bluuebunny/crossref_metadata_2025 huggingface dataset:
Dose‐dependent Induction of Mammary Carcinomas in Female Sprague‐Dawley Rats with 2‐Amino‐l‐methyl‐6‐phenylimidazo[4,5‐b]pyridine
Authors: Imaida Katsumi , Hagiwara Akihiro , Yada Hideaki , Masui Tsuneo , Hasegawa Ryohei , Hirose Masao , Sugimura Takashi , Ito Nobuyuki , Shirai Tomoyuki
Published: August 2005

The dose‐dependence of 2‐amino‐1‐methyl‐6‐phenylimidazo[4,5‐b]pyridine (PhIP) induction of mammary carcinomas was investigated in female Sprague‐Dawley (SD) rats given PhIP in the diet for 48 weeks at concentrations of 0, 25, 100 and 200 ppm in experiment 1, and 0, 12.5, 50 in experiment 2. Yields of ductular lesions, including intraductal papillomas and carcinomas, as well as papillo‐tubular and solid‐tubular carcinomas, showed dependence on the dose, with the respective total incidences being 0, 4.8, 25, 72.2 and 0, 10 and 35%. There was thus no apparent carcinogen exposure threshold. The present results confirmed the carcinogenicity demonstrated in a previous study using F344 rats and revealed the SD rat strain to be more susceptible.
p53 Mutations in Two Patients with Intraductal Papillary Adenoma of the Pancreas
Authors: Kaino Miyuki , Kondoh Satoshi , Okita Satoshi , Ryozawa Syomei , Hatano Satoko , Shiraishi Kei , Kaino Seiji , Akiyama Tetsuji , Okita Kiwamu , Kawano Toyokazu
Published: August 2005

There has been no report on p53 gene mutation in benign human pancreatic intraductal tumors. We examined pancreatic juice and tissue specimens from two patients with intraductal papillary adenoma of the pancreas by polymerase chain reaction‐single‐strand conformation polymorphism analysis and direct sequencing and found point mutations of p53 gene resulting in amino acid substitutions in exons 6 and 8. Thus, p53 gene mutation may be an early event in the neoplastic process of some pancreatic intraductal tumors and may play an important role in tumorigenesis.
Growth Characteristics in the Initial Stage of Micrometastasis Formation by Bacterial LacZ Gene‐tagged Rat Prostatic Adenocarcinoma Cells
Authors: Kobayashi Kiyoshi , Nakanishi Hayao , Inada Ken‐ichi , Fujimitsu Yasunobu , Yamachika Takasuke , Shirai Tomoyuki , Tatematsu Masae
Published: August 2005

A micrometastasis model was established using a rat differentiated prostatic adenocarcinoma, designated PLS301Z, transfected with the lacZ gene encoding a bacterial β‐galactosidase. The morphology, tumorigenicity and metastatic ability of PLS301Z were comparable to those of the parental cells. Micrometastatic foci could be specifically detected at the single cell level after X‐Gal staining with a dissecting microscope. After intravenous injection, the number of X‐Gal positive foci in the lung decreased progressively to a steady‐state level (less than 1% of injected cells) by 4–7 days, while the size of persisting positive foci started to increase from 4 days after inoculation, as demonstrated by image analysis. X‐Gal and BrdU double staining revealed that BrdU labeling indices of X‐Gal‐positive cells decreased transiently at the 2‐day time point and increased again from 4 days after inoculation. Type IV collagen immunostaining showed the tumor cells to be surrounded by a basement membrane intravascularly at the time point when they started new growth. Electron microscopy confirmed that, 2 days post injection, most tumor cells were degenerative or dead, but on day 4, persisting tumor cells formed multicellular clumps in contact with the vascular basement membrane inside vessels. These results indicate that PLS301Z cells begin to grow intravascularly depending upon the presence of a basement membrane before extravasation at the initial stage of micrometastasis formation.
CD44H Plays an Important Role in Peritoneal Dissemination of Scirrhous Gastric Cancer Cells
Authors: Nishimura Shigehiko , Chung Yong‐Suk , Yashiro Masakazu , Inoue Tohru , Sowa Michio
Published: August 2005

The role of the adhesion molecule CD44H in the peritoneal adhesion and invasion of cancer cells was assessed using cell lines with low and high peritoneal seeding ability, OCUM‐2M (2M) and OCUM‐2MD3 (2MD3), respectively. The in vitro binding ability to peritoneal components (mesothelial cells, fibroricetin and type I collagen) and invasive ability of 2MD3 cells were higher than those of 2M cells. The expression level of CD44H on 2MD3 cells was higher than that on 2M cells as determined by western blot analysis and flow cytometry. The adhesiveness of 2MD3 cells to hyaluronic acid, which is expressed on the surfaces of mesothelial cells, was greater than that of 2M cells. The binding ability of 2MD3 cells to mesothelial cells was inhibited in the presence of anti‐CD44H monoclonal antibody, but that of 2M cells was not. These results suggested that the 2MD3 cell binding to mesothelial cells is regulated by the CD44‐hyaluronic acid dependent system. The in vitro binding to submesothelial components and the invasiveness of 2MD3 cells were also inhibited in the presence of anti‐CD44H antibody. The in vivo inoculation of 2MD3 cells treated with an anti‐CD44H antibody resulted in a significant prolongation of survival time as compared with control mice that were inoculated with 2MD3 cells alone. In conclusion, CD44H was associated with attachment not only to hyaluronic acid on mesothelial cells, but also to peritoneal stromal components. Thus, CD44H may play an important role in cancer cell binding and invasion in the peritoneal dissemination of scirrhous gastric cancer cells.
Antitumor Effect of Streptococcus pyogenes by Inducing Hydrogen Peroxide Production
Authors: Higuchi Yoshihiro
Published: August 2005

The Su strain of Streptococcus pyogenes (S‐coccus)‐derived anticancer preparation, OK‐432, which is immobilized by heating in the presence of penicillin G, is well known to have an immunopotentiating activity through activation of natural killer cells in vivo. In this study, a streptococcal anticancer preparation stronger than OK‐432 was prepared. Live streptococci (S‐cocci) of the Su strain were induced to acquire H2O2‐producing ability by treatment with serum under aerobic conditions. The resulting preparation no longer possessed hemolytic activity, and was not viable. The serum‐treated S‐coccus preparation activated natural killer cells as well as OK‐432 did, and had stronger antitumor activity than OK‐432 did. These results suggest that the serum‐treated S‐coccus preparation would be a useful tool for chemotherapy, in addition to immunotherapy, for the treatment of cancer.
No Alteration in DNA Topoisomerase I Gene Related to CPT‐11 Resistance in Human Lung Cancer
Authors: Ohashi Nobuyuki , Fujiwara Yasuhiro , Yamaoka Naoki , Katoh Osamu , Satow Yukio , Yamakido Michio
Published: August 2005

Mutations and reduced expression of DNA topoisomerase I (topo I) gene have been shown to be important in the acquisition of resistance to camptothecin and its analogues in vitro, but their significance has not been verified in vivo. We investigated possible topo I gene mutations and topo I mRNA expression levels in 127 samples obtained from 56 patients with lung tumors, including patients who had developed clinical resistance to topo I inhibitors. No mutations were detected in any of the samples examined and expression levels did not differ significantly between clinically resistant cases and others. However, the topo I mRNA expression level was significantly higher in small cell lung carcinomas than in non‐small cell lung carcinomas (P<0.05). These results suggest that topo I mRNA levels may affect CPT‐11 sensitivity in human lung cancer. However, topo I gene mutations and reduced topo I mRNA expression may not be the main mechanism of clinically acquired resistance to camptothecin analogues in vivo.
Dose‐dependent Induction of 8‐Hydroxyguanine and Preneoplastic Foci in Rat Liver by a Food‐derived Carcinogen, 2‐Amino‐3,8‐dimethylimidazo[4,5‐f]quinoxaline, at Low Dose Levels
Authors: Kato Toshio , Hasegawa Ryohei , Nakae Dai , Hirose Masao , Yaono Makoto , Cui Lin , Kobayashi Yozo , Konishi Yoichi , Ito Nobuyuki , Shirai Tomoyuki
Published: August 2005

Male F344 rats were administered 2‐amino‐3,8‐dimethylimidazo[4,5‐f]quinoxaline (MeIQx) in the diet at doses of 200, 50, 12.5, 3.2, 0.8, 0.2 and 0.05 ppm for 6 weeks, and partially hepatectomized 1 week after the beginning of MeIQx administration. Quantitative values for glutathione S‐transferase placental form (GST‐P)‐positiye foci in the liver were dose‐dependently increased by the MeIQx treatment. 8‐Hydroxyguanine (8‐OHG) levels assessed after 1 week of dietary MeIQx administration were also dose‐dependently increased, although the effect was no longer observed at the end of the treatment period. The correlation between numbers of GST‐P‐positive foci at week 6 and 8‐OHG levels at week 1 was linear, values for both parameters being higher than the control levels even in the 0.8 ppm dose group. These findings indicate that, in addition to the previously reported MeIQx‐DNA adduct formation, DNA modifications due to oxidative damage may play an important role in MeIQx liver carcinogenesis in rats.
Establishment of a Hepatocyte Cell Line Producing Growth‐promoting Factors for Liver‐colonizing Tumor Cells
Authors: Yamori Takao , Shimada Koji , Kanda Hiroaki , Nishizuru Yumiko , Komi Akiko , Yamazaki Kanami , Asanoma Keiko , Ogawa Masako , Nomura Kimie , Nemoto Nobuo , Kumada Kaoru , Tsuruo Takashi
Published: August 2005

A hepatocyte‐derived cell line designated MLE‐15A2 was established from a primary culture of mouse hepatocytes. The MLE‐15A2 cells appeared to retain the basic nature of hepatocytes in that they showed morphology of an epithelial cell type and secreted albumin into the culture medium. These cells were grown on collagen‐coated plates and could be easily expanded to a large‐scale culture. Therefore, MLE‐15A2 cells may provide a more useful model for studying liver microenvironments than primary cultures of hepatocytes. We found that conditioned media from MLE‐15A2 cells, as well as from primary cultures of hepatocytes, promoted the proliferation of highly liver‐colonizing colon 26 NL‐17 cells better than the poorly liver‐colonizing colon 26 NL‐4 cells. Moreover, the conditioned media stimulated the growth of some human colon cancer cell lines. These results indicate that MLE‐15A2 cells secrete growth factors that selectively stimulate certain tumor cell types. Hepatocyte‐derived growth factors may regulate selective survival and colonization of tumor cells in the process of liver metastasis. The growth‐promoting activity was unaffected by dialysis, was stable at 80°C for 30 min and was bound to a heparin‐Sepharose column. The major activity was eluted from the column with 0.7–0.75 M NaCl, and some minor activities eluted with lower concentrations of NaCl. These results suggest that the active components are heterogeneous heparin‐binding proteins with lower affinity to heparin than platelet‐derived and fibroblast growth factors.
Cell Death Induced by Baicalein in Human Hepatocellular Carcinoma Cell Lines
Authors: Matsuzaki Yuko , Kurokawa Norie , Terai Shuji , Matsumura Yasuhiro , Kobayashi Nobuyuki , Okita Kiwamu
Published: August 2005

We examined the action of baicalein, a flavonoid contained in the herbal medicine sho‐saiko‐to (TJ‐9), on three cell lines of human hepatocellular carcinoma (HCC). Treatment with baicalein strongly inhibited the activity of topoisomerase II and suppressed the proliferation of all three HCC cell lines. But the mode of cell death induced by baicalein differed according to the cell line. Baicalein induced apoptosis in a concentration‐dependent mannner in only one cell line, and an increased concentration of baicalein produced cell death via necrosis in the other two lines. These results suggest that the inhibition of topoisomerase II is not by itself sufficient for induction of apoptosis, and that there is a more important mechanism which can account for the difference in susceptibility of cells to apoptosis induced by baicalein.
Suppressive Effects of S‐Methyl Methanethiosulfonate on Promotion Stage of Diethylnitrosamine‐initiated and Phenobarbital‐promoted Hepatocarcinogenesis Model
Authors: Sugie Shigeyuki , Okamoto Kiyohisa , Ohnishi Masami , Makita Hiroki , Kawamori Toshihiko , Watanabe Tomoyuki , Tanaka Takuji , Nakamura Yasushi K. , Nakamura Yoshiyuki , Tomita Isao , Mori Hideki
Published: August 2005

Modifying effects of S‐methyl methanethiosulfonate (MMTS) on diethylnitrosamine (DEN)‐initiated and phenobarbital (PB)‐promoted hepatocarcinogenesis were examined in rats. Five‐week‐old male F344 rats were divided into 8 groups. After a week, groups 1–5 were given DEN (100 mg/kg body weight, i.p.) once a week for 3 weeks, whereas groups 6–8 received vehicle treatment. Group 2 was given 100 ppm MMTS containing diet in the initiation phase. From 4 weeks after the start of experiment, groups 3 and 5 were fed MMTS, and groups 1–3 and 7 received drinking water containing 500 ppm PB. Group 6 was given MMTS diet alone throughout the experiment (24 weeks). The incidences of hepatocellular adenoma and total liver tumors were significantly smaller in group 3 than those of group 1. The average numbers of hepatocellular adenoma, carcinoma and total tumors in group 3 were significantly smaller than in group 1. Glutathione, S‐transferase placental form‐positive foci were also significantly decreased by MMTS treatment in the promotion phase. MMTS treatment in the initiation or promotion phase reduced ornithine decarboxylase activity in the liver of rats given DEN. The antioxidant activity against lipid peroxidation of MMTS was confirmed in tests with rabbit erythrocyte membrane ghosts or rat hepatocytes. These results suggest that MMTS is a promising chemopreventive agent for liver neoplasia when concurrently administered with PB.
Hypersensitivity to Mosquito Bites Conceals Clonal Lymphoproliferation of Epstein‐Barr Viral DNA‐positive Natural Killer Cells
Authors: Ishihara Shigehiko , Ohshima Koichi , Tokura Yoshiki , Yabuta Reiko , Imaishi Hidenori , Wakiguchi Hiroshi , Kurashige Takanobu , Kishimoto Hiroshi , Katayama Ichiro , Okada Shintaro , Kawa‐Ha Keisei
Published: August 2005

In order to clarify the relationship between Epstein‐Barr (EB) virus and hypersensitivity to mosquito bites (HMB), and to search for the mechanism which induces EB virus‐associated lymphoproliferative diseases, we investigated patients with HMB, using hematological, immunological and virological techniques. Among 5 cases of HMB, CD56+ cells had proliferated and CD3+ cells were diminished in 4 cases. Although anti‐EB virus antibody titers were not consistent with chronic active EB virus infection, EB viral DNA was detected in the peripheral blood mononuclear cells in all 5 cases. Moreover, EB viral DNA‐positive cells had proliferated monoclonally in 4 cases, and hiclonally in 1 case. It was proved that most of the EB viral DNA existed in natural killer (NK) cells through polymerase chain reaction analysis. These findings suggest that the basis of HMB may be clonal lymphoproliferation of EB viral DNA‐positive NK cells and this hematological abnormality may induce the characteristic symptoms of HMB. In some cases, the proliferating NK cells can metamorphose into leukemic cells, and hemophagocytic syndrome, which has been assumed to be a complication of HMB, may then occur.
Overexpression of Cyclin‐dependent Kinase‐activating CDC25B Phosphatase in Human Gastric Carcinomas
Authors: Kudo Yasusei , Yasui Wataru , Ue Teruyoshi , Yamamoto Soichiro , Yokozaki Hiroshi , Nikai Hiromasa , Tahara Eiichi
Published: August 2005

CDC25 phosphatases activate cyclin‐dependent kinases by removing inhibitory phosphate groups on the molecules and positively regulate the cell cycle progression. The expression of CDC25A, B and C was examined in gastric carcinoma cell lines and gastric carcinoma tissues by northern blotting and immunohistochemistry. The gastric carcinoma cell lines expressed CDC25A, B and C mRNA at various levels. The expression levels of CDC25B were generally higher than those of CDC25A and C. Of the 40 gastric carcinomas, 70% of the tumors expressed CDC25B mRNA at higher levels than the corresponding normal mucosas, while 38% overexpressed CDC25A mRNA. The CDC25C expression was at very low or undetectable levels. No obvious correlation was detected between the expression of CDC25B and p53 gene mutations. Inununohistochemically, CDC25‐positive tumor cells were detected in 43 (78%) of 55 gastric carcinoma cases, of which 27 (49%) were strongly positive. Strong expression of CDC25B protein was associated with advanced stage and deep invasion. Furthermore, the incidence of strong expression was significantly higher in carcinomas with nodal metastasis than in those without metastasis. These findings suggest that Overexpression of CDC25B may favor development and progression and may be an indicator of malignant behavior of gastric carcinomas.
Expression of Interleukin‐6 and Its Effect on the Cell Growth of Gastric Carcinoma Cell Lines
Authors: Ito Reiko , Yasui Wataru , Kuniyasu Hiroki , Yokozaki Hiroshi , Tahara Eiichi
Published: August 2005

The expression and the effect of IL‐6 were examined in human gastric carcinoma cell lines to determine whether IL‐6 serves as a growth stimulator. The expression of IL‐6 mRNA was detected in three (TMK‐1, MKN‐1, MKN‐7) of 8 gastric carcinoma cell lines. All three cell lines secreted IL‐6 into the culture fluid, in large amounts in the cases of MKN‐1 and MKN‐7 cells. Scatchard plot analysis of IL‐6 binding revealed that MKN‐1 and MKN‐7 cells had both high‐ and low‐affinity receptors. Cell growth of MKN‐1 and MKN‐7 cells was stimulated by IL‐6, while anti‐IL‐6 antibody inhibited growth. The expression of IL‐la mRNA by these three cell lines was induced by IL‐6. IL‐la increased the expression of mRNA for IL‐6 by TMK‐1 cells. These findings indicate that IL‐6 induced by IL‐la is an autocrine growth factor for some gastric carcinomas
Increased Telomerase Activities in Human Pancreatic Duct Adenocarcinomas
Authors: Tsutsumi Masahiro , Tsujiuchi Toshifumi , Ishikawa Osamu , Majima Toshimitsu , Yoshimoto Masatoshi , Sasaki Yasutaka , Fukuda Tomokazu , Oohigashi Hiroaki , Konishi Yoichi
Published: August 2005

Telomerase is a key enzyme with regard to immortalization of cancer cells and increased activity has been demonstrated in various human malignant neoplasms. Since little is known of its role in pancreatic cancers, we investigated changes in telomerase activity in human pancreatic duct adenocar‐cinomas and compared the frequency of increased telomerase activity with the presence of K‐ras gene mutations. The samples were obtained from 38 pancreatic duct adenocarcinomas and 7 tumor surrounding tissues at surgical resection. Telomerase activity was examined by telomeric repeat amplification protocol assay and terminal restriction fragment (TRF) length was examined by Southern analysis. K‐ras mutation was examined by means of polymerase chain reaction‐single strand conformation polymorphism analysis. Among 38 pancreatic carcinomas, 32 (84%) exhibited increased telomerase activities with no apparent relation to the histological type of tumor, tumor size, regional lymphnode involvement and distant metastasis or clinical stage. In tissue surrounding the tumor, telomerase activity was not detected. TRF length tended to be reduced in pancreatic carcinomas. Mutations of K‐ras gene were found in 24 out of the 38 (63%) cases. Among the 38 cases, 14 showed increased telomerase activity without K‐ras mutation and 4 cases showed K‐ras mutation without telomerase activity. These results suggest that increased telomerase activity might be a sensitive genetic diagnostic marker and could be a target for future therapy of pancreatic duct carcinomas.
In vitro Enhancement of Antitumor Activity of a Water‐soluble Duocarmycin Derivative, KW‐2189, by Caffeine‐mediated DNA‐repair Inhibition in Human Lung Cancer Cells
Authors: Ogasawara Hayato , Nishio Kazuto , Ishida Tomoyuki , Arioka Hitoshi , Fukuoka Kazuya , Saijo Nagahiro
Published: August 2005

Duocarmycins, including KW‐2189, bind in the minor groove of double‐stranded DNA at A‐T‐rich sequences, followed by covalent bonding with N‐3 of adenine in preferred sequences. We examined the effect of DNA‐repair modulators, such as caffeine and aphidicolin, on the cytotoxicity of duocarmycins towards human lung cancer cells, as determined by dye formation assay. Caffeine (0.5 or 1 mM), but not aphidicolin, enhanced the growth‐inhibitory activity of KW‐2189, DU‐86, and duocarmycin SA. Caffeine inhibited repair of DNA strand breaks induced by KW‐2189, as assayed by the alkaline elution technique. This suggests that duocarmycin‐induced DNA strand breaks, which are potentially lethal to cells, are repaired through a caffeine‐sensitive pathway.
Inhibitory Effect of NS‐398, a Selective Cyclooxygenase‐2 Inhibitor, on Azoxymethane‐induced Aberrant Crypt Foci in Colon Carcinogenesis of F344 Rats
Authors: Yoshimi Naoki , Kawabata Kunihiro , Hara Akira , Matsunaga Kengo , Yamada Yasuhiro , Mori Hideki
Published: August 2005

Prostaglandin E2, which is produced by cyclooxygenase (COX) during arachidonic acid metabolism, is considered to be related to colon Carcinogenesis. Therefore, the effect of NS‐398 (N‐(2‐cyclohexyl‐oxy‐4‐nitrophenyl)methanesulfonamide), a COX‐2 inhibitor, was examined in azoxymethane (AOM)‐induced colon Carcinogenesis in rats in this study. In the first experiment, groups 1–3 were treated with AOM (15 mg/kg, s.c.) 3 times at intervals of a week from 5 weeks of age. Groups 2 and 3 were respectively given 1 mg/kg and 10 mg/kg of NS‐398 in 5% gum arable aqueous solution 3 times per week by oral garage during the experiment. Six weeks after the first exposure to AOM, aberrant crypt foci (ACF) were counted in the colonic mucosa of all rats. The mean occurrence of ACF per length in rats given 1 mg/kg b.w. or 10 mg/kg b.w. of NS‐398 was reduced to 65.7% or 52.8%, respectively, of that in rats treated with only AOM. Levels of COX‐2 mRNA expression in groups treated with AOM, regardless of NS‐398, were slightly higher than that in the group treated with NS‐398 alone as judged from reverse transcription‐polymerase chain reaction analysis. In the second experiment, the effect of NS‐398 at different times, i.e., during initiation and post‐initiation, was examined. Treatment with NS‐398 in both phases significantly inhibited the appearance of ACF. The results imply that NS‐398 might have a chemopreventive potential.
In vitro Efficacy of Anti‐glial Fibrillary Acidic Protein Monoclonal Antibodies against Human Malignant Glioma Cell Lines
Authors: Abaza Mohamed‐Salah I. , Narayan Raj K. , Atassi M. Z.
Published: August 2005

Our studies have confirmed the presence of large concentrations of various intermediate filament proteins (IFPs) in glioma tissue compared to normal brain. This avenue of research was extended to assess the anti‐proliferative activity of anti‐intermediate filament protein monoclonal antibodies (anti‐IFP mAbs)against human glioma cells. In this study, anti‐proliferative activity of glial fibrillary acidic protein monoclonal antibodies (anti‐GFAP mAbs) has been tested in vitro, using glioma cell lines prepared and established from freshly resected brain tumors. One anaplastic astrocytoma (AA), two glioblastoma multiforme (GB1 and GB2) cell lines and three anti‐GFAP mAbs (Bi2C4, B12B4 and B6C6, all IgG15, kappa) were used. Immunofiuorescence study indicated the ability of anti‐GFAP mAbs to recognize the cell surface of glioma cells and the inhibition study showed that mAb B12B4 inhibited the proliferation of GB1, (96%), GB2(85%)and AA(93%) at a concentration of 3.2x 10−10M. mAb B12C4 inhibited the proliferation of GB1 (95%), GB2 (86%) and A A (91%) at a concentration of 3.26 X10‐10M and mAb B6C5 inhibited the proliferation of GB1 (75%), GB2 (75%) and AA (91%) at a concentration of 2.074 X10 −19M. Thymidine release assay demonstrated the cytolytic activities of anti‐GFAP mAbs towards these glioma cell lines, and this observation was confirmed by dye exclusion, which indicated the lysis of glioma cells after anti‐GFAP mAbs treatment. Anti‐GFAP mAbs had little effect (<120%) on normal human lymphocyte, liver and intestine cell lines. These results look promising for radioimaging and immunotherapy of human gliomas.
Dual Infection of Rabbits with Human T Cell Lymphotropic Virus Types I and II
Authors: Machida Hisanori , Morishita Nobumasa , Ikezoe Takayuki , Sawada Takashi , Ohtsuki Yuji , Miyoshi Isao
Published: August 2005

Attempts were made to generate a rabbit model of dual infection with human T lymphotropic virus (HTLV) types I and II. Four groups (A, B, C, and D) of three rabbits each were used. Group A was inoculated with the RW‐1 cell line coinfected with HTLV‐I and HTLV‐II and group B was transfused from a dually infected rabbit. Polymerase chain reaction (PCR) using primers specific for the pol region of each virus detected both HTLV‐I and HTLV‐II in all group A and two group B rabbits, but HTLV‐II only in the remaining group B rabbit. Groups C and D already infected with HTLV‐I and HTLV‐II, respectively, were inoculated with an HTLV‐II‐ or HTLV‐I‐producing cell line. One group C rabbit became PCR‐positive for both viruses but the other five resisted superinfection with the respective viruses. During prolonged observation, three of the six dually infected rabbits converted to single (HTLV‐I or HTLV‐II) infection. The in vivo dual infection was confirmed by in vitro establishment of a lymphoid cell line coinfected with HTLV‐I and HTLV‐II. It was also possible to establish coinfected lymphoid cell lines from HTLV‐I‐infected rabbits by coculture with lethally irradiated HTLV‐II‐producing cells and vice versa. The mechanism of viral elimination in dually infected rabbits, as well as that of protective immunity against superinfection, remains to be elucidated.
Direct Intratumoral Gene Transfer of the Herpes Simplex Virus Thymidine Kinase Gene with DNA‐liposome Complexes: Growth Inhibition of Tumors and Lack of Localization in Normal Tissues
Authors: Takakuwa Koicni , Fujita Kazuyuki , Kikuchi Akira , Sugaya Susumu , Yahata Tetsuro , Aida Hiroshi , Kurabayashi Takumi , Hasegawa Isao , Tanaka Kenichi
Published: August 2005

To constitute the site‐specific expression of the herpes simplex virus thymidine‐kinase (HSV‐TK) gene in tumor cells, we have assessed the promoter function of the simian virus 40 (SV40) promoter and the 5’flanking region of c‐erbB‐2 gene using a luciferase‐expressing reporter plasmid. After the transfec‐tion of the luciferase plasmid directed by the promoter region of c‐erbB‐2 gene, a large amount of luciferase activity was observed in c‐erbB‐2‐expressing cells (Colo201, MCF‐7, and HEC1‐A), while none was detected in cells with no expression of c‐erbB‐2 protein (HRA and KF cells). On the other hand, a high level of luciferase activity was detected in all tumor cell lines tested, when the transfection was performed with SV40 promoter. The repeated transfection of the liposome‐conjugated HSV‐TK gene regulated by the SV40 promoter or by the promoter region of c‐erbB‐2 gene with cultivation in 100 μg/ml of aciclovir for 5 days in vitro resulted in growth inhibition for all four cell lines examined or for only c‐erbB‐2‐expressing cells in the presence of SV40 promoter or c‐erbB‐2 promoter, respectively. Finally, direct injection of the DNA‐liposome complex into established tumors in the presence of 50 mg/kg of aciclovir led to significant tumor volume reduction in all three tumors tested when SV40 promoter was employed. However, this anti‐tumor effect was noted only in c‐erbB‐2‐positive cells (Colo201 cells) upon intratumoral injection of HSV‐TK gene regulated by c‐erbB‐2 promoter. In the case of intratumoral gene transfer, foreign DNA was detected in only one of seven mice by polymerase chain reaction (PCR) analysis performed 7 days following injection. When PCR analysis was carried out at 14 or 21 days following injection, no DNA signal was found at all. However, DNA was detected in several normal tissues at all three times tested in the case of intravenous injection. No abnormalities were seen in histologic examinations of normal tissues or in serum biochemical parameters following DNA liposome delivery. These results suggest that the direct gene transfer of HSV‐TK gene regulated by tumor‐specific transcriptional units may be one of the most clinically promising of the selective genetic strategies against cancer.
Histocompatibility Leukocyte Antigen‐A2402‐restricted Cytotoxic T Lymphocytes Recognizing Adenocarcinoma in Tumor‐infiltrating Lymphocytes of Patients with Colon Cancer
Authors: Gohara Rumi , Nakao Masanobu , Ogata Yutaka , Isomoto Hiroharu , Oizumi Kotaro , Itoh Kyogo
Published: August 2005

To cast light on T cell‐mediated specific immunity at the tumor site of colon cancer, we investigated whether interleukin‐2 (IL‐2)‐activated tumor‐infiltrating lymphocytes (TIL) from colon cancer show histocompatibility leukocyte antigen (HLA)‐class I‐restricted cytotoxicity against adenocarcinoma. IL 2‐activated TIL from all four HLA‐A24 patients examined lysed HLA‐A2402+ adenocarcinomas, but not HLA‐A2402 tumors. Those of two of the four cases also lysed HLA‐A2402+ squamous cell carcinomas. CD8+ cytotoxic T lymphocyte (CTL) clones recognizing HLA‐A2402+ adenocarcinomas were established from one CTL line. This CTL line produced IFN‐γ upon recognition of an HLA‐A2402‐ adenocarcinoma transfected with HLA‐A2402 cDNA. These results suggest the presence of HLA‐A2402‐restricted CTL recognizing adenocarcinoma at the tumor site of colon cancer. Furthermore, HLA‐A31‐restricted CTL activity was found in IL‐2‐activated TIL from one of two HLA‐A31+ patients, suggesting the existence of HLA‐class I‐restricted CTL involving an allele other than A24
Decreased Renal Accumulation of Biotinylated Chimeric Monoclonal Antibody‐Neocarzinostatin Conjugate after Administration of Avidin
Authors: Otsuji Eigo , Yamaguchi Toshiharu , Yamamoto Kazuhito , Matsumura Hiroomi , Tsuruta Hiroshi , Yata Yoshihiro , Nishi Hiroshi , Okamoto Kazuma , Kitamura Kazuya , Takahashi Toshio
Published: August 2005

Murine monoclonal antibodies (mAbs) such as A7 administered to humans induce a human anti‐mouse antibody response. Moreover, because Fab fragments of mAbs are able to penetrate target tumors easily, they may be more suitable than intact mAb to be carriers of anticancer agents such as neocarzinostatin (NCS), which are rapidly inactivated in the blood. To address these problems, chimeric A7 Fab fragment‐NCS conjugate (chA7Fab‐NCS) was produced. However, large amounts of 12SI‐labeled chA7Fab‐NCS accumulate in the kidney and can lead to renal dysfunction. To decrease renal accumulation of chA7Fab‐NCS, chA7Fab was biotinylated and administered with a subsequent injection of avidin. Human pancreatic carcinoma‐bearing nude mice were injected with 125I‐labeled biotinylated chA7Fab‐NCS with or without subsequent administration of avidin. The accumulation of 125I‐labeled biotinylated chA7Fab‐NCS in tissue samples was measured at appropriate time intervals. 12SI‐labeled biotinylated chA7Fab‐NCS was cleared more rapidly from the blood and the kidney with the administration of avidin than without it. There was no difference between tumor accumulation in these groups. The tumor/blood ratio of radioactivity of 125I‐labeled biotinylated chA7Fab‐NCS was significantly higher with subsequent administration of avidin than without avidin. The administration of biotinylated chA7Fab‐NCS followed by avidin may enhance safety and permit the administration of larger doses of NCS without the subsequent development of renal failure. A larger amount of 12SI‐labeled biotinylated chA7Fab‐NCS was retained in the liver and spleen with the subsequent administration of avidin than without avidin
RAPID COMMUNICATION
Authors: Suzuki Kazufumi , Daigo Yataro , Fukuda Seisuke , Tokino Takashi , Isomura Minoru , Isono Kaichi , Wainwright Brandon , Nakamura Yusuke
Published: August 2005

The high frequency of loss of heterozygosity that has been observed on the distal region of the long arm of chromosome 9 in squamous cell carcinomas of esophagus, lung, uterus, and head and neck indicates the presence of a tumor suppressor gene(s) in this region. To investigate the possible role of the PTC gene on chromosome 9q22.3, that was identified as the cause of nevoid basal cell carcinoma syndrome, during carcinogenesis in esophagus and lung, we examined 20 esophageal squamous cell carcinomas and 10 squamous cell carcinomas of the lung for mutations in any coding exon of PTC. Using single‐strand conformation polymorphism and direct sequencing, we detected no mutations other than two non‐deleterious polymorphisms. Our results suggest that inactivation of some tumor suppressor gene(s) on 9q other than PTC contributes to the development of squamous cell carcinomas in these tissues.
Strain Differences in Sensitivity to the Promoting Effect of Sodium L‐Ascorbate in a Two‐stage Rat Urinary Bladder Carcinogenesis Model
Authors: Murai Takashi , Mori Satoru , Hosono Motoko , Takashima Akira , Machine Setsuko , Oohara Tadao , Yamashita Hirofumi , Makino Susumu , Matsuda Tsutomu , Wanibuchi Hideki , Fukushima Shoji
Published: August 2005

Rat strain differences in sensitivity to the promoting effect of sodium L‐ascorbate (SA) on the development of urinary bladder tumors were investigated. In experiment 1, WS/Shi (WS), ODS/Shi‐od/od (ODS), and LEW/Crj (LEW) rats were initiated with 0.05%N‐butyl‐N‐(4‐hydroxybutyl)‐nitrosamine (BBN) in their drinking water and subsequently given basal Oriental MF diet (M) with or without a 5% SA supplement. In LEW rats the SA treatment increased the induction of neoplastic lesions in the urinary bladder, whereas WS and ODS animals proved unresponsive to its promoting effects. In experiment 2, WS and F344 rats were maintained on two kinds of commercial basal diets, M and CLEA CA‐1 (C), during administration of SA, since dietary factors can influence promoting effects. Feeding M during the promotion period in F344 rats yielded significantly more neoplastic lesions than feeding C, but in WS rats no such dietary influence was apparent. In experiment 3, strain differences in biosynthesis of α‐2u‐globulin (α2u‐g) were assessed because both α2u‐g in the urine and administration of sodium salts of organic acids such as SA have been reported to be involved in tumor promotion. Immunohistochemical analysis of renal tubules and Western blotting analysis of urine revealed the presence of α2u‐g in all three strains examined. These data suggest that differences in susceptibility to promotion are due to genetic factors rather than dietary factors and the ability to synthesize . α2u‐g.
Immunoreaction at 43 kDa with Anti‐ubiquitin Antibody in Breast Neoplasms
Authors: Iwaya Keiichi , Nishibori Hideki , Osada Takuya , Matsuno Yoshihiro , Tsuda Hitoshi , Sato Seiji , Kono Hiroaki , Fukutomi Takashi , Suzuki Minoru , Torikata Chikao , Iwamatsu Akihiro , Hirohashi Setsuo
Published: August 2005

Protein ubiquitination has been implicated in ATP‐dependent protein turnover and normal cell proliferation. To investigate whether the ubiqnitin‐mediated system is functionally involved in the cancerous state, we examined changes in protein ubiquitination in 52 surgically resected primary breast tumors. Imnmnohistochcmically, ubiquitin (Ub) was identified in the cytoplasm of cancer cells, which were stained more strongly than adjacent normal ductal epithelium. Corresponding immunoblot analysis of normal and neoplastic regions of human breast showed that the immunoreaction for Ub at about 43 kDa was increased in all of the tumors (100%), regardless of the clinical stage or histologic grade. This protein, which gave a single spot on two‐dimensional gel electrophoresis, had partial amino acid sequences which were identical to those of actin family members. Our results suggest that ubiquitination of this 43‐kDa protein may be involved in the carcinogenesis or biological characteristics of human breast neoplasms.
Bacillus Calmette‐Guérin Enhances Production and Secretion of Type IV Collagenases in Peripheral Blood Mononuclear Cells
Authors: Kageyama Yukio , Kawakami Satoru , Fujii Yasuhisa , Kihara Kazunori , Oshitna Hiroyuki
Published: August 2005

Intravesical administration of bacillus Calmette‐Guérin (BCG) is an effective and widely accepted treatment for superficial bladder cancer. Rapid progression of the disease after BCG therapy, however, has been reported in some cases refractory to the treatment. We examined whether BCG treatment and coexistence of peripheral blood mononuclear cells (PBMCs) alter the invasive potential of bladder cancer cells. Production and secretion of two type IV collagenases, matrix metalloproteinase (MMP) 2 and MMP 9, by PBMCs from five healthy donors or bladder cancer cells (T24, JTC 30, and JTC 32) were evaluated by gelatin zymography, western blot analysis, and northern blot analysis. Invasion of bladder cancer cells was also examined using reconstituted basement membrane (Matrigel). BCG (5, 50, and 500 μg/ml) had no effect on secretion of MMP 2 and MMP 9 by bladder cancer cells, but increased the production and secretion of MMP 9 by PBMCs in a dose‐dependent manner. The coexistence of PBMCs increased invasion of T24 cells and BCG further enhanced the invasion. Thus, BCG promotes invasion of bladder cancer cells under certain conditions. An increase in the secretion of MMP 9 by PBMCs may account in part for the effect.
Antitumor Effects of 5′‐Deoxy‐S‐fluorouridine in Combination with Recombinant Human Interleukin 2 on Murine Colon Carcinoma 26
Authors: Midoro Katsuo , Gotoh Kazunori , Houkan Takashi , Yukishige Koichi , Fujiwara Kôsaku , Ootsu Koichiro
Published: August 2005

The antitumor activity of rccombinant human interleukin 2 (rIL‐2) in combination with 5′‐deoxy‐5‐fluorouridine (doxifluridine; 5′‐DFUR) against murine colon carcinoma 26 (Colon 26) was studied. BALB/c mice were treated daily for 15 days with S'‐DFUR, rIL‐2 or both, beginning on day 7 after subcutaneous transplantation of Colon 26. While mice treated with 5′‐DFUR or rIL‐2 alone died of tumor growth with pulmonary metastases within 9 weeks posttransplantation, the survival time was significantly prolonged in mice treated with both 5′‐DFUR and rIL‐2. Most of the combination‐treated animals showed the regression of local tumors and the inhibition of pulmonary metastasis. Histopathologically, many tumor cells were degenerated and necrotized, with marked infiltration of mononuclear cells including large granular lymphocytes (LGLs) with periodic acid‐Schiff‐positive cytoplasmic granules. The cells were positive for CD3s, asialo GM1 and NK1.1. Spleen cells from the combination‐treated mice showed high activities of natural killer (NK) cytotoxicity as well as growth inhibition of Colon 26 and Meth A fibrosarcoma in mice. The results suggest that the combination therapy of 5′‐DFUR plus rIL‐2 enhanced non‐specific cytotoxicity of LGL/NK cells for Colon 26 in tumor‐bearing mice and was effective in the inhibition of tumor growth.
Inhibition by Green Tea Extract of Diethylnitrosamine–initiated but Not Cholinedeficient, L–Amino Acid–defined Diet–associated Development of Putative Preneo–plastic, Glutathione S–Transferase Placental Form–positive Lesions in Rat Liver
Authors: Tamura Kazutoshi , Nakae Dai , Horiguchi Kohsuke , Akai Hiroyuki , Kobayashi Yozo , Satoh Hiroshi , Tsujiuchi Toshifumi , Denda Ayumi , Konishi Yoichi
Published: August 2005

The effects of green tea extract (GTE) on exogenous and endogenous models of rat liver carcinogenesis using diethylnitrosamine (DEN) and a choline–deficient, L–amino acid–defined (CDAA) diet were studied. For the exogenous carcinogenesis study, male Fischer 344 rats, 6 weeks old, were given a single intraperitoneal dose of 200 mg/Kg body weight of DEN, partially hepatectomized at week 3, and administered GTE at doses of 0, 0.01 and 0.1% in the drinking water from week 2 for 10 weeks. For the endogenous carcinogenesis study, rate were fed the CDAA diet and simultaneously given GTE for 12 weeks. All rats were killed at the end of week 12. After DEN–initiation, the apparent numbers of glutathione S–transferase placental form–positive foci, assayed as putative preneoplastic lesions, were decreased by the administration of GTE, though their sizes were not altered. In contrast, GTE did not significantly reduce the numbers of the lesions induced by the CDAA diet or affect their sizes. While the levels of 8–hydroxyguanine, a parameter of oxidative DNA damage, were reduced by the GTE administration in both experimental models, GTE did not protect against the CDAA–diet–associated liver tissue damage in terms of either histology or plasma marker enzyme levels. We conclude that, while GTE may be a possible chemopreventive agent for nitrosamine–initiated hepato–carcinogenesis in the absence of chronic hepatocyte damage, it does not significantly inhibit lesion development in hepatocarcinogenesis associated with the CDAA diet, a cirrhosis–associated model.
PRLTS Gene Alterations in Human Prostate Cancer
Authors: Komiya Akira , Suzuki Hiroyoshi , Ueda Takeshi , Aida Sara , Ito Naomi , Shiraishi Taizo , Yatani Ryuichi , Emi Mitsuru , Yasuda Kosaku , Shimazaki Jun , Ito Haruo
Published: August 2005

Since loss of heterozygosity on 8p22‐p21,3 has been found frequently in prostate cancer, the status of a candidate tumor suppressor gene named PRLTS gene, recently cloned from the same region in some human malignancies, was examined in the present study. DNAs were isolated from 69 Japanese prostate cancer patients (37 localized and 32 cancer‐death cases). Loss of heterozygosity at this gene locus was observed in 15 of 36 (42%) localized prostate cancer patients and 22 of 32 (69%) cancer‐death patients. One cancer‐death patient had a missense mutation, ACG→ATG (Thr→Met) at codon 64 in metastatic tumor tissues of pelvic lymph node and liver, and these tissues showed loss of the homologous allele, indicating that “two‐hit” mutation of the PRLTS gene had occurred in this case. The others did not show any mutation, regardless of the presence or absence of loss of heterozygosity. Although loss of heterozygosity at the PRLTS gene locus is a relatively common abnormality, mutation of this gene is rare in prostate cancer.
Radioimmunodetection with 111In–labeled Monoclonal Antibody Nd2 in Patients with Pancreatic Cancer
Authors: Chung Yong‐Suk , Sawada Tetsuji , Kondo Yasuyuki , Hirayama Koji , Inui Akimasa , Yamashita Yoshito , Nakata Bunzo , Okamura Terue , Ochi Hironobu , Ho Jenny J. L. , Kim Young S. , Sowa Michio
Published: August 2005

This report summarizes results from an initial clinical evaluation of radioimnumodetection (RAID) in patients with pancreatic cancer using murine monoclonal antibody Nd2, directed against mucins from pancreatic cancer.Nd2 (2 mg) was labeled with 111ln (2 mCi) and injected into 19 patients suspected of having pancreatic cancer. Planar scintigrams were taken 3 days post–infusion. As for final diagnoses after surgery, 14 cases were pancreatic cancer, and one case each was chronic pancreatitis, neurilem–moma, islet cell carcinoma, cholangioma, and apparent absence of suspected recurrent lesion of pancreatic cancer. Of 14– patients with pancreatic cancer, RAID was positive in 10 cases (71.4%). Cases other than pancreatic cancer were all negative, so the specificity was 100%. These results demonstrate that RAID using 111In–Nd2 can be useful in differentiating exocrine pancreatic cancer from benign conditions and other types of carcinomas in the pancreatoduodenal regions.
Pharmacokinetic Study of Carboplatin Given on a 5‐Day Intravenous Schedule
Authors: Ando Yuichi , Minami Hironobu , Saka Hideo , Ando Masahiko , Sugiura Seiji , Sakai Shuzo , Shimokata Kaoru
Published: August 2005

We investigated whether carboplatin pharmacokinetics is altered when the drug is delivered daily over 5 days, compared to a single‐day infusion. Carboplatin was infused in 11 patients with lung cancer, who were randomly assigned to 2 groups. In the first group, the agent was administered on a conventional single‐day schedule in the first course and then on a 5‐day schedule in the second course. In the second group, the order was reversed (crossover design). The dose was calculated using Calvert's formula with 24 h creatinine clearance (Ccr, ml/min) as a substitute for glomerular filtration rate (GFR): carboplatin (mg) = AUC X (Ccr+25), where AUC denotes the area under the concentration versus time curve (mg ml–1 min). No difference of carboplatin clearance between the single‐day and 5‐day schedule was observed (94.8± 19.9 versus 96.1+29.9 ml/min, P=0.818, paired t test). The formula systematically overestimated the carboplatin clearance; the ratio of estimated clearance/ observed clearance ranged from 1.01 to 1.58 (median 1.28; 95% confidence interval, 1.18 to 1.39). We concluded that the individual dosing strategy based on renal function can be applied with a 5‐day schedule as well as a single‐day schedule. Carboplatin is overdosed when Ccr is substituted for GFR in Calvert's formula.
Inhibition of Azoxymethane‐initiated Colon Tumor by Bovine Lactoferrin Administration in F344 Rats
Authors: Sekine Kazunori , Watanabe Emiko , Nakamura Joe , Takasuka Nobuo , Kim Dae Joong , Asamoto Makoto , Krutovskikh Vladimir , Baba‐Toriyama Hiroyasu , Ota Tomonori , Moore Malcolm A. , Masuda Mitsuharu , Sugimoto Hajime , Nishino Hoyoku , Kakizoe Tadao , Tsuda Hiroyuki
Published: August 2005

The influence of bovine lactoferrin (bLF) on colon carcinogenesis was investigated in male F344 rats treated with azoxymethane (AOM). Following three weekly injections of AOM, the animals received 2 or 0.2% bLF for 36 weeks. No effects indicative of toxicity were noted, but significant reduction in both the incidence and number of adenocarcinomas of the large intestine was observed with both doses. Thus, the incidences of adenocarcinomas in the groups receiving 2% and 0.2% bLF were 15% and 25%, respectively, in contrast to the 57.5% control value (P < 0.01 and P<0.05, respectively). The results indicate that bLF might find application for chemoprevention of colon cancer.
Protective Effect of Recombinant Human Leukemia Inhibitory Factor against Thrombocytopenia in Carboplatin‐treated Mice
Authors: Akiyama Yasuto , Kikuchi Yasufumi , Matsuzaki Junichi , Kajimura Naoko , Ohue Chiharu , Yamaguchi Ken
Published: August 2005

The protective effect of recombinant human leukemia inhibitory factor (rhLIF) against development of throrabocytopenia in carboplatin (CBDCA; cis‐diammine‐l, l‐cyclobutane dicarboxylate platinum II)‐treated mice was examined. The optimal dose of rhLIF was determined by administering doses of rhLIF ranging from 0.2 to 20 μg/day intraperitoneally to normal BALB/c mice, and by analyzing platelet counts. Platelet counts significantly increased at doses over 0.2 μg/day. When 20 μg/day rhLIF was injected for 7 days, platelet counts increased to 240% of control. A remarkable weight loss occurred in the 20μg/day group, but no weight loss was detected at doses of less than 10 μg/day. The rhLIF dose of 4 μg/day was therefore used to examine the protective effect against carboplatin‐induced thrombocytopenia in mice. The intravenous injection of 100 mg/kg carboplatin caused a significant thrombocytopenia with a nadir on day 8. The administration of 4 μg/day rhLIF intraperitoneally for 7 days promoted the recovery of platelet counts from day 5 after the injection of carboplatin. These results suggest that rhLIF at a suboptimal dose might be a useful therapeutic agent for chemotherapy‐induced thrombocytopenia.