt p53. In cells with wt p53, X‐irradiation‐induced apoptosis was not inhibited by the caspase‐3 inhibitor acetyl‐L‐aspartyl‐L‐methionyl‐L‐glutaminyl‐L‐aspartyl‐aldehyde (Ac‐DMQD‐CHO) and caspase‐3 activity was not elevated following X‐irradiation, although induction of p53 and p21/WAF‐l protein was observed. In contrast, irradiated cells with mt p53 showed 89% inhibition of cell death with Ac‐DMQD‐CHO and 98% inhibition with the antioxidant N‐acetyl‐L‐cysteine (NAC). In cells with mt p53, caspase‐3 activity was increased approximately 5 times beyond baseline activity at 24 h after irradiation. This increase was almost completely inhibited by NAC. However, inhibition of caspase‐3 by Ac‐DMQD‐CHO failed to decrease production of reactive oxygen species by cells with mt p53. Differential involvement of caspase‐3 is a reason for differences in sensitivity to X‐irradiation in cells with different p53 status. Caspase‐3 activation appears to occur downstream from generation of reactive oxygen species occurring independently of wt p53 during X‐irradiation‐induced cell death.
Expression of CYP2A6 in Tumor Cells Augments Cellular Sensitivity to Tegafur
Authors: Murayama Norie , Sai Kimie , Nakajima Yukiko , Kaniwa Naoko , Ozawa Shogo , Ohno Yasuo , Sawada Jun‐ichi
Published: August 2005

To examine the role of cytochrome P450 2A6 (CYP2A6) in the cellular sensitivity to an anti‐tumor prodrug, tegafur (FT), a CYP2A6 cDNA construct was transfected into cells of a colon cancer cell line, DLD‐1. CYP2A6‐expressing cells (DLD‐1/CYP2A6 cells) more efficiently catalyzed the conversion of FT to 5‐fluorouracil (5‐FU) (2.6‐fold) and the 7‐hydroxylation of coumarin (7.9‐fold) than cells transfected with a null construct (DLD‐1/null cells). These results indicated that the expressed CYP2A6 was functionally active. The extent of growth inhibition of the DLD‐1/CYP2A6 cells by FT was greater than that of DLD‐1/null cells; the difference between the DLD‐1/CYP2A6 and DLD‐1/null cells was statistically significant at the concentrations of 250, 500 and 1000 μM. 5‐FU, an active metabolite of FT, inhibited the growth of both types of cells to the same extent. Thus, intracellular expression of CYP2A6 can sensitize cells to FT.
UCN‐01 (7‐Hydoxystaurosporine) Inhibits in vivo Growth of Human Cancer Cells through Selective Perturbation of G1 Phase Checkpoint Machinery
Authors: Abe Sadanori , Kubota Tetsuro , Otani Yoshihide , Furukawa Toshiharu , Watanabe Masahiko , Kumai Koichiro , Akiyama Tadakazu , Akinaga Shiro , Kitajima Masaki
Published: August 2005

Mechanisms underlying tumor sensitivity to the antitumor agent UCN‐01 (7‐hydroxystaurosporine) were examined in the nude mouse model using three human tumor xenografts, two pancreatic cancers (PAN‐3‐JCK and CRL 1420) and a breast cancer (MX‐1). UCN‐01 antitumor activity was evaluated in terms of relative tumor weights in treated and untreated mice bearing the tumor xenografts. The activity of cyclin‐dependent kinase 2 (CDK2), levels of p21 and p27 proteins, pRb status and cell cycle were evaluated. Induction of p21 and apoptosis were also assessed immuno‐histochemically in CRL 1420. UCN‐01 was administered intraperitoneally at a dose of either 5 or 10 mg/kg daily for 5 days followed by a further 5 injections after an interval of 2 days. UCN‐01 significantly suppressed the growth of both pancreatic cancers, but was ineffective against MX‐1. p21 protein expression was markedly induced in the UCN‐01‐sensitive pancreatic carcinoma xenografts at both doses, but p21 induction was only evident in the UCN‐01‐resistant MX‐1 at 10 mg/kg. MX‐1 exhibited CDK2 activity that was 6‐fold higher than that of pancreatic cancer strains, which may explain the resistance of MX‐1 to UCN‐01 despite the induction of p21 at the dose of 10 mg/kg. The UCN‐01‐sensitive tumors exhibited G1 arrest and increased levels of apoptosis, changes not observed in resistant MX‐1. In conclusion, it appears that a determining factor ofin vivoUCN‐01 sensitivity involves the balance of CDK2 kinase activity and p21 protein induction, resulting in augmented pRb phosphorylation, G1 cell cycle arrest and apoptosis.
Monitoring of Intracellular l‐βD‐Arabinofuranosylcytosine 5′‐Triphosphate in 1‐β‐D‐Arabinofuranosylcytosine Therapy at Low and Conventional Doses
Authors: Yamauchi Takahiro , Kawai Yasukazu , Kishi Shinji , Goto Nobuyuki , Urasaki Yoshimasa , Imamura Shin , Fukushima Toshihiro , Yoshida Akira , Iwasaki Hiromichi , Tsutani Hiroshi , Masada Mikio , Ueda Takanori
Published: August 2005

1‐β‐D‐Arabinofuranosylcytosine (ara‐C) is used empirically at a low, conventional, or high dose. Ara‐C therapy may be optimal if it is directed by the clinical pharmacokinetics of the intracellular active metabolite of ara‐C, 1‐β‐D‐arabinofuranosylcytosine 5′‐triphosphate (ara‐CTP). However, ara‐CTP has seldom been monitored during low‐ and conventional‐dose ara‐C therapies because detection methods were insufficiently sensitive. Here, with the use of our newly established method (Cancer Res., 56, 1800‐1804 (1996), ara‐CTP was monitored in leukemic cells from acute myelog‐enous leukemia patients receiving low‐ or conventional‐dose ara‐C [subcutaneous ara‐C administration (10 mg/m2) (3 patients), continuous ara‐C infusion (20 or 70 mg/m2/24 h) (7 patients), 2‐h ara‐C infusion (70 mg/m2) (4 patients), and 2‐h infusion of N4‐behenoyl‐l‐β‐D‐arabinofuranosylcy‐tosine, a deaminase‐resistant ara‐C derivative (70 mg/m2) (6 patients)]. Ara‐CTP could be determined at levels under 1μM. There was a close correlation between the elimination half‐life values of the plasma ara‐C and the intracellular ara‐CTP. The presence of ara‐C in the plasma was important to maintain ara‐CTP. The continuous ara‐C and the 2‐h N4‐behenoyl‐l‐β‐D‐arabinofura‐nosylcytosine infusions maintained ara‐CTP and the plasma ara‐C longer than the subcutaneous ara‐C or the 2‐h ara‐C infusion. They also afforded relatively higher ara‐CTP concentrations, and consequently produced ara‐CTP more efficiently than the 2‐h ara‐C infusion. Different administration methods produced different quantities of ara‐CTP even at the same dose.
CPT‐11 Alters the Circadian Rhythm of Dihydropyrimidine Dehydrogenase mRNA in Mouse Liver
Authors: Shimizu Mikiko , Tamura Tomohide , Yamada Yasuhide , Akiyama Yoshiko , Saijo Nagahiro , Nishio Kazuto
Published: August 2005

Combination chemotherapy consisting of 5‐fluorouracil (5‐FU) and 7‐ethyl‐10‐[4‐(l‐piperidino)‐l‐piperidino]carboxycamptothecin (CPT‐11) is a promising regimen for gastrointestinal cancer. The circadian‐dependent efficacy and toxicity of 5‐FU are related to the circadian variation in the activity of dihydropyrimidine dehydrogenase (DPD), which is a rate‐limiting enzyme in the pyrimidine catabolic pathway. To optimize the schedule of the CPT‐11 plus 5‐FU combination, we investigated the effect of CPT‐11 on the circadian rhythm of DPD in vivo. In control mice, the DPD mRNA level in the liver was significantly higher at 14:00 than that at 02:00. After intravenous administration of CPT‐11 (30 mg/kg) at 20:00, the circadian rhythm of the DPD mRNA level in the liver was no longer observed 18 h later (14:00), but it was unaffected 6 and 18 h later (at 14:00 and 02:00) when CPT‐11 was given at 08:00. In addition, a dose‐dependent lengthening of the period of the circadian rhythm of DPD was observed for 42 h after intravenous injection of CPT‐11 at 20:00. The levels of DPD protein and activity at 21 h after administration of CPT‐11 (at 17:00) were significantly higher than at 9 h (at 05:00). These results suggest that CPT‐11 may influence the circadian rhythm of DPD at the transcriptional level. Modulation of the circadian rhythm of DPD by CPT‐11 may be a factor in optimizing the combination of 5‐FU and CPT‐11.
Frameshift Mutations at Mononucleotide Repeats in RAD50 Recombinational DNA Repair Gene in Colorectal Cancers with Microsatellite Instability
Authors: Ikenoue Tsuneo , Togo Goichi , Nagai Keiichi , Ijichi Hideaki , Kato Jun , Yamaji Yutaka , Okamoto Makoto , Kato Naoya , Kawabe Takao , Tanaka Atsushi , Matsumura Masayuki , Shiratori Yasushi , Omata Masao
Published: August 2005

To identify additional genes targeted for microsatellite instability (MSI), we search for human genes which contain mononucleotide repeats in their coding region, selected 7 genes (RAD50, DNA‐PKcs, FLASH, Apaf‐1, XPG, CtIP, and MLSN1), and analyzed frameshift mutations in them. Here we report that 60% (3 out of 5) of human colorectal cancer cell lines exhibiting a high frequency of MSI (MSI‐H) and 46% (6 out of 13) of MSI‐H primary colorectal tumors had mutations in the (A)9 repeat of RAD50 recombinational repair gene. In contrast, no frameshift mutations were found in any of the 5 MSI‐negative colorectal cancer cell lines, 8 colorectal tumors exhibiting a low frequency of MSI (MSI‐L), or 28 MSI‐negative colorectal tumors. No mutations were found in the mononucleotide repeats of 6 other genes, even in MSI‐H cancers. These results suggest that RAD50 frameshift mutations may play a role in the tumorigenesis of MSI‐H colorectal cancers.
Suppression of Occurrence and Advancement of β‐Catenin‐accumulated Crypts, Possible Premalignant Lesions of Colon Cancer, by Selective Cyclooxygenase‐2 Inhibitor, Celecoxib
Authors: Yamada Yasuhiro , Yoshimi Naoki , Hirose Yoshinobu , Hara Akira , Shimizu Masahito , Kuno Toshiya , Katayama Masaki , Qiao Zheng , Mori Hideki
Published: August 2005

Suppression of occurrence and advancement of premalignant lesions is important for cancer prevention. Our previous studies clarified that β‐catenin‐accumulated crypts, independent of aberrant crypt foci (ACF), are probably direct precursors of colon cancers in rats. Here we investigated the effects of a selective cyclooxygenase‐2 inhibitor, celecoxib, on the development of β‐catenin‐accumulated crypts in comparison with those on ACF. Male F344 rats were divided into 4 groups. Groups 1‐3 were administered azoxymethane (AOM) s.c. at a dose of 15 mg/kg body weight, once weekly for 3 weeks to induce β‐catenin‐accumulated crypts. Groups 2 and 3 also received experimental diet containing celecoxib (500 and 1500 ppm, respectively) for 8 weeks, starting a week before the first dosing of AOM. At termination, the frequency and crypt multiplicity (number of crypts/lesion) of β‐catenin‐accumulated crypts of groups 2 and 3 were significantly less than that of group 1. Furthermore, numbers of silver‐stained nucleolar organizer regions (AgNORs)/nucleus in β‐catenin‐accumulated crypts were also decreased by exposure to celecoxib. In this study, celecoxib had greater effects on the frequency and growth of β‐catenin‐accumulated crypts than on those of ACF. These findings represent additional evidence that β‐catenin‐accumulated crypts are premalignant lesions of colon cancer. The results also suggest that β‐catenin‐accumulated crypts could be a novel target for evaluation of possible chemopreventive agents against colon carcino‐genesis, and indicate that possible chemopreventive effects of celecoxib on the initial stage of colon carcinogenesis may be related to modulation of cell proliferation activity in such early lesions.
Quantitative Analysis of BRCA1 and BRCA2 mRNA Expression in Sporadic Breast Carcinomas and Its Relationship with Clinicopathological Characteristics
Authors: Egawa Chiyomi , Miyoshi Yasuo , Taguchi Tetsuya , Tamaki Yasuhiro , Noguchi Shinzaburo
Published: August 2005

BRCA1 and BRCA2 mRNA expression in sporadic breast cancers was quantified by a real‐time reverse transcriptase‐polymerase chain reaction (RT‐PCR), and the relationship of their expression with various Clinicopathological factors was studied. BRCA2 mRNA levels (0.993±1.395, mean±SD (BRCA2/β‐glucuronidase mRNA ratios)) were significantly (P<0.01) higher than BRCA1 mRNA levels (0.519±0.570 (BRCAl/p‐glucuronidase mRNA ratios)), and a weak but significant (r=0.390, P<0.01) correlation was observed between BRCA1 and BRCA2 mRNA expression levels. There was no significant association between BRCA1 mRNA expression and Clinicopathological factors such as menstrual status, tumor size, lymph node status, estrogen and progesterone receptor status, and histological grade. On the other hand, there was a significant association between higher BRCA2 mRNA expression and estrogen receptor (ER) negativity (P<0.01) or progesterone receptor (PR) negativity (P<0.01) or high histological grade (P<0.01). These results suggest a differential contribution of BRCA1 and BRCA2 in the pathogenesis of sporadic breast cancers. BRCA2 mRNA is speculated to be up‐regulated in response to proliferation and genomic instability in high histological grade tumors.
Expression of Apoptosis Regulatory Proteins in the Skeletal Muscle of Tumorbearing Rabbits
Authors: Yoshida Hiroyuki , Ishiko Osamu , Sumi Toshiyuki , Honda Kenichi , Hirai Kouzo , Ogita Sachio
Published: August 2005

We reported finding that apoptosis occurred in skeletal muscle in the early stage after implantation. In the present study, we investigated expression of the apoptosis‐related proteins Bax and Bcl‐2 to determine the mechanism of the apoptosis. In the early stage of tumor bearing, 20 days after implantation, lean body mass (LBM) was reduced by 5.06±1.10% in the tumor‐bearing group, compared with an increase of 4.96±1.26% in the control group. The apoptotic index (AI) of the skeletal muscle in the tumor‐bearing group increased to 40.5±3.20% but was 0% in the control group, and Bax expression was strongly positive in 5 of the 10 rabbits in the tumor‐bearing group, and significantly stronger than in the control group (P=0.0002). In the late stage of tumor bearing, 40 days after implantation, the AI had declined to 0.93±0.96% in the tumor‐bearing group, but was still 0% in the control group. Bax expression was rarely detected in either the tumor‐bearing group or the control group, and there was no significant difference between the two groups (P=0.706). No significant changes in Bcl‐2 were observed in either group. The above results showed that apoptosis via Bax played a role in muscle wasting associated with progression of the malignant tumor. However, the apoptosis and expression of Bax were seen only in the early stage, within 20 days after implantation, not in the late stage. This suggested that the muscle wasting in the early stage might be caused by a different mechanism from that in the late stage.
Large‐scale Analysis of Mutations in RET Exon 16 in Sporadic Medullary Thyroid Carcinomas in Japan
Authors: Takano Torn , Miyauchi Akira , Yoshida Hiroshi , Hasegawa Yukiko , Kuma Kanji , Amino Nobuyuki
Published: August 2005

Germline mutations in the RET proto‐oncogene are the cause of multiple endocrine neoplasia type 2 (MEN 2A and 2B) and familial medullary thyroid carcinoma (FMTC). Some cases of sporadic medullary thyroid carcinoma (MTC) have also been reported to have mutations in the RET gene. However, two previous reports have given discrepant results on the frequency of the mutations in RET in sporadic MTCs in Japan. To clarify this problem, we analyzed mutations in RET exon 16 in 72 sporadic MTCs by means of the two methods used in the previous studies, direct sequencing and polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP). Mutations in exon 16 were detected in only 2 of 72 cases of sporadic MTC. These results suggest that when a MTC has a mutation in RET exon 16, it is more likely to be a hereditary MTC than a sporadic one in Japan.
A Novel Method for Detecting Single Glandular Intestinal Metaplasia in the Mucosal Surface of the Fixed Stomach Using Methylene Blue
Authors: Nomura Sachiyo , Tatemichi Masayuki , Kaminishi Michio , Esumi Hiroyasu
Published: August 2005

A close association between intestinal metaplasia of the stomach and the well‐differentiated type of gastric cancer is well recognized. The etiological relationship and how intestinal metaplasia contributes to gastric carcinogenesis are, however, still unclear. In order to answer this question, precise mapping and identification of the smallest lesion of intestinal metaplasia are desired. Establishment of an accurate and easy method for detecting intestinal metaplasia was the goal of this study. Surgical specimens of stomachs resected for gastric cancer were used. The specimens were stained with methylene blue, an oxidation‐reduction marker, in whole mount, after fixation with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4), and observed under a stereo‐microscope. Normal gastric mucosa was stained blue, whereas intestinal metaplasia mucosa was not stained and had white or sky‐blue island‐like features. Intestinal metaplasia of complete type was unstained and showed white island‐like features, while intestinal metaplasia of incomplete type showed sky‐blue staining. With this method, we were able to detect even intestinal metaplasia composed of a single gland, when the intestinal metaplasia was of complete type. When stomach samples were stained in the presence of diphenyleneiodonium chloride (DPI), an inhibitor of nicotineamide adenine dinucleotide phosphate reduced form (NADPH) reductase, all the samples were homogeneously stained blue. Loss of the color of methylene blue was caused by the reductase activity of NADPH reductase, which is strongly and specifically expressed in intestinal metaplasia. A novel method for detecting intestinal metaplasia, even a single gland, was established.
Coordinate Involvement of Cell Cycle Arrest and Apoptosis Strengthen the Effect of FTY720
Authors: Nagahara Yukitoshi , Matsuoka Yumiko , Saito Kaoru , Ikekita Masahiko , Higuchi Shigesada , Shinomiya Takahisa
Published: August 2005

A novel reagent, FTY720 (2‐amino‐2‐[2‐(4‐octylphenyl)ethyl]‐l,3‐propanediol hydrochloride), has been shown to induce a significant decrease of lymphocytes and lymphoma cells and is expected to be a potent immunosuppressant and anti‐tumor drug. The decrease in lymphocytes and lymphoma cells is mainly the result of FTY720‐induced apoptosis. FTY720 directly affects mitochondria and induces cell death. Moreover, FTY720 activates protein phosphatase (PP) 2A and affects anti‐apo‐ptotic intracellular signal transduction proteins to attenuate the anti‐apoptotic effect. In this study, we examined the relationship between FTY720‐induced apoptosis and cell cycle regulation. FTY720 induced apoptosis significantly at the GO/G1 phase and caused GO/G1 cell cycle arrest of the human lymphoma cell lines HL‐60RG and Jurkat. Simultaneously, retinoblastoma protein (pRB) was dephosphorylated, suggesting that dephosphorylation of pRB was related to FTY720‐induced GO/G1 cell cycle arrest. Because this dephosphorylation was completely blocked by a specific PP1/ 2A inhibitor, okadaic acid, it appears that FTY720‐activated PP2A is essential for FTY720‐induced cell cycle arrest. FTY720‐induced apoptosis was inhibited by Bcl‐2 overexpression in Jurkat cells, but this did not prevent FTY720‐induced cell cycle arrest, suggesting that the mechanism of FTY720‐induced cell cycle arrest is independent of the mechanism of FTY720‐induced apoptosis. These two independent pathways strengthen the effect of FTY720.
Treatable Subsets in Cancer of Unknown Primary Origin
Authors: Sumi Hajime , Itoh Kuniaki , Onozawa Yusuke , Shigeoka Yasushi , Kodama Keiji , Ishizawa Kenichi , Fujii Hirofumi , Minami Hironobu , Igarashi Tadahiko , Sasaki Yasutsuna
Published: August 2005

The purpose of this study was to investigate the treatable subsets in cancer of unknown primary origin (CUP). Fifty patients (27 males and 23 females; median age, 53 years) with CUP diagnosed between April 1992 and June 1999 were analyzed retrospectively. Of the 50 patients, 39 received chemotherapy: platinum‐based in 31, non‐platinum‐based in 4, and clinical trials of new agents in 4. Of the 39 patients, 13 (33.3%; 95% confidence interval: 19.1–50.2%) showed objective responses, with 4 complete responders. Patients with poorly differentiated carcinomas in whom p‐subunit of human chorionic gonadotropin (β‐HCG) was elevated more than 10 mlU/ml and female patients with peritoneal adenocarcinomatosis achieved high response rates (83.3% and 80%, respectively) with platinum‐based chemotherapy, as compared with only a 15.3% response rate in the remaining patients. Platinum‐based chemotherapy provided promising results in patients with poorly differentiated carcinomas and in female patients with peritoneal adenocarcinomatosis. Significantly elevated serum levels of β‐HCG in patients with poorly differentiated carcinoma might predict a better response to platinum‐based chemotherapy. However, the investigation of novel chemotherapeutic approaches is warranted for other groups of patients with CUP.
X‐Irradiation Induces Up‐regulation of ATM Gene Expression in Wild‐type Lymphoblastoid Cell Lines, but Not in Their Heterozygous or Homozygous Ataxia‐telangiectasia Counterparts
Authors: Hirai Yuko , Hayashi Tomonori , Kubo Yoshiko , Hoki Yuko , Arita Izumi , Tatsumi Kouichi , Seyama Toshio
Published: August 2005

Ataxia‐telangiectasia (AT) is an autosomal recessive disease. The relevant gene has been cloned and designated ATM. We studied the expression of both ATM mRNA and the ATM protein in unirradi‐ated and X‐irradiated EBV (Epstein‐Barr virus)‐transformed lymphoblastoid cell lines (LCLs) derived from donors who were normal (ATM+/+), AT heterozygotes (ATM+/−), or AT homozy‐gotes (ATM−/−), respectively. In ATM+/+ LCLs, the levels of ATM mRNA were found to have increased by approximately 1.5‐fold within 1 h of exposure to 10 Gy of X‐rays, while the ATM protein levels had increased by 1.5‐ to 2.0‐fold within 2 to 3 h of irradiation. The wild‐type mRNA and protein levels both returned to their basal values fairly quickly after this tune. The results obtained with the ATM+/− LCLs were quite different, however: neither the mRNA nor protein levels were found to have increased as a consequence of X‐irradiation in any ATM+/− LCL. Twelve of the mutations in the ATM−/− LCLs we used were truncating mutations, and we suspected that the corresponding truncated ATM proteins would be too labile to be detected by western blot analysis. However, five of the ATM−/− LCLs produced mutant ATM proteins that were identical in molecular weight to the wild‐type ATM protein. When cells from three of these five clones were exposed to X‐rays, transcription of the mutant ATM genes appeared to reduce somewhat, as were the levels of protein being produced. These results suggest that the normal ATM gene responds to ionizing radiation by up‐regulating its activity, whereas none of the mutant ATM genes we studied were able to respond in this way.
Contribution to Promoting Cancer Epidemiology in Japan and to the Activities of the UICC and Others in Cancer Epidemiology and Prevention
Authors: Aoki Kunio
Published: August 2005

On the occasion of being awarding the 5th Nagayo Mataro Prize for contributions to promoting cancer epidemiology and international involvement in the UICC (International Union against Cancer), the Monbusho Overseas Field Research (International Scientific Research)‐Special Cancer Study and others, it is my pleasure to give a concise description of activities in which I have played a role. My achievements in administration and management are only a small part of the whole of what has been accomplished in cancer epidemiology and prevention, but I hope that a comprehensive coverage of the projects in which it has been my good fortune to participate may provide orientation and suggest priorities for research in the coming decades. I have taken part in many meetings that stimulated the interest of young scientists and physicians in epidemiology and prevention. It can be said that efficient administration and management are indispensable for modern scientific research in order to promote information collection and exchange. I, therefore, humbly hope that my experience may be illuminating.
Preventive Effects of Isoflavones, Genistein and Daidzein, on Estradiol‐17β‐related Endometrial Carcinogenesis in Mice
Authors: Lian Zenglin , Niwa Kenji , Tagami Keiko , Hashimoto Midori , Gao Jingchun , Yokoyama Yasuhiro , Mori Hideki , Tamaya Teruhiko
Published: August 2005

The effects of isoflavones (genistein and daidzein) on endometrial carcinogenesis in mice were investigated in two experiments. In the short‐term experiment (2 weeks), single subcutaneous (s.c.) administration of genistein [1 mg/30 g body weight (b.w.)] significantly decreased the levels of estradiol‐l7β (E2) (5 ppm in diet)‐induced expression of c‐jun, interleukin‐lα (IL‐lα) and tumor necrosis factor‐a (TNF‐a) mRNAs in the uteri of ovariectomized mice (P<0.005, P<0.05 and P<0.01, respectively). Daidzein significantly inhibited E2‐induced expression of c‐fos and IL‐lα (P<0.01, P<0.01 respectively). In the long‐term experiment (30 weeks), 140 female ICR mice were given N‐methyl‐N‐nitrosourea‐containing solution (1 mg/100 g b.w.) and normal saline (as controls) into their left and right uterine corpora, respectively. They were divided into six groups; group 1 was given E2 (in diet) alone. Group 2 was given E2 and genistein (1 mg/30 g b.w., s.c., every four weeks). Group 3 was exposed to E2 and daidzein (1 mg/30 g b.w., s.c., every four weeks). Groups 4 and 5 respectively received genistein and daidzein, and were kept on the basal diet. Group 6 was kept on the basal diet and served as a control. At the termination of the experiment, incidences of endometrial adenocarcinoma and atypical endometrial hyperplasia of the group given E2 and genistein or daidzein were significantly lower than of the group with E2 alone (P<0.01 and P<0.05, respectively). It is suggested that both genistein and daidzein have an inhibitory effect on estrogen‐related endometrial carcinogenesis in mice, possibly by suppressing expression of estrogen‐induced estrogen‐related genes c‐fos and c‐jun, and internal cytokines IL‐lα and TNF‐α through a cytokine and estrogen receptor‐mediated pathway.
A Novel Experimental Mouse Model of Peritoneal Dissemination of Human Gastric Cancer Cells: Analysis of the Mechanism of Peritoneal Dissemination Using cDNA Macroarrays
Authors: Nomura Hiroki , Nishimori Hidefumi , Yasoshima Takahiro , Hata Fumitake , Sogahata Katsuya , Tanaka Hiroshi , Nakajima Futoshi , Ikeda Shinichiro , Kamiguchi Kenjiro , Isomura Hiroshi , Sato Noriyuki , Denno Ryuichi , Hirata Koichi
Published: August 2005

We established a new cell line, NUGC‐3P4T, with high peritoneal metastatic disseminating potential in nude mice. NUGC‐3P4T cells were derived from the human gastric carcinoma line NUGC‐3, which has low capacity for peritoneal dissemination. NUGC‐3P4T cells developed peritoneal dissemination in 10/10 (100%) mice, whereas the parental NUGC‐3 cells developed dissemination in 1/5 (20.0%) mice. The metastatic foci in the peritoneum showed essentially the same histological appearance as those induced by parental cells. The tumorigenicity, the motile activity and the adhesive activity to the laminin of NUGC‐3P4T cells were stronger than those of NUGC‐3 cells. Production of IL‐8 was significantly higher in NUGC‐3P4T than in NUGC‐3. cDNA macroarrays analysis showed that a variety of cytokines, interleukins, and other immunomodulators and their receptors were up‐ or down‐regulated at the mRNA level in NUGC‐3P4T cells, compared with NUGC‐3 cells. Thus, this unique cell line and in vivo model might be useful to study the biology of peritoneal dissemination of human gastric cancer.
Phenotypic Expression of Colorectal Adenocarcinomas with Reference to Tumor Development and Biological Behavior
Authors: Yao Takashi , Tsutsumi Shuichi , Akaiwa Yuji , Takata Miyuki , Nishiyama Ken‐ichi , Kabashima Akira , Tsuneyoshi Masazumi
Published: August 2005

The purpose of this study is to clarify the correlation between cell differentiation and tumor development, including tumor aggressiveness and biological behavior. Eighty‐three cases of advanced colorectal adenocarcinoma were randomly selected. Using immunohistochemical staining with antibodies to CD10, MUC2 and human gastric mucin (HGM), the colorectal adenocarcinomas could be classified into five types (18 small intestinal, 27 large intestinal, 2 gastric, 9 mixed and 27 unclassified). Each type had characteristic features. The small‐intestinal type showed a relatively lower incidence of lymphatic permeation and higher venous invasion. The large‐intestinal type showed a low incidence of venous invasion and lymph node metastasis. The mixed type revealed female and right‐side‐dominant distribution, large tumor size, high incidence of mucinous carcinoma, and low incidence of venous invasion. Gastric type was seen in only two cases (2%), which exhibited high histologic grade, lymphatic permeation and lymph node metastasis with no venous invasion. Such phenotypic classifications are considered to be useful not only for evaluation of the biological behavior of the carcinoma, but also for analysis of tumorigenesis.
Identification of Cyclophilin B‐derived Peptides Capable of Inducing Histocompatibility Leukocyte Antigen‐A2‐restricted and Tumor‐specific Cytotoxic T Lymphocytes
Authors: Tamura Mayumi , Nishizaka Shinya , Maeda Yoshiaki , Ito Masaaki , Harashima Nanae , Harada Mamoru , Shichijo Shigeki , Itoh Kyogo
Published: August 2005

We recently suggested that cyclophilin B (Cyp‐B) is a tumor antigen recognized by histocompatibility leukocyte antigen (HLA)‐A24‐restricted and tumor‐specific cytotoxic T lymphocytes (CTLs). In this study, we tried to identify Cyp‐B‐derived epitopes, which can induce HLA‐A2‐restricted and tumor‐specific CTLs in cancer patients. The tumor‐infiltrating lymphocytes (TILs) from an HLA‐A0207 patient with colon cancer were found to respond to COS‐7 cells when co‐transfected with the Cyp‐B gene and either HLA‐A0201, ‐A0206, or ‐A0207 cDNA. These TILs contained CTLs capable of recognizing either the Cyp‐B129–138 or the Cyp‐B172–179 peptide among 28 different peptides, all of which were prepared based on the HLA‐A2 binding motif. Both Cyp‐B peptides possessed the ability to induce tumor‐specific CTLs in HLA‐A2+ cancer patients. Cyp‐B172–180 (v), which is a 9‐mer peptide with valine added at the C terminus, showed no clear superiority over the parental Cyp‐B172‐179 peptide in an in vitro sensitization experiment. In vitro ‐sensitized T cells with these peptides responded to cancer cells in an HLA‐A2‐restricted manner. These two Cyp‐B peptides could be useful for specific immunotherapy of HLA‐A2+ cancer patients.
Enhancement of in vivo Antitumor Activity of a Novel Antimitotic 1‐Phenylpropenone Derivative, AM‐132, by Tumor Necrosis Factor‐cc or Interleukin‐6
Authors: Tatsumi Yasuaki , Arioka Hitoshi , Ikeda Shun‐ichi , Fukumoto Hisao , Miyamoto Ken‐ichi , Fukuoka Kazuya , Ohe Yuichiro , Saijo Nagahiro , Nishio Kazuto
Published: August 2005

TK5048 and its derivatives, AM‐132, AM‐138, and AM‐97, are recently developed antimitotic (AM) compounds. These 1‐phenylpropenone derivatives induce cell cycle arrest at the G2/M phase of the cell cycle. TK5048 inhibited tubulin polymerization in human lung cancer PC‐14 cells in a concentration‐dependent manner. In a polymerization assay using bovine brain tubulin, AM‐132 and AM‐138 were quite strong, AM‐97 was moderately strong, and TK5048 was a relatively weak inhibitor of tubulin polymerization. A murine leukemia cell line resistant to a sulfonamide antimitotic agent, E7010, which binds to colchicine‐binding sites on tubulin, was cross‐resistant to the in vitro growth‐inhibitory effect of AM compounds. Inhibition of tubulin polymerization is therefore one of the mechanisms of action of these AM compounds against tumor cells. To profile the antitumor effect of AM compounds, the in vivo antitumor effect of AM‐132 was evaluated against cytokine‐secreting Lewis lung carcinoma (LLC). Tumor‐bearing mice were treated with intravenous AM‐132 using three different treatment schedules. LLC tumors expressing tumor necrosis factor‐a (TNF‐α), granulocyte macrophage colony‐stimulating factor (GM‐CSF), or interleukin (TL)‐6 were very sensitive to AM‐132. In particular, LLC tumors expressing IL‐6 were markedly reduced by AM‐132 treatment, and showed coloring of the tumor surface and unusual hemorrhagic necrosis. These results suggest a combined effect of AM‐132 and cytokines on the blood supply to tumors.
A New Quinoline Derivative MS‐209 Reverses Multidrug Resistance and Inhibits Multiorgan Metastases by P‐glycoprotein‐expressing Human Small Cell Lung Cancer Cells
Authors: Nokihara Hiroshi , Yano Seiji , Nishioka Yasuhiko , Hanibuchi Masaki , Higasida Tsuyoshi , Tsuruo Takashi , Sone Saburo
Published: August 2005

Development of distant metastases and acquired multidrug resistance (MDR) are major problems in therapy for human small cell lung cancer (SCLC). MS‐209 is a novel quinoline compound, which reverses P‐glycoprotein (P‐gp)‐mediated MDR. We previously reported that MS‐209 reversed in vitro MDR of human SCLC (SBC‐3/ADM and H69/VP) cells expressing P‐gp. In the present study, we determined the therapeutic effect of MS‐209 in combination with chemotherapy against multiorgan metastases of MDR SCLC cells. SBC‐3/ADM cells expressing P‐gp were highly resistant to etoposide (VP‐16), adriamycin (ADM), and vincristine (VCR) in vitro, compared with parental SBC‐3 cells lacking P‐gp expression. MS‐209 restored chemosensitivity of SBC‐3/ADM cells to VP‐16, ADM, and VCR in a dose‐dependent manner in vitro. Intravenous injection with SBC‐3 or SBC‐3/ADM cells produced metastatic colonies in the liver, kidneys and lymph nodes in natural killer (NK) cell‐depleted severe combined immunodeficiency (SCID) mice, though SBC‐3/ ADM cells more rapidly produced metastases than did SBC‐3 cells. Treatment with VP‐16 and ADM reduced metastasis formation by SBC‐3 cells, whereas the same treatment did not affect metastasis by SBC‐3/ADM cells. Although MS‐209 alone had no effect on metastasis by SBC‐3 or SBC‐3/ADM cells, combined use of MS‐209 with VP‐16 or ADM resulted in marked inhibition of metastasis formation by SBC‐3/ADM cells to multiple organs. These findings suggest that MS‐209 reversed the MDR of SBC‐3/ADM cells, but not SBC‐3 cells, growing in the various organs, and inhibited metastasis formation in vivo. Therefore, this chemosensitizing agent, MS‐209, may be useful for treatment of refractory SCLC patients with multiorgan metastases.
Expression of ICAD‐L and ICAD‐S in Human Brain Tumor and Its Cleavage upon Activation of Apoptosis by Anti‐Fas Antibody
Authors: Masuoka Jun , Shiraishi Tetsuya , Ichinose Makoto , Mineta Toshihiro , Tabuchi Kazuo
Published: August 2005

ICAD/DFF is a downstream molecule of caspases, participating in nuclear DNA fragmentation during apoptosis. ICAD/DFF binds CAD/DFF40 and inhibits its DNase activity. ICAD/DFF has two alternative isoforms, long isoform (ICAD‐L/DFF45) and short isoform (ICAD‐S/DFF35). We have studied the presence and functional status of ICAD/DFF in human glioma cell lines. All cell lines tested expressed both ICAD‐L and ICAD‐S. When the cultured glioma cells were exposed to anti‐Fas antibody, these isoforms were degraded prior to the fragmentation of the nuclear DNA, indicating that the ICAD/DFF expressed in cultured glioma cells was potentially functional. In primary brain tumors and normal brain tissues, there was a difference in the expression level between ICAD‐L and ICAD‐S. In glioblastomas, ICAD‐S was more abundant than ICAD‐L. In contrast, ICAD‐L was more abundant than ICAD‐S in medulloblastomas. The present findings suggest that primary brain tumors and normal brain constitutively express ICAD/DFF, and that there is a difference between the expression levels of ICAD‐L and ICAD‐S.
A Simple Method for Classification of Cell Death by Use of Thin Layer Collagen Gel for the Detection of Apoptosis and/or Necrosis after Cancer Chemotherapy
Authors: Matsuo Atsushi , Watanabe Atsushi , Takahashi Takao , Futamura Manabu , Mori Shigeru , Sugiyama Yasuyuki , Takahashi Yuzo , Saji Shigetoyo
Published: August 2005

To assess the efficacy of cancer chemotherapy, an important index is apoptosis of the target cells, which can usually be confirmed by electron microscopy (EM). We established a new experimental technique, whereby cancer cells (MKN45) were distributed in thin collagen gel as one or two cell layers, and cultured with anti‐cancer drugs (5‐FU and CDDP). The cells were stained with fluorescent Hoechst 33258 (Ho) and photographed, then with hematoxylin and eosin (H&E) and again photographed, and processed for EM. This approach allowed us to characterize the patterns of death of single cells in detail. There were six patterns of cell damage: two patterns of apoptosis, early peripheral condensation of chromatin and late apoptotic bodies, two patterns of necrosis, cytoplasmic swelling and washed‐out images, and two further patterns, with morphological features of both apoptosis and necrosis, neither classified into necrosis nor apoptosis. The results show that cell death patterns can be mostly determined by combining observations of Ho and H&E‐stained cells without the necessity for EM observation.
Expression of 5α‐Reductases in Human Epithelial Ovarian Cancer: Its Correlation with Androgen Receptor Status
Authors: Akahira Jun‐Ichi , Suzuki Takashi , Ito Kiyoshi , Darnel Rew D. , Moriya Takuya , Sato Shinji , Yaegashi Nobuo , Okamura Kunihiro , Sasano Hironobu
Published: August 2005

Androgen metabolism and possible actions are considered to play some roles in human epithelial ovarian neoplasms, but the details have not been well studied. We have examined the expression of 5α‐reductase type 1 and type 2, which catalyze the conversion of testosterone to more active androgen, 5α‐dehydrotestosterone, and androgen receptor (AR), using immunohistochemistry (104 cases) and reverse transcription‐polymerase chain reaction (RT‐PCR) (16 cases) as a first step toward understanding the metabolism and possible actions of androgens in human common epithelial ovarian carcinoma. 5α‐Reductase type 1 was immunopositive in 75/104 cases (72.0%), and 5a‐reductase type 2 in 52/104 cases (50.0%) (P<0.001). There was no significant correlation between patterns of immunolocalization and clinicopathological parameters examined. Median labeling index (LI) for AR was 17.8% (range 0‐84.4%) which was significantly higher in serous carcinoma than other histological types (P<0.001). There was a significant positive correlation between 5α‐reductase type 1 immunoreactivity and AR LI (P=0.0027), but no significant correlation was detected in 5α‐reductase type 2. Results of RT‐PCR analysis were also consistent with those of immunohistochemistry. The relatively wide distribution of 5a‐reductase type 1, and its correlation to AR status in human epithelial ovarian malignancies suggest that this isozyme plays important roles in androgen metabolism and actions in these tumors.