ognizant Communication Corporation


VOLUME 12, NUMBER 4, 2000

Oncology Research, Volume 12, pp. 169-172, 2000
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Copyright © 2000 Cognizant Comm. Corp.
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Induction of Hepatic Metallothioneins in Tumor-Bearing IL-6-Null Mice

Andrei Molotkov, Masahiko Satoh, and Chiharu Tohyama

Environmental Health Sciences Division, National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba 305-0053, Japan

Interleukin-6 (IL-6) has been thought to play a key role in the induction of hepatic metallothionein (MT) synthesis in tumor-bearing animals. In order to clarify the role of IL-6 and to distinguish its effect from those of other cytokines, we inoculated IL-6-null and B6J129Sv (wild-type control) mice SC with 1 x 107 Ehrlich carcinoma cells and examined tumor growth, hepatic MT, and serum cytokines. We have found that tumor growth was followed by an increase of hepatic MT in both IL-6-null and wild-type mice and that the two strains of mice had a similar hepatic MT induction followed by zinc accumulation in the liver. These results could be explained by our finding that tumor-bearing IL-6-null mice had a high level of tumor-secreted IL-6 in the blood. In conclusion, by utilizing tumor-bearing IL-6-null mice, we have demonstrated that IL-6 secreted from the tumor cells is responsible for the tumor-evoked increase of hepatic MT level.

Key words: IL-6-null mice; Metallotioneins; Ehrlich ascites carcinoma; Tumor-bearing mice

Address correspondence to Chiharu Tohyama, Ph.D., Environmental Health Sciences Division, National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba 305-0053, Japan. Tel: +81-298-50-2336; Fax: +81-298-50-2574; E-mail: ctohyama@nies.go.jp

Oncology Research, Volume 12, pp. 173-179, 2000
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Copyright © 2000 Cognizant Comm. Corp.
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Human Interferon-g Enhances Expression of Ganglioside GM2 on Human Lung Cancer Cells and Their Susceptibility for Antiganglioside GM2 Monoclonal Antibody-Dependent Cellular Cytotoxicity

Masaki Hanibuchi, Yasuhiko Nishioka, Hiroaki Yanagawa, Seiji Yano, Prahlad Parajuli, Mika Bando, and Saburo Sone

Third Department of Internal Medicine, The University of Tokushima School of Medicine, Tokushima, Japan

Interferons are known to modulate several cellular functions by the induction of various proteins. In this study, we demonstrated that human interferon-g (HuIFN-g) enhanced the expression of ganglioside GM2 (GM2), which is a kind of tumor-associated antigen substantially expressed in human lung cancer and that human lung cancer cells expressing GM2 became more susceptible to anti-GM2 monoclonal antibody (mAb)-dependent tumor cell killing mediated by human effector cells after HuIFN-g treatment. GM2 expression on human lung cancer cells treated with or without HuIFN-g was measured by flow cytometry. The antibody-dependent cellular cytotoxicity (ADCC) activity was assessed by 4-h 51Cr release assay. HuIFN-g enhanced GM2 expression on human small-cell lung cancer (SCLC), SBC-3, and human non-small-cell lung cancer (NSCLC), A549 cells in a dose-dependent manner. The optimal concentration of HuIFN-g was 1000 U/ml. The effect of HuIFN-g reached maximum after 4 days of culture. HuIFN-g did not have any effect to enhance the expression of other gangliosides in SBC-3 cells. No other cytokines used in this study modulated GM2 expression in SBC-3 cells. Anti-GM2 mAb-dependent ADCC activities induced by lymphocytes and monocytes were more potent against IFN-g-treated SBC-3 and A549 cells than nontreated cells. Taken together, HuIFN-g combined with anti-GM2 mAb may be useful for immunotherapy against GM2-positive human lung cancer.

Key words: Human interferon-g; Ganglioside GM2; Lung cancer

Address correspondence to Saburo Sone, Third Department of Internal Medicine, The University of Tokushima School of Medicine, 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan. Tel: +81-886-33-7127; Fax: +81-886-33-2134; E-mail: ssone@clin.med.tokushima-u.ac.jp

Oncology Research, Volume 12, pp. 181-183, 2000
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Expression of the ATF3 Gene on Cell Lines and Surgically Excised Specimens

Tatsuaki Ishiguro and Hirokazu Nagawa

Department of Surgical Oncology, Faculty of Medicine, The University of Tokyo, 7-3-1 Yayoi Bunkyo-ku, Tokyo 113-8655, Japan

TI-241 (ATF3 homologue gene) is a metastasis-associated gene that is expressed at higher levels in the high-metastatic mouse B16 melanoma subline B16-FIO than in the low-metastatic clone Fl. We studied ATF3 expression in established cell lines of human colon and stomach cancer and surgically excised human colon cancer. ATF3 was expressed at higher levels in the cell lines that were established from metastatic sites than in those from original tumor sites. Also ATF3 was expressed at higher levels in tumor specimens than in adjacent normal mucosa, especially in the tumors that had invaded the lymphatic ducts and/or the vessels, while there was not a significant relationship between its expression and the other pathological features. As metastasis is the most important factor in determining prognosis, these results show the potential for ATF3 in making the prognosis.

Key words: ATF3 gene; Metastasis; Gene expression

Address correspondence to Dr. Tatsuaki Ishiguro, Department of Surgical Oncology, Faculty of Medicine, University of Tokyo, 7-3-1 Yayoi Bunkyo-ku, Tokyo 113-8655, Japan. Tel: +81-3-3811-6822; Fax: +81-3-3591-8246; E-mail: ti227241@monbu.go.jp

Oncology Research, Volume 12, pp. 185-192, 2000
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4-[3-(2-Nitro-1-imidazolyl)propylamino]-7-chloroquinoline Hydrochloride (NLCQ-1), a Novel Bioreductive Compound as a Hypoxia-Selective Cytotoxin

Maria V. Papadopoulou, Ming Ji, Mira K. Rao, and William D. Bloomer

The Radiation Medicine Institute at Evanston Northwestern Healthcare, 2650 Ridge Avenue, Evanston, IL 60201

A novel weakly DNA-intercalative bioreductive compound, 4-[3-(2-nitro-1-imidazolyl)-propylamino]-7-chloroquinoline hydrochloride (NLCQ-1), has been synthesized and studied as a hypoxia-selective cytotoxin in vitro. NLCQ-1, which shares a similar structure with the DNA-intercalative antimalarial drug chloroquine, bound more strongly to DNA than the nonchlorinated analog NLQ-1 (4-[3-(2-nitro-1-imidazolyl)propylamino]-quinaldine hydrochloride). Thus, NLCQ-1 exhibited a C50 [concentration for 50% displacement of the ethidium bromide (EB) from a DNA-EB complex] of 44 mM, whereas a C50 value could not be reached for NLQ-1 up to 225 mM. NLCQ-1 demonstrated significant hypoxic selectivity in several rodent (V79, EMT6, SCCVII) or human (A549, OVCAR-3) tumor cell lines. Its potency as a hypoxic cytotoxin (expressed as the product of exposure time and concentration for 50% survival) ranged between 10 and 136 mM·h, for the cell lines tested, at 30 mM input concentration. Because uptake in all cell lines was similar, the differences in potency may reflect differences in the enzymatic profile or damage repair processes among the cell lines. In addition, however, the most striking feature of NLCQ-1 was that hypoxic selectivity increased with exposure time, a common feature normally found in only bis-bioreductive agents carrying two moieties with different redox potentials. Thus, hypoxic selectivity of NLCQ-1 in V79 cells at 50% survival was increased from fivefold up to 388-fold by increasing exposure time from 1 to 4.5 h, as the result of a concomitant increase and decrease in its hypoxic and aerobic potency, respectively, over time. Because the nonchlorinated analog NLQ-1 did not demonstrate similar behavior, we hypothesized that the C-7 chlorine of NLCQ-1 might play a significant role in this phenomenon.

Key words: NLCQ-1; Bioreductive drug; Hypoxic cytotoxin; Weak DNA intercalation

Address correspondence to Maria V. Papadopoulou, Evanston Northwestern Healthcare, Department of Radiation Medicine, 2650 Ridge Avenue, Evanston, IL 60201. Tel: (847) 570-2262; Fax: (847) 570-1878; E-mail: m-papadopoulou@nwu.edu

Oncology Research, Volume 12, pp. 193-201, 2000
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Printed in the USA. All rights reserved.
Sodium Butyrate-Induced Differentiation of Human LIM2537 Colon Cancer Cells Decreases GSK-3b Activity and Increases Levels of Both Membrane-Bound and APC/Axin/GSK-3b Complex-Associated Pools of b-Catenin

Elizabeth Vincan,1 Christine S. F. Leet,1 Nancy I. Reyes,1 Rodney J. Dilley,2 Robert J. S. Thomas,1 and Wayne A. Phillips1

1Trescowthick Research Laboratories, Peter MacCallum Cancer Institute, East Melbourne, Victoria 3002, Australia
2Baker Medical Research Institute, Prahran, Victoria 3181, Australia

Analysis of the glycogen synthase kinase-3b (GSK-3b) activity in several colon cancer cell lines suggested a correlation between comparatively low enzyme activity and moderate to high differentiation status. Treatment of LIM2537 cells, a poorly differentiated colon cancer cell line, with the potent differentiating agent sodium butyrate resulted in 34% reduction in GSK-3b activity in the treated cells (P < 0.028, n = 3). Decreases in GSK-3b activity were paralleled by stabilization of cytoplasmic b-catenin, a hallmark of Wnt signaling. However, in contrast to Wnt signaling, expression of the b-catenin/TCF target genes c-myc and cyclin D1 did not appear to be increased in the sodium butyrate-treated cells. Interestingly, expression of membrane-bound b-catenin was increased in the sodium butyrate-treated cells. This suggests that, in the context of cellular differentiation, increases in b-catenin expression may be sequestered at the cell membrane and suggests that a possible role of sodium butyrate in promoting differentiation may be via increasing the levels of b-catenin available for cell adhesion.

Key words: b-Catenin; GSK-3b; Differentiation; Colon; Epithelium; Cancer

Address correspondence to Elizabeth Vincan, Ph.D., Trescowthick Research Laboratories, Peter MacCallum Cancer Institute, Locked Bag 1, A'Beckett Street, Melbourne, Victoria 8006, Australia. Tel: +61-3-9656 1842; Fax: +61-3-9656 1411; E-mail: e.vincan@pmci.unimelb.edu.au

Oncology Research, Volume 12, pp. 203-208, 2000
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Angiogenesis and p53 Expression in the Colorectal Adenoma-Carcinoma Sequence

Stylianos A. Kakolyris,1 Loukas G. Kaklamanis,2 Michael J. Koukourakis,3 Alexandra Giatromanolaki,4 Maria Rousomoustakaki,5 John C. Souglakos,1 Despina K. Reppa,1 Vassilis A. Georgoulias,1 Kevin C. Gatter,6 and Adrian L. Harris7

1Department of Clinical Oncology, 3Department of Radiotherapy, and 5Department of Gastrenterology, University Hospital of Heraklion, PO Box 1352, Heraklion 71110, Crete, Greece
2Department of Pathology, Onassis Cardiac Surgery Center, 356 Siggrou Avenue, Athens, 17674, Greece
4Histopathology Department, Democritus University, Alexandroupolis, Thrace, Greece
6Department of Cellular Science, and 7ICRF Molecular Oncology Laboratory, John Radcliffe Hospital, University of Oxford, OX3 9DU, UK

Angiogenesis, the formation of new vessels, is essential for tumor growth and metastasis. Mutations of p53 tumor suppressor gene are frequent and play an important role in colorectal oncogenesis. A role of p53 as an angiogenesis inhibitor has also been proposed. We evaluated angiogenesis and p53 expression in 16 hyperplastic polyps, 35 solitary tubular and tubulovillous adenomas, and 47 cases of sporadic colorectal carcinomas arising on the basis of preexisting adenomas, with standard immunohistochemical techniques. The mean microvessel density (MVD) in carcinomas was significantly higher compared with the respective adenomatous part of the same tumor (27.9 vs. 7; P = 0.0001). Linear regression analysis of MVD between cancerous and adenomatous areas showed a significant correlation (P = 0.0001, r = 0.56), raising the possibility that carcinomas arising from better vascularized adenomas might show increased vascularity. The MVD was significantly higher in stage C compared with stage A cases (P = 0.04). p53 positivity was detected in 26 of 47 cancerous (55%) and in 14 of 47 adenomatous areas (30%; P = 0.0002). All carcinomas arising from p53-positive adenomas were also p53 positive. p53 positivity associated with a higher MVD in adenomas (P = 0.02), but not in carcinomas (P = 0.78). We conclude that angiogenesis and p53 play a critical role in colorectal neoplasia, and the process of malignant transformation in tumors arising from highly angiogenic adenomas, particularly those carrying p53 mutations, is accelerated with rapid tumor progression from stage to stage, indicating a more aggressive tumor phenotype.

Key words: Angiogenesis; p53; Colorectal cancer; Immunohistochemistry

Address correspondence to Stylianos A. Kakolyris, M.D., Ph.D., Department of Clinical Oncology, University Hospital of Heraklion, Heraklion 71110, PO Box 1352, Crete, Greece. Tel: 0030-81-392747; Fax: 0030-81-392802; E-mail: GeorgSec@med.uch.gr