bluuebunny/crossref_metadata_2025 huggingface dataset:
Release of Cytokines from Human Umbilical Vein Endothelial Cells Treated with Platinum Compounds in vitro
Authors: Shi Yucheng , Inoue Shoichi , Shinozaki Reiko , Fukue Kaoru , Kougo Tohru
Published: August 2005

Endothelial cells (EC) produce cytokines, such as interleukin (IL)‐1, IL‐6, IL‐8 and granulocyte‐macrophage colony‐stimulating factor (GM‐CSF). These cytokines have an important role in the proliferation and differentiation of hematopoietic progenitor cells. On the other hand, anticancer agents generally cause hematopoietic disorders. However, little is known about the effects of chemotherapeutic agents on the secretion of cytokines from EC. Therefore, we investigated if treatment with platinum compounds may stimulate EC to secrete cytokines. EC newly isolated from a human umbilical vein were exposed to cisplatin, carboplatin, or TRK‐710 for 80 min, then the cells were washed and placed in fresh medium. The levels of cytokines in the fresh medium were measured by the ELISA method, the levels of intracellular hydrogen peroxide (H2O2) were measured by flow cytometry, and the rhodamine 123‐stained live mitochondria of the EC were observed under a confocal laser microscope. Platinum compounds induced cytokine production in human EC: cisplatin most prominently induced the release of IL‐1 and IL‐6, and TRK‐710 had the greatest ability to induce the release of GM‐CSF. Intracellular H2O2 production and IL‐8 release were transiently induced immediately after treatment with platinum compounds, leading to IL‐1 release when H2O2 production was eliminated. These results may provide new insights into the hematological toxicity induced by anticancer agents and the role of IL‐1 and IL‐6 secreted from EC in this toxicity.
Antitumor Immunity Induction by Intracellular Hyperthermia Using Magnetite Cationic Liposomes
Authors: Yanase Mitsugu , Shinkai Masashige , Honda Hiroyuki , Wakabayashi Toshihiko , Yoshida Jun , Kobayashi Takeshi
Published: August 2005

Induction of antitumor immunity to T‐9 rat glioma by intracellular hyperthermia using functional magnetic particles was investigated. Magnetite cationic liposomes (MCLs), which have a positive surface charge, were used as heating mediators for intracellular hyperthermia. Solid T‐9 glioma tissues were formed subcutaneously on both femurs of female F344 rats, and MCLs were injected via a needle only into the left solid tumors (treatment side). The rats were then divided into two groups, which received no irradiation, or irradiation for 30 min given three times at 24‐h intervals with an alternating magnetic field (118 kHz, 384 Oe). On the treatment side, the tumor tissue disappeared completely in many rats exposed to the magnetic field. The tumor tissue on the opposite side also disappeared completely, even though MCLs were not injected into the right solid tumors. To examine whether a long‐lasting and tumor‐specific immunity could be generated, the rats that had been cured by the hyperthermia treatment were rechallenged with T‐9 cells 3 months later. After a period of transient growth, all tumors disappeared. Furthermore, immuno‐cytochemical assay revealed that the immune response induced by the hyperthermia treatment was mediated by both CD8+ and CD4+ T cells and accompanied by a marked augmentation of tumor‐selective cytotoxic T lymphocyte activity. These results suggest that our magnetic particles are potentially effective tools for hyperthermic treatment of solid tumors, because in addition to killing of the tumor cells by heat, a host immune response is induced.
Fluorescence in situ Hybridization Analysis of 12;21 Translocation in Japanese Childhood Acute Lymphoblastic Leukemia
Authors: Eguchi‐Ishimae Minenori , Eguchi Mariko , Tanaka Kimio , Hamamoto Kazuko , Ohki Misao , Ueda Kazuhiro , Kamada Nanao
Published: August 2005

Fluorescence in situ hybridization (FISH) analysis was applied to detect t(12;21) using two yeast artificial chromosome probes and cosmid probes covering the TEL(ETV6) and the AML1 gene to clarify the incidence of abnormality of t(12;21) in Japanese childhood acute lymphoblastic leukemia (ALL). We detected seven TEL/AML1 fusion positive patients (9.5%), all of whom were diagnosed as B‐lineage ALL, among 74 childhood ALL. On the other hand, no TEL/AML1 fusion positive patients were found among 37 adult ALL. The incidence among Japanese seemed to be lower than that among other nations. Of the seven patients with the TEL/AML1 fusion, five exhibited normal karyotype, one was t(8;12)(q11;p13), i(21q) and the remaining one exhibited a near‐triploid karyotype in conventional G‐banding. The FISH method clearly demonstrated that all patients with the TEL/AML1 fusion had subpopulations of leukemic cells with deletion of the normal TEL allele, which is significant for understanding the progression of leukemia with t(12;21).
Inhibitory Effects of Nabumetone, a Cyclooxygenase‐2 Inhibitor, and Esculetin, a Lipoxygenase Inhibitor, on N‐Methyl‐N‐nitrosourea‐induced Mammary Carcinogenesis in Rats
Authors: Matsunaga Kengo , Yoshimi Naoki , Yamada Yasuhiro , Shimizu Masahito , Kawabata Kunihiro , Ozawa Yuki , Hara Akira , Mori Hideki
Published: August 2005

We investigated the modifying effects of nabumetone, a relatively selective cyclooxygenase‐2 inhibitor, and esculetin, a lipoxygenase inhibitor, on N‐methyl‐N‐nitrosourea(MNU)‐induced mammary carcinogenesis in female Sprague‐Dawley rats. A total of 124 rats, 6 weeks old, were divided into 6 groups. At 50 days of age, groups 1, 2, and 3 were treated with MNU (50 mg/kg body weight) by subcutaneous injection. From the age of 8 weeks, groups 2 and 4 were given 0.03% nabumetone in the diet and groups 3 and 5 were given 0.03% esculetin in the diet. All rats were necropsied at the termination (25 weeks after the start of experiment). The incidence and multiplicity of neoplasms in group 2 were significantly smaller than those in group 1 (P<0.005 and P<0.001, respectively). The incidence of neoplasms in group 3 was also significantly smaller than that in group 1 (P<0.05). These results indicate that the intake of nabumetone or esculetin during the time corresponding to the post initiation phase has a chemopreventive effect on MNU‐induced mammary carcinogenesis in rats.
Effects of Tamoxifen on Endometrial Carcinogenesis in Mice
Authors: Niwa Kenji , Morishita Shigeo , Hashimoto Midori , Itoh Tsuneo , Fujimoto Jiro , Mori Hideki , Tamaya Teruhiko
Published: August 2005

Two experiments were conducted to determine the effect of tamoxifen (TAM) in mouse endometrium in comparison with that of 17β‐estradiol (E2). In a medium‐term assay, TAM as well as E2 treatment semi‐dose‐dependently increased the levels of fos/jun mRNA and their oncoproteins (Fos/Jun). The long‐term effect of TAM on mouse endometrial carcinogenesis was also examined in the following model. A total of 150 female ICR mice, 12–13 weeks of age, were used. Of these, 125 mice received an injection of N‐methyl‐N‐nitrosourea (MNU) solution (1 mg/100 g body weight) into their left uterine tube and saline into the right. One week later, they were divided into four groups: groups 1 (35 mice) and 2 (30 mice) were given 25 ppm and 5 ppm E2‐containing diet, respectively, while group 3 (30 mice) was fed 5 ppm TAM‐containing diet. Group 5 (30 mice) was fed basal diet alone. The remaining 25 mice (group 4) received 5 ppm TAM‐containing diet alone. At the termination of the experiment (30 weeks), endometrial carcinomas were confirmed to be present in the groups exposed to MNU. TAM increased the incidence of preneoplastic lesions of the endometrium, while E2 enhanced the occurrence of the carcinoma. No carcinomas were found in the group given TAM alone. In the ovaries, corpora lutea were lacking in most of the mice exposed to TAM, suggesting that the animals were not cycling. Such findings indicate that TAM has an enhancing effect on endometrial carcinogenesis in mice, probably via a mechanism involving overexpression of Fos/Jun proteins.
Site‐specific Localization of Epstein‐Barr Virus in Pharyngeal Carcinomas
Authors: Kojya Shizuo , Itokazu Tetsuo , Noda Yutaka , Ezaki Mitsuhiko , Tomita Yasuhiko , Ohsawa Masahiko , Aozasa Katsuyuki
Published: August 2005

In this study, the correlations of factors with Epstein‐Barr virus (EBV)‐association were investigated in 50 patients with nasopharyngeal carcinoma (NPC), 61 with oropharyngeal carcinoma (OPC), and 55 with hypopharyngeal carcinoma (HPC) in Okinawa and Osaka prefectures in Japan. The incidence of pharyngeal carcinoma in Okinawa was previously found to be higher than that in Osaka; the incidence of OPC was approximately 6 times higher and that of HPC was two times higher. The EBV genome was detected in the tumor cells of the present patients; 83% of the Okinawa and 92% of the Osaka NPC patients. The EBV genome was not detected in OPC or HPC. A univariate analysis showed that sex, the location of the tumor, histology, and the degree of lymphocytic infiltration correlated with the EBV‐positive rate. A multivariate analysis revealed that only the location of the tumor was independently correlated with the EBV‐positive rate. Histology and tumor size were factors affecting the prognosis of the patients with NPC. The NPC of poorly differentiated type frequently showed the EBV genome, and NPC with lymphocytic infiltration showed a more favorable prognosis compared to the other NPC types. These findings suggest that latent genes of EBV expressed in cancer cells might trigger a cytotoxic T cell reaction against the cancer.
Characterization of a New Human Embryonal Rhabdomyosarcoma Cell Line, RMS‐GR
Authors: Fernández Juan Emilio , Prados Jose , Melguizo Consolación , Arena Nicola , Malavasi Fabio , Álvarez Luis , Aránega Antonia
Published: August 2005

A human tumor cell line designated RMS‐GR was established from an embryonal rhabdomyosarcoma. The monolayer cells were polygonal, round or spindle‐shaped. The RMS‐GR cell line became stable with a doubling time of 42 h. Tumorigenicity of the cells was confirmed by hetero‐transplantion into nude mice. Electron microscopic images showed typical cytoplasmic inclusion of aggregated intermediate filaments and myofibril‐like thin filaments. The expression of desmin, vimentin, actin and human myoglobin was recognized by cytofluorometric analyses, and a large fraction of CK‐MM and small fractions of CK‐BB and MCK‐1 isoenzymes were found. Chromosomal analysis showed that the modal chromosome number was consistently near triploid with structural abnormalities mostly involving chromosomes 1, 3 and 8, and additional unidentified markers. No alteration of chromosome 2 was observed. The RMS‐GR cell line may provide a system to identify genes which are involved in the pathogenic mechanism of rhabdomyosarcomas, and to investigate the modulation of myogenic differentiation.
Association of Loss of Heterozygosity at the p53 Locus with Chemoresistance in Osteosarcomas
Authors: Goto Akiteru , Kanda Hiroaki , Ishikawa Yuichi , Matsumoto Seiichi , Kawaguchi Noriyoshi , Machinami Rikuo , Kato Yo , Kitagawa Tomoyuki
Published: August 2005

Although the osteosarcoma is considered to be among the most chemosensitive malignancies and preoperative chemotherapy is commonly applied, an appreciable proportion of cases are in fact quite insensitive. Predictive markers for chemosensitivity are therefore desirable in order to develop effective treatment strategies. Thirty‐two cases of conventional osteosarcomas treated at the Cancer Institute Hospital, Tokyo, were analyzed. The sensitivity to preoperative chemotherapy was investigated with reference to loss of heterozygosity (LOH) at the 17p13 (p53) and 13q14 (Rb) loci and expression of the cell‐cycle associated proteins, p53, Rb, p21/Waf‐1, mdm‐2 and Ki‐67, as detected immunohistochemically. LOH was detected by analyzing polymerase chain reaction products at marker microsatellite loci. The efficacy of chemotherapy was evaluated both radiologically and histologically. LOH at p53 or Rb loci was seen in 54% (13/24) and 58% (14/24) of cases, respectively. Only 15% of osteosarcomas with LOH at the p53 locus were sensitive to preoperative chemotherapy, as compared to 64% of tumors without such loss (P<0.05). A similar but much less distinct tendency was observed with LOH at the Rb locus. No relationship was evident between chemosensitivity and immunohistochemical staining patterns for p53, Rb, p21/Waf‐1, mdm‐2 or Ki‐67. The results suggest that p53 gene deletion, but not the other parameters investigated, may be useful for predicting chemoresistance of osteosarcomas.
Increased Proliferation of Myoblasts after Cyclic Plasma Perfusion of Tumor‐bearing Rabbits
Authors: Nakata Shin‐ichi , Ishiko Osamu , Honda Ken‐ichi , Hirai Kouzo , Deguchi Masaaki , Ogita Sachio
Published: August 2005

Rabbits implanted with VX2 carcinoma and treated by cyclic plasma perfusion showed lower weight loss and longer survival than untreated rabbits. The urinary 3‐methylhistidine/creatinine ratio was less elevated in the treated rabbits than in the untreated rabbits. Myoblasts from untreated rabbits with VX2 carcinoma showed lower proliferation than those from normal rabbits, while those from rabbits treated by cyclic plasma perfusion showed intermediate proliferation. Furthermore, the DNA synthesis rate of myoblasts from treated rabbits was much better maintained than in the case of untreated rabbits.
Two Germline Missense Mutations at Codons 804 and 806 of the RET Proto‐oncogene in the Same Allele in a Patient with Multiple Endocrine Neoplasia Type 2B without Codon 918 Mutation
Authors: Miyauchi Akira , Futami Hitoyasu , Hai Noritaka , Yokozawa Tamotsu , Kuma Kanji , Aoki Norihiko , Kosugi Shinji , Sugano Kokichi , Yamaguchi Ken
Published: August 2005

Multiple endocrine neoplasia (MEN) type 2B is a clinically distinct entity among the autosomal dominant MEN 2 syndromes. Most patients with MEN 2B carry a germline mutation (M918T) of the RET proto‐oncogene, while a few carry A883F. We examined a patient with MEN 2B, but without M918T or A883F, and her relatives. Here, we report the presence in this patient of 2 germline mutations, V804M and Y806C in the same allele. While the novel Y806C was inherited from her father, its carriers (her father and brother) was not affected by MEN 2. In contrast, V804M was a de novo mutation, that has been reported in patients with familial medullary thyroid carcinoma. Combinations of mutations of the RET proto‐oncogene may cause oncogenic activities different from those of single mutations.
Human T‐cell Leukemia Virus Type 1 Can Infect a Wide Variety of Cells in Mice
Authors: Feng Renqing , Tanaka Masakazu , Abe Hideaki , Arashi Noriko , Sun Binlian , Uchida Kazuhiko , Miwa Masanao
Published: August 2005

Analysis of human T‐cell leukemia virus type 1 (HTLV‐1)‐infected cell types and the interplay of these infected cells in vivo should provide valuable information to elucidate the pathogenesis of HTLV‐1‐associated diseases in humans and in animal models. In this study, HTLV‐1‐infected cell types were identified in HTLV‐1‐infected C3H/HeJ mice. Pan T, CD+4, CD8+8, granulocyte and pan B cell fractions in the splenocytes of MT‐2 cell‐inoculated mice were sorted by use of their cell surface high‐density expression of CD3e, CD4, CD8, Gr‐1 and B220 antigens, respectively, with a fluorescence‐activated cell sorter. the pX sequence of HTLV‐1 provirus in the lysate of each fraction was amplified by polymerase chain reaction and detected by Southern hybridization. Interestingly, in addition to the CD+4 cell fraction, the pX sequence was also found in CD8+ cell, B cell and granulocyte fractions. The broad cell spectrum of HTLV‐1 infection in mice is consistent with the situation in humans. Our findings indicate that HTLV‐1 receptor or coreceptor is widely distributed among different cell types in mice.
Mutations of the β‐Catenin Gene in Endometrial Carcinomas
Authors: Kobayashi Kanji , Sagae Satoru , Nishioka Yoshihiro , Tokino Takashi , Kudo Ryuichi
Published: August 2005

To investigate the contribution of β‐catenin to the development of endometrial carcinoma, we searched for genetic alterations of the β‐catenin gene in primary endometrial carcinomas. Mutational analysis of exon 3 of the β‐catenin gene, encoding the serine/threonine residues for GSK‐3β phosphorylation, was performed for 35 tumors. Nucleotide sequencing analysis revealed that 5 tumors (5/35, 14%) contained mutations (S33C, S37C, S37F, T41A) that altered potential GSK‐3β phosphorylation sites. Each of the mutations resulted in the substitution of serine/threonine residues that have been implicated in the down‐regulation of β‐catenin through phosphorylation by GSK‐3β kinase. Furthermore, the incidence of β‐catenin mutations was significantly higher in early‐onset (3 of 5) than that in late‐onset tumors (2 of 30) (P=0.014, Fisher's exact test). Replication error (RER)‐positive phenotype was not detected in tumors with the β‐catenin gene mutation, although 10 of 35 tumors revealed RER. We performed immunohistochemistry of β‐catenin in 17 cases for which tissue samples were available. We confirmed accumulation of β‐catenin protein in both the nucleus and cytoplasm in 3 tumors, including two in which amino acid alterations had occurred at codon 33 and 37. The other case had no mutation in exon 3. Our results suggested that mutations at serine/threonine residues involved in phosphorylation by GSK‐3β affected the stability of β‐catenin. Accumulation of mutant β‐catenin could contribute to the development of a subset of endometrial carcinomas, particularly those of the early‐onset type.
Growth Inhibition of A549 Human Lung Adenocarcinoma Cells by L‐Canavanine Is Associated with p21/WAF1 Induction
Authors: Ding Yi , Matsukawa Yoshizumi , Ohtani‐Fujita Naoko , Kato Daishiro , Dao Su , Fujii Takaaki , Naito Yuji , Yoshikawa Toshikazu , Sakai Toshiyuki , Rosenthal Gerald A.
Published: August 2005

L‐Canavanine (CAV) is a higher plant nonprotein amino acid and a potent L‐arginine antimetabolite. CAV can inhibit the proliferation of tumor cells in vitro and in vivo, but little is known regarding the molecular mechanisms mediating these effects. We demonstrated that the treatment of human lung adenocarcinoma A549 cells with CAV caused growth inhibition; G1 phase arrest is accompanied by accumulation of an incompletely phosphorylated form of the retinoblastoma protein, whose phosphorylation is necessary for cell cycle progression from G1 to S phase. In addition, CAV induces the expression of p53 and subsequent expression of a cyclin‐dependent kinase inhibitor, p21/WAF1. The p53–dependent induction of p21/WAF1 and the following dephosphorylation of the retinoblastoma protein by CAV could account for the observed CAV‐mediated G1 phase arrest.
Wide Spectrum of Antitumor Activity of a Neutralizing Monoclonal Antibody to Human Vascular Endothelial Growth Factor
Authors: Asano Makoto , Yukita Ayako , Suzuki Hideo
Published: August 2005

Vascular endothelial growth factor (VEGF) is known as an angiogenic factor for tumor angiogene—sis. We developed a neutralizing anti—VEGF monoclonal antibody (MAb), MV833, and examined its antitumor activity against 27 human tumor cell lines transplanted in nude mice. All the tumor cell lines used in this study secreted various amounts of VEGF into culture medium in vitro. However, the growth of the cell lines, including three which expressed VEGF receptor, was not affected by exogenously added MV833 in vitro. All tumor cell lines including colon, lung, breast, pancreas, and melanoma, grew subcutaneously in nude mice. The growth of HeLa/v5, which had been transformed by human VEGF121 gene and secreted large amounts of VEGF, was significantly faster than that of the control vector transformant. Although the amounts of VEGF secreted from two HeLa transformants differed greatly, MV833 completely inhibited the growth of both tumors. Moreover, the growth of the other 25 human tumor cell lines transplanted into nude mice was also strongly suppressed by MV833. Neither the amount of VEGF secreted from each tumor cell line in vitro nor the expression of VEGF receptor correlated with the antitumor activity of MV833. MV833, administered when tumor volumes reached 400 mm3, completely inhibited the growth of some tumor lines. The results show VEGF to be a critical angiogenic factor for many tumors. VEGF—neutralizing antibody could be applicable as an antitumor agent for a wide range of tumors.
Preventive Effect of Matrix Metalloproteinase Inhibitor, R‐94138, in Combination with Mitomycin C or Cisplatin on Peritoneal Dissemination of Human Gastric Cancer Cell Line TMK‐1 in Nude Mice
Authors: Igarashi Naoki , Kubota Tetsuro , Otani Yoshihide , Matsuzaki Shinjiro Wilson , Watanabe Masahiko , Teramoto Tatsuo , Kumai Koichiro , Tamaki Kazuhiko , Tanzawa Kazuhiko , Kobayashi Tomowo , Kitajima Masaki
Published: August 2005

R‐94138, a matrix metalloproteinase inhibitor, was examined for the ability to prevent peritoneal dissemination of a human gastric cancer xenograft, TMK‐1. When the supernatant of a co‐culture of TMK‐1 cells and human normal fibroblast cells was subjected to gelatin zymography, it was clear that the protein expression of MMP‐2 had been inhibited by R‐94138. When TMK‐1 was injected intraperitoneally (i.p.) into nude mice at 5×105 cells/body, the resulting peritoneal dissemination mimicked clinical carcinomatous peritonitis. When the maximum tolerated dose of mitomycin C (MMC) or cisplatin (DDP) was given 12 h after the tumor inoculation, peritoneal dissemination was completely inhibited, while the effect of R‐94138 was limited when it was given i.p. at a dose of 20 mg/kg in a schedule of q.d. ×5 starting 12 h after tumor injection. MMC and DDP also suppressed peritoneal dissemination when they were administered 1 week after the tumor inoculation at a single dose of 2 and 3 mg/kg i.p., respectively. R‐94138 inhibited peritoneal dissemination when it was administered i.p. at a dose of 30 mg/kg in a schedule of q.d. ×5 starting from 1 week after tumor injection. The combination of MMC and R‐94138 increased the preventive effect on peritoneal dissemination. R‐94138 seems to be a promising candidate to prevent peritoneal dissemination of gastric cancer.
Chemoprevention by the Oxygenated Carotenoid β‐Cryptoxanthin of N‐Methylnitrosourea‐induced Colon Carcinogenesis in F344 Rats
Authors: Narisawa Tomio , Fukaura Yoko , Oshima Shunji , Inakuma Takahiro , Yano Masamichi , Nishino Hoyoku
Published: August 2005

β‐Cryptoxanthin (βCx), one of 4 major carotenoids in the blood, was investigated for anticarcinogenic activity in F344 rats. Four groups of 25 rats each received an intrarectal dose of 2 mg of N‐methylnitrosourea 3 times a week for 5 weeks, and were fed the diet supplemented with 0 ppm (control), 25 ppm, 5 ppm or 1 ppm βCx throughout the experiment. The colon cancer incidence at week 30 was significantly lower in the βCx (25 ppm) diet group, but not in the βCx (5 ppm) and βCx (1 ppm) diet groups, than in the control diet group: 68%, 84%, 80% vs. 96%. The results suggested that dietary βCx may affect colon carcinogenesis after accumulation in the colonic mucosa, perhaps due to absorption from the colon as well as the small intestine.
Epidermal Growth Factor‐dependent Dissociation of CrkII Proto‐oncogene Product from the Epidermal Growth Factor Receptor in Human Glioma Cells
Authors: Katayama Haruko , Hashimoto Yuko , Kiyokawa Etsuko , Nakaya Mie , Sakamoto Atsuhiko , Machinami Rikuo , Kurata Takeshi , Mochizuki Naoki , Matsuda Michiyuki
Published: August 2005

Human glioma cells frequently overexpress epidermal growth factor receptor (EGFR). We found that the CrkII proto‐oncogene product was associated with the EGFR in human glioma cells in the absence of epidermal growth factor (EGF). EGF stimulation of glioma cells induced the phosphorylation of tyrosine 221 of the CrkII protein, which correlates with its dissociation from the EGFR. By contrast, Shc and Grb2 were inducibly associated with the EGFR in response to EGF stimulation of glioma cells. In A431 cells, epidermoid carcinoma cells which overexpress EGFR, CrkII was tyrosine‐phosphorylated and associated with the EGFR in an EGF‐dependent manner. Therefore, the dissociation of CrkII from the EGFR upon stimulation with EGF appears to be specific to glioma cells. The Cbl oncogene product was also tyrosine‐phosphorylated in U87MG glioma cells upon EGF stimulation. However, unlike in other cell lines, CrkII was not inducibly bound to Cbl in U87MG glioma cells. Thus, EGF‐dependent binding of CrkII to phosphotyrosine‐containing proteins appears to be suppressed in glioma cells. To evaluate the physiological role of dissociation of CrkII from EGFR, we expressed the CrkII‐23 mutant in glioma cells. CrkII‐23 mutant, which was isolated as a suppressor gene of the EGF‐dependent transformation of NRK cells, binds constitutively to EGFR. We found that expression of CrkII‐23 inhibited the anchorage‐independent growth of the glioma cells in the presence of EGF. Taken together, these data implicate EGF‐dependent dissociation of CrkII from EGFR in the oncogenicity of human glioma cells.
Antitumor Efficacy of Hypothemycin, A New Ras‐signaling Inhibitor
Authors: Tanaka Hidekazu , Nishida Kazuyo , Sugita Kenji , Yoshioka Takayuki
Published: August 2005

We have devised a new drug screening assay to discover anti‐cancer drugs which inhibit Ras‐mediated cellular signals, by utilizing a Ras‐responsive element (RRE)‐driven reporter gene system. We found that hypothemycin, an anti‐bacterial, reduces RRE‐dependent transcription. Treatment of tumor cells with hypothemycin resulted in reduced expression of Ras‐inducible genes, including MMP (matrix metalloproteinase)‐1, MMP‐9, transforming growth factor‐β (TGF‐β), and vascular endothelial growth factor (VEGF), but not that of the constitutively expressed gene, MMP‐2. The results of zymography demonstrated that hypothemycin reduced the production of MMP‐9 and MMP‐3, another Ras‐inducible MMP, in the culture medium. Hypothemycin selectively inhibits anchorage‐independent growth of Ras‐transformed cells in comparison with anchorage‐dependent growth. These findings suggest that hypothemycin inhibits Ras‐mediated cellular signaling. Daily treatment of tumor‐bearing mice with hypothemycin resulted in significant inhibition of tumor growth. Since MMP‐1, MMP‐3 and MMP‐9 play important roles in tumor invasion and TGF‐? and VEGF are involved in tumor angiogenesis, hypothemycin is considered to be an example of a new class of antitumor drugs, whose antitumor efficacy can be at least partly attributed to inhibition of Ras‐inducible genes.
In vivo Cisplatin Resistance Depending upon Canalicular Multispecific Organic Anion Transporter (cMOAT)
Authors: Minamino Tomoyoshi , Tamai Mitsuo , Itoh Yoshie , Tatsumi Yasuaki , Nomura Masaaki , Yokogawa Koichi , Suzuki Hiroshi , Sugiyama Yuichi , Ohshima Tohru , Miyamoto Ken‐ichi
Published: August 2005

The in vitro sensitivities to cisplatin of AH66 and AH66F cells, a variant obtained from AH66 cells, were very similar, when assayed in a medium containing 5% fetal bovine serum (FBS), whereas in the in vivo experiments AH66F cells were sensitive and AH66 cells were highly resistant to cisplatin. In this study, we examined the mechanism of the in vivo cisplatin resistance of AH66 cells. The in vitro cisplatin sensitivity of AH66 cells was lowered by changing FBS to 5% ascites fluid (ASF) in the assay medium and the sensitivity in FBS by treatment with buthioninesulfoximine (BSO). The sensitivity of AH66F cells was not changed by these treatments. Moreover, after culture in 5% ASF for 48 h, the accumulation of cisplatin in AH66 cells was decreased and the efflux of cisplatin from the cells was accelerated. The accumulation of cisplatin in AH66 cells in ASF was increased by pretreatment with BSO, sodium azide or probenecid. Then, we examined the expression of the glutathione (GSH) conjugate efflux pump family. Among them, only the expression of canalicular multispecific organic anion transporter (cMOAT) in AH66 cells was decreased by culture in FBS and enhanced by ASF. These results suggest that some substances contained in ASF enhanced the expression of cMOAT in the plasma membrane of AH66 cells and this transporter actively extruded cisplatin‐GSH conjugate from the cells. Consequently, AH66 cells afford a cisplatin‐resistant tumor in the host.
Establishment of an in vivo Highly Metastatic Rat Hepatocellular Carcinoma Model
Authors: Futakuchi Mitsuru , Hirose Masao , Ogiso Tadashi , Kato Koji , Sano Masashi , Ogawa Kumiko , Shirai Tomoyuki
Published: August 2005

We previously found by chance that N‐nitrosomorpholine (NMOR) given after a multi‐carcinogenic treatment induces liver carcinomas with 56% lung metastasis, and it was confirmed that hepatocellular carcinoma (HCC) with 100% lung metastasis was produced by 24‐week treatment with NMOR and additional treatment with diethylnitrosamine (DEN). In the present study, we modified the duration of NMOR to establish an animal model with a simple experimental protocol and an appropriate experimental duration which would facilitate further study of the mechanisms of metastasis and antimetastatic agents. The results revealed DEN exposure followed by a 16‐week treatment with NMOR to be a most efficient method for the induction of HCC metastasizing to the lung. Loss of cadherin, demonstrated immunohistochemically, occurred in an early stage of carcinogenesis, and this was reflected in malignant conversion of primary lesions. This model, with its essential similarities to malignant tumor behavior in man, should find application not only for elucidation of the mechanisms underlying metastasis, but also in the development of anti‐metastatic agents.
Difference in Target Organs in Carcinogenesis with a Heterocyclic Amine, 2‐Amino‐3,4‐dimethylimidazo[4,5‐f]quinoline, in Different Strains of Mice
Authors: Fujita Hirofumi , Nagano Kasuke , Ochiai Masako , Ushijima Toshikazu , Sugimura Takashi , Nagao Minako , Matsushima Taijiro
Published: August 2005

2‐Amino‐3,4‐dimethylimidazo[4,5‐f]quinoline (MeIQ) induces cancers in the forestomach and liver, but not in the colon, of CDF1 male and female mice, which are thought to be resistant to induction of colon cancer by 1,2‐dimethylhydrazine. In this study, we examined the carcinogenicity of MeIQ in C57BL/6N female mice, which are susceptible to 1,2‐dimethylhydrazine. This strain of mice developed carcinomas of the cecum, colon and liver, but not the forestomach, when given a diet containing 300 ppm of MeIQ. This fact indicates that the target organs of a chemical carcinogen change depending on the strain of a given animal species.
Gastric Tumor Induction by 1,2‐Dimethylhydrazine in Wistar Rats with Intestinal Metaplasia Caused by X‐Irradiation
Authors: Watanabe Hiromitsu , Uesaka Toshihiro , Kido Shoichirou , Ishimura Yoshimasa , Shiraki Kazuhisa , Kuramoto Ken , Hirata Shitau , Shoji Shuneki , Katoh Osamu , Fujimoto Nariaki
Published: August 2005

Five‐week‐old male Wistar rats were X‐irradiated with a total of 20 Gy in 2 equal fractions at a 3‐day interval. 1,2‐Dimethylhydrazine (DMH) solution was injected i.m. into the back musculature at a dose of 20 mg/kg body weight weekly for 10 weeks, beginning 20 weeks after the final irradiation. Twelve months after the initial carcinogen treatment, tumors in the fundus of the glandular stomach were observed in 5 of 23 animals receiving both X‐irradiation and DMH treatment. No tumors of the glandular stomach were observed in the DMH and X‐ray alone or nontreatment groups. It is concluded that the presence of intestinal metaplasia may increase sensitivity to the induction of gastric tumors by carcinogens like DMH.
Differential Expression of Facilitative Glucose Transporter (GLUT) Genes in Primary Lung Cancers and Their Liver Metastases
Authors: Kurata Takayasu , Oguri Tetsuya , Isobe Takeshi , Ishioka Shin‐ichi , Yamakido Michio
Published: August 2005

Glucose uptake is mediated by members of the facilitative glucose transporter (GLUT) family. Malignant cells take up more glucose than their normal counterparts. The aim of this study was to investigate the gene expression levels of the GLUT family, especially GLUT1, GLUT3, and GLUT5 in primary lung cancer, metastatic liver tumors, and normal lung tissues, and to compare the expression levels of primary and metastatic tumors with those of normal tissues. We analyzed 105 autopsy samples (35 primary lung tumors, 35 corresponding normal lung tissues, 25 normal liver tissues, and 10 metastatic liver tumors) from 35 patients using the quantitative reverse transcription polymerase chain reaction. The GLUT1 gene expression levels in primary lung tumors were significantly higher than those in normal lung tissues. In liver metastatic lesions, the GLUT3 and GLUT5 gene expression levels were significantly higher than those in primary lung tumors, but there were no differences in GLUT1 expression levels between primary and metastatic liver tumors. Our results show that the gene expression pattern of the GLUT family is different between primary and metastatic liver tumors and suggest that the energy transporters in metastatic tumors may be different from those in primary tumors.
Expression of Thymidylate Synthase in Human Non‐small Cell Lung Cancer
Authors: Otake Yosuke , Tanaka Fumihiro , Yanagihara Kazuhiro , Hitomi Shigeki , Okabe Hiroyuki , Fukushima Masakazu , Wada Hiromi
Published: August 2005

5‐Fluorouracil (5‐FU) has been used worldwide, and the correlation between its effects and thymidylate synthase (TS) expression has been reported in gastrointestinal malignancy. But the significance of TS expression for 5‐FU‐based chemotherapy has rarely been reported in non‐small cell lung cancer (NSCLC). We investigated surgically resected specimens of 23 consecutive patients with previously untreated NSCLC. We used immunohistochemistry and western blot analysis with anti‐TS polyclonal antibody to evaluate the existence of TS, and fluorodeoxyuridine‐5′‐monophosphate (FdUMP) binding assay to evaluate the enzymic activity of TS. We found that 14 samples (60.9%) were positive immunohistochemically, and that the results of immunohistochemistry closely reflected the enzymic activity measured by FdUMP binding assay (ranging from 1.8 to 56.9 pmol/g protein). These results seem to support our experience that 5‐FU and its derivatives are clinically significantly effective as a postoperative adjuvant chemotherapy against NSCLC.
Protein Expression of Cell Cycle Regulator, p27Kip1, Correlates with Histopathological Grade of Non‐Hodgkin's Lymphoma
Authors: Kudoh Shiori , Kumaravel Tirukalikundram S. , Kuramavel Bharathy , Eguchi Minenori , Asaoku Hideki , Dohy Hiroo , Fujiwara Megumi , Sasaki Naomi , Tanaka Kimio , Kamada Nanao
Published: August 2005

The protein p27Kp1 is one of the cyclin‐dependent kinase inhibitors that are known to play important roles in the regulation of cell‐cycle progression. Low levels of p27 expression in malignant cells are associated with poor prognosis in patients with breast, lung, colorectal and gastric cancers. To determine the relation of cyclin‐dependent kinase inhibitors to histopathological grades of B‐cell non‐Hodgkin's lymphomas, the expression of p27, cyclin D1 and cyclin E in lymph node tissues was investigated in 56 patients with B‐cell non‐Hodgkin's lymphomas by western blotting and immunohistochemical techniques. High levels of p27 expression were observed in most lymph node tissue samples (93%) obtained from patients with low grade B‐cell non‐Hodgkin's lymphomas, while expression was low in lymph node tissue taken from all patients with intermediate and high grade B‐cell non‐Hodgkin's lymphomas. The difference in p27 expression in lymphoma tissues was significant among the different histopathological grades of B‐cell non‐Hodgkin's lymphomas (P<0.01). The analysis of the survival time of patients showed that the reduction of p27 expression correlated with poor prognosis. Cyclin D1, showed a high level of expression in mantle cell lymphomas and high grade B‐cell non‐Hodgkin's lymphomas. Cyclin E showed limited expression in 18 of 31 lymphoma tissues. Both cyclin D1 and E protein expression were not significantly different among the grades of B‐cell non‐Hodgkin's lymphomas. These results demonstrate that the level of p27 expression in lymphoma tissue is an important parameter in the classification of B‐cell non‐Hodgkin's lymphomas and in the prediction of prognosis.
C‐kit Gene Abnormalities in Gastrointestinal Stromal Tumors (Tumors of Interstitial Cells of Cajal)
Authors: Sakurai Shinji , Fukasawa Tomoki , Chong Ja‐Mun , Tanaka Akira , Fukayama Masashi
Published: August 2005

Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the GI tract, and expresses KIT and CD34 in most cases. Gain‐of‐function mutation of the c‐kit proto‐oncogene has been described, but its significance in GIST has not yet been fully evaluated. Mutation in exon 11 of the c‐kit gene was determined by both polymerase chain reaction‐single strand conformation polymorphism (PCR‐SSCP) analysis and direct sequencing in primary and metastatic GISTs and esophageal leiomyomas in Japanese subjects. C‐kit gene mutation was identified in 15 of 48 primary GISTs (31%), four of seven metastatic GISTs, but none of the leiomyomas. Three mutations were mis‐sense point mutations, and 16 were in‐frame deletions of 3–48 bp. C‐kit gene mutation was observed equally in low‐ and high‐risk groups, and was not related to any clinical and pathologic factors, phenotypes or Ki‐67 labeling index (LI) of tumor cells. In five of 15 deletion mutations (four in primary tumors and one in a metastatic tumor), the mutations were present at the distal location of exon 11 of the c‐kit gene, which was a minor mutation in previous reports from Finland and the USA. C‐kit gene mutations in GIST are not always related to a poor prognosis, but further comparative studies are necessary in Western and Japanese populations.
Cyclooxygenase‐2 (COX‐2) mRNA Expression Levels in Normal Lung Tissues and Non‐small Cell Lung Cancers
Authors: Ochiai Mari , Oguri Tetsuya , Isobe Takeshi , Ishioka Shinichi , Yamakido Michio
Published: August 2005

One of the cyclooxygenase (COX) isoforms, COX‐2, is over expressed in various human cancers. In this study, we examined the gene expression levels of COX‐2 in primary non‐small cell lung cancers (NSCLC), metastatic lymph nodes, and normal lung tissues. The expression levels of the COX‐2 gene were assessed by means of the reverse transcription polymerase chain reaction in 76 autopsy samples (29 primary NSCLC, 29 corresponding normal lung tissues, and 9 metastatic lymph nodes). The expression levels in NSCLC (both adenocarcinomas and squamous cell carcinomas) were significantly higher than in normal lung tissues and were significantly higher in adenocarcinomas than in squamous cell carcinomas. Differences between the levels of expression of COX‐2 in primary tumors and their corresponding metastatic lymph nodes in 9 adenocarcinoma patients were not significant. Our results indicate that COX‐2 may be associated with carcinogenesis of NSCLC, and that it may be a target for the treatment of NSCLC.
Expression of Platelet‐derived Endothelial Cell Growth Factor/Thymidine Phosphorylase in Human Bladder Cancer
Authors: Tanaka Tsutomu , Yoshiki Tatsuhiro , Arai Yoichi , Higuchi Kayoko , Kageyama Susumu , Ogawa Yasutaka , Isono Takahiro , Okada Yusaku
Published: August 2005

We investigated the expression of platelet‐derived endothelial cell growth factor/thymidine phosphorylase (PD‐ECGF/TP) in primary bladder cancer, its association with clinicopathologic findings, and their prognostic value. mRNA was extracted from 20 bladder cancer specimens and 6 normal bladder mucosal tissues. Relative amounts of PD‐ECGF/TP mRNA were evaluated by reverse transcriptase‐polymerase chain reaction (RT‐PCR) and compared with the level of glycer‐aldehyde‐3‐phosphate dehydrogenase mRNA (used as an internal standard). PD‐ECGF/TP expression was examined by immunohistochemistry in 85 patients who underwent cystectomy for bladder cancer. Serum PD‐ECGF/TP levels were measured in 23 patients using a sandwich‐type enzyme‐linked immunosorbent assay. By RT‐PCR analysis, expression of PD‐ECGF/TP was found to be 7‐fold higher in invasive tumors than in superficial tumors (P<0.01) and 9‐fold higher than in normal bladder (P<0.01). Out of 85 transitional cell carcinoma tissue samples, 69 (81%) were evaluated as PD‐ECGF/TP‐positive by immunohistochemical staining. PD‐ECGF/TP expression correlated significantly with tumor grade (P=0.001), depth of invasion (P=0.012), and lymphatic invasion (P=0.01). No correlation was found between expression of PD‐ECGF/TP and the number of tumors, tumor configuration, lymph node involvement, venous invasion, c‐erbB‐2 expression, or overall survival. We could not detect a significant serum level of PD‐ECGF/TP in any patient. The results suggest that PD‐ECGF/TP might give valuable information for bladder cancer management, though it may not be a good new tumor marker for bladder cancer.
Cisplatin‐resistant HeLa Cells Are Resistant to Apoptosis via p53‐dependent and ‐independent Pathways
Authors: Minagawa Yukihisa , Kigawa Junzo , Itamochi Hiroaki , Kanamori Yasunobu , Shimada Muneaki , Takahashi Masakuni , Terakawa Naoki
Published: August 2005

Since HeLa cells possess very little functional p53 activity, they could be originally resistant to genotoxic stress‐induced apoptosis. Therefore, it is likely that the drug‐resistant cells derived from HeLa cells are more resistant to apoptosis. The aim of this study was to determine whether cisplatin‐resistant cells derived from HeLa cells have an apoptosis‐resistant phenotype. A cisplatin‐resistant cell subline, HeLa/CDDP cells, showed a 19‐fold resistance to cisplatin compared with the parent cells. The subline showed a collateral sensitivity to paclitaxel. An equitoxic dose (IC50) of cisplatin produced DNA fragmentation in HeLa cells but not in HeLa/CDDP cells. Transfection of wild‐type p53gene enhanced the cytotoxicity of cisplatin and cisplatin‐induced apoptosis in HeLa cells but not in HeLa/CDDP cells, although it caused p53 overexpression in both cell lines. The expression of caspase 1 (interleukin‐1β‐converting enzyme, ICE) mRNA and the overexpression of bax protein were observed only in HeLa cells. Paclitaxel‐induced DNA fragmentation appeared less in HeLa/CDDP cells than in HeLa cells. p53gene transfection did not affect the extent of DNA fragmentation in either cell line, suggesting that paclitaxel may induce p53‐independent apoptosis. These findings suggest that HeLa/CDDP cells may have an acquired phenotype that is resistant to p53‐dependent and ‐independent apoptosis.
Association between Smoking and Tumor Progression in Japanese Women with Adenocarcinoma of the Lung
Authors: Sekine Ikuo , Nagai Kanji , Tsugane Shoichiro , Yokose Tomoyuki , Kodama Tetsuro , Nishiwaki Yutaka , Suzuki Kenji , Kuriyama Takayuki
Published: August 2005

We studied the effect of smoking on tumor progression in 3312 patients with lung cancer registered at the National Matsudo Hospital and National Cancer Center Hospital East between 1977 and 1996. The odds ratios of the following variables for tumor extent (localized versus advanced disease) and hazard ratios for survival were calculated in both sexes separately using the logistic regression and Cox proportional hazard models, respectively: smoking history, number of cigarettes smoked per day, pack‐years smoked, age, histological type, and the year of admission. Of the 943 women, 367 (38.9%) were smokers and 694 (73.6%) had adenocarcinoma, whereas of the 2369 men, 2255 (95.2%) were smokers and 1010 (42.6%) had adenocarcinoma. In female adenocarcinoma patients, the odds ratio (95% confidence interval) for advanced disease and the hazard ratio (95% confidence interval) for survival with an increase of 30 cigarettes smoked per day were 2.86 (1.49‐5.49) and 1.52 (1.13‐2.04), respectively, but in those with non‐adenocarcinoma, the odds ratio and hazard ratio were 0.96 (0.41‐2.23) and 1.13(0.75‐1.70), respectively. In male patients, smoking history influenced tumor progression regardless of histological type, but the odds ratios and hazard ratios were lower than those for women with adenocarcinoma. In conclusion, smoking habit was closelycorrelated with progression of adenocarcinoma in women. This association was not observed in women with non‐adenocarcinoma and was weaker in men, suggesting various effects of smoking on lung cancer development depending on gender and the histological typeof the tumor
Tea Polyphenol Intake and Changes in Serum Pepsinogen Levels
Authors: Hamajima Nobuyuki , Tajima Kazuo , Tominaga Suketami , Matsuura Akira , Kuwabara Masayoshi , Okuma Katashi
Published: August 2005

Following a phase I study, a phase II study was conducted to evaluate the effects of two different doses of tea polyphenols on serum pepsinogen levels. Subjects were patients aged 40 to 69 years who had undergone gastroscopy between 1995 and 1997 at Aichi Cancer Center Hospital, and had been found to have no disease requiring medication. Those with pepsinogen I <70 ng/ml and pepsinogen I/II ratio ?6 were included in this study. Capsules containing 100 mg of tea polyphenols were administered for 1 year: 1 capsule per day for 101 patients (42 males and 59 females), and 6 capsules (equivalent to 10 cups) per day for 83 patients (30 males and 53 females). The enrollment of the 1 capsule group preceded that of the 6 capsule group, in which re‐participation was allowed. Blood samples were obtained 1 year after participation from 86 participants of the 1 capsule group and 77 participants (43 new participants and 34 re‐participants) of the 6 capsule group. The compliance in polyphenol capsule intake ranged from 11.4 to 105.7% (87.6% on average) of the scheduled amount for the 1 capsule group and 3.2 to 112.3% (77.8% on average) for the 6 capsule group. No serious polyphenol‐related adverse effects were reported. The difference in pepsinogen I between before and after 1 year intake of the polyphenol was 3.1 ng/ml for the 43 participants of the 6 capsule group, but 3.5 ng/ml for the 1 capsule group. The mean pepsinogen I/II ratio for the 43 participants increased from 2.37 by 0.08. This increase was not larger than that for the 1 capsule group (from 2.61 by 0.11). Among 34 participants in both interventions, no significant increase in pepsinogen I and I/II ratio for the 6 capsule intervention was observed. This result suggests that additional polyphenol intake for 1 year in Japanese does not improve pepsinogen levels, which are considered to reflect stomach atrophy, a high‐risk condition for stomach cancer.
Detection of Apoptotic Cells in Human Colorectal Cancer by Two Different in situ Methods: Antibody against Single‐stranded DNA and Terminal Deoxynucleotidyl Transferase‐mediated dUTP‐biotin Nick End‐labeling (TUNEL) Methods
Authors: Watanabe Ichizo , Toyoda Masao , Okuda Junji , Tenjo Toshiyuki , Tanaka Keitaro , Yamamoto Tetsuhisa , Kawasaki Hiroshi , Sugiyama Toshihiro , Kawarada Yoshihiko , Tanigawa Nobuhiko
Published: August 2005

Wecomparatively investigated the extent of apoptotic cell loss in human colorectal cancers evaluated by two methods, namely the conventional terminal deoxynucleotidyl transferase (TdT)‐ mediated deoxyuridine triphosphate (dUTP)‐biotin nick end‐labeling (TUNEL) method and immunohistochemistry for single‐stranded (ss) DNA. The apoptotic index (AI) obtained with the TUNEL method was higher than that shown by the immunohistochemistry for ssDNA. However, a significant correlation in AIs evaluated by these methods was found. The AIs obtained by both methods were significantly higher in the advanced cancers than in the early cancers. Cellular proliferation activity was assessed in terms of positivity rate (PR) for expression of proliferating cell nuclear antigen (PCNA). The PR of advanced cancers was significantly higher than that of early cancers. The present results indicate that immunohistochemistry for ssDNA is useful (as is the TUNEL method) for evaluation of apoptotic tumor cells in colorectal carcinomas. In addition, it was confirmed that there is a remarkable increase of not only proliferation activity, but also tumor cell apoptosis in the process of progression of colon cancer from early to advanced stages of the disease.