ognizant Communication Corporation



Oncology Research, Volume 14, pp. 407-414
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

4-[3,5-bis(trimethylsilyl)benzamido] Benzoic Acid Inhibits Angiogenesis in Colon Cancer Through Reduced Expression of Vascular Endothelial Growth Factor

Noritaka Minagawa,1 Yoshifumi Nakayama,1 Yuzuru Inoue,1 Koji Onitsuka,1 Takefumi Katsuki,1 Yosuke Tsurudome,1 Kazunori Shibao,1 Keiji Hirata,1 Tatsuhiko Sako,1 Naoki Nagata,1 Shinji Ohie,3 Kimitoshi Kohno,2 and Hideaki Itoh1

1Department of Surgery 1 and 2Department of Molecular Biology, University of Occupational and Environmental Health, School of Medicine, 1-1 Iseigaoka, Yahatanish-ku, Kita-kyushu 807-8555, Japan
3Cancer Research Laboratory, Hanno Research Center, Taiho Pharmaceutical Co., Ltd., 1-27 Misugidai, Hanno, Saitama 357-8527, Japan

4-[3,5-bis(trimethylsilyl)benzamido] Benzoic acid (TAC-101) has potent antiproliferative, antiangiogenic, and antitumor effects in vitro and in vivo. These effects might be due to TAC-101 binding to retinoic acid receptor a (RAR-a) and interfering with the binding of activator protein-1 (AP-1) to DNA. However, little is known about the detailed mechanism of TAC-101 function. We investigated the mechanism of the antiangiogenic effect of TAC-101 using a rat hepatic metastatic model in vivo and DLD-1 human colon cancer cells in vitro. Liver metastases were induced by portal injection of RCN-9 rat colonic cancer cells into F344 rats. TAC-101 (8 mg/kg) was orally administered 5 days per week for 4 weeks and then hepatic tumors were immunohistochemically evaluated for microvessel density (MVD) and vascular endothelial growth factor (VEGF). TAC-101 significantly reduced both MVD and VEGF expression. Northern blot analysis and ELISA indicated that TAC-101 efficiently inhibited production of VEGF mRNA and protein in DLD-1 cells in a time- and dose-dependent manner. These findings suggest that TAC-101 may inhibit progression and metastasis in colon cancer by interfering with tumor production of VEGF.

Key words: TAC-101; Vascular endothelial growth factor (VEGF); Colon cancer

Address correspondence to Noritaka Minagawa M.D., Department of Surgery 1, University of Occupational and Environmental Health, 1-1 Iseigaoka, Yahatanishi-ku, Kitakyushu 807-8555, Japan. Tel: 81-93-691-7441; Fax: 81-93-603-2361; E-mail: minagawa@med.uoeh-u.ac.jp

Oncology Research, Volume 14, pp. 415-426
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Resveratrol and Propolis as Necrosis or Apoptosis Inducers in Human Prostate Carcinoma Cells

Christian Scifo,1 Venera Cardile,2 Alessandra Russo,1 Rosanna Consoli,1 Carlo Vancheri,3 Francesco Capasso,4 Angelo Vanella,1 and Marcella Renis1

1Department of Biological Chemistry, Medicinal Chemistry and Molecular Biology, and 2Department of Physiological Sciences, Viale Andrea Doria, 6 and 3Department of Internal and Specialistic Medicine, Via Passo Gravina 187- University of Catania, 95125 Catania, Italy
4Department of Experimental Pharmacology, Via D. Montesano 49, University of Naples Federico II, 80131 Naples, Italy

Vegetables and fruit help the prevention and the therapy of several kinds of cancer because they contain micronutrients, a class of substances that have been shown to exhibit chemopreventive and chemotherapeutic activities. In the present study the effects of resveratrol (100 and 200 mM), a phytoalexin found in grapes, and of the ethanolic extract of propolis (50 and 100 mmg/ml), a natural honeybee hive product, were tested in androgen-resistant prostate cancer cells (DU145), a cell line resembling the last stage of prostate carcinoma. A comparison between the activity of these micronutrients and vinorelbine bitartrate (Navelbine®), a semisynthetic drug normally used in the therapy of prostate cancer, was conducted. Several biochemical parameters were tested, such as cell viability (MTT assay), cell membrane integrity (lactate dehydrogenase release), cell redox status (nitric oxide formation, reactive oxygen species production, reduced glutathione levels), genomic DNA fragmentation (COMET assay) with special attention on the presence of apoptotic DNA damage (TUNEL test), and possible mitochondrial transmembrane potential alteration (DY). Our results point out the anticancer activity of resveratrol and propolis extract in human prostate cancer, exerting their cytotoxicity through two different types of cell death: necrosis and apoptosis, respectively. The data obtained suggest the possible use of these micronutrients both in alternative to classic chemotherapy, and in combination with very low dosage of vinorelbine (5 mM).

Key words: Human prostate cancer; DU 145; Resveratrol; Propolis; Vinorelbine

Address correspondence to Dr. Marcella Renis, Associate Professor, Department of Biological Chemistry, Medicinal Chemistry and Molecular Biology, University of Catania, Viale Andrea Doria, 6, 95125 Catania, Italy. Tel: (+39) 095 7384081; Fax: (+39) 095 7384222; E-mail: renis@unict.it

Oncology Research, Volume 14, pp. 427-438
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Modulation of Wnt-Specific Colon Cancer Cell Kill by Butyrate and Lithium

Michael Bordonaro, Darina L. Lazarova, Rocco Carbone, and Alan C. Sartorelli

Department of Pharmacology and Developmental Therapeutics Section, Cancer Center, Yale University School of Medicine, New Haven, CT

Colorectal cancer (CRC) may be particularly amenable to gene therapy because CRCs exhibit constitutive upregulation of Wnt signaling. We have previously demonstrated that butyrate, found in the colonic lumen, modulates Wnt signaling and nonspecifically upregulates transcription from minimal promoters. Because both of these actions may influence the efficiency and specificity of Wnt-targeted expression, the effects of butyrate on Wnt-targeted gene therapy were determined. Lithium is another agent known to upregulate Wnt activity in HCT-116 CRC cells and therefore may induce Wnt-targeted CRC cell kill. CRC cells were cotransfected with an expression vector for green fluorescent protein (GFP) and different versions of vectors coupling Wnt-sensitive promoters to FADD or diphtheria toxin A-chain (DT) effector genes. Cells were treated with butyrate and/or lithium chloride and flow cytometry was used to determine the percentage of remaining transfected (GFP-positive) cells. We demonstrate that promoter and cell type-specific differences occur in Wnt-specific cell kill induced by FADD and DT. A Wnt-sensitive version of the CMV promoter (TcfCMV) exhibited the optimum combination of efficient SW620 CRC cell kill and Wnt specificity; in addition, treatment with a physiologically relevant concentration of butyrate enhanced cell kill induced by TcfCMV-FADD while maintaining specificity. In HCT-116 CRC cells, optimum results were achieved utilizing TcfFos-DT constructs and cotreatment with both butyrate and lithium. The findings suggest that effective CRC cell kill can be achieved by gene therapy through modulation of Wnt signaling by butyrate and/or lithium.

Key words: Wnt; Gene therapy; Butyrate; Lithium; Tcf; Colon cancer

Address correspondence to Michael Bordonaro, Department of Pharmacology, Yale University School of Medicine, 333 Cedar Street, PO Box 208066, New Haven, CT 06520. Tel: + 203-785-4390; Fax: + 203-785-2494; E-mail: michaelbordonaro@yahoo.com.

Oncology Research, Volume 14, pp. 439-445
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Cell-Free DNA and RNA in Plasma as a New Molecular Marker for Prostate Cancer

Eirini Papadopoulou,1 Elias Davilas,2 Vasilios Sotiriou,2 Alexander Koliopanos,3 Filipos Aggelakis,4 Konstantinos Dardoufas,4 Niki J. Agnanti,5 Irini Karydas,6 and Georgios Nasioulas1

1Molecular Biology Research Center HYGEIA "Antonis Papayannis," Kiffisias Ave. & 4 Erythrou Stavrou Str., 151 23 Marousi, Athens, Hellas
2Urologic clinic, Diagnostic & Therapeutic Center of Athens, "HYGEIA," Kiffisias Ave. & 4 Erythrou Stavrou Str., 151 23 Marousi, Athens, Hellas
3Surgical Clinic, General State Hospital of Athens "G. Gennimatas," Mesogeion Avenue, Athens, Hellas
4Centre of Radiation Oncology, Diagnostic & Therapeutic Center of Athens, "HYGEIA," Kiffisias Ave. & 4 Erythrou Stavrou Str., 151 23 Marousi, Athens, Hellas
5Department of Pathology, Medical School, University of Ioannina, Hellas
6Breast Center, Diagnostic & Therapeutic Center of Athens, "HYGEIA," Kiffisias Ave. & 4 Erythrou Stavrou Str., 151 23 Marousi, Athens, Hellas

Extracellular nucleic acids could serve as molecular markers in the early detection of cancer and in the prediction of disease outcome. In this study we examined six molecular markers, such as: variations in the quantity of DNA in plasma, glutathione-S-transferase P1 (GSTP1) gene methylation status in plasma, carcinoembryonic antigen (CEA) and prostate-specific membrane antigen (PSMA) mRNA in peripheral blood mononuclear cells (PBMC), and plasma samples from prostate cancer patients in different stages. The combination of DNA load and GSTP1 promoter methylation status identified 83% (10/12) of the prostate cancer patients before therapy. This study shows that free circulating DNA can be detected in patients with prostate cancer compared with disease-free individuals, and suggests a new, noninvasive approach for early detection of prostate cancer.

Keywords: Prostate-specific membrane antigen (PSMA), Glutathione-S-transferase P1 (GSTP1); Carcinoembryonic antigen (CEA); DNA quantification

Address correspondence to Dr. Georgios Nasioulas, Molecular Biology Research Center HYGEIA "Antonis Papayannis," Kiffisias Ave. & 4 Erythrou Stavrou Str., 151 23 Marousi, Athens, Hellas. E-mail: g.nasioul@hygeia.gr

Oncology Research, Volume 14, pp. 447-454
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Radiosensitization of Colon Cancer Cell Lines by Docetaxel: Mechanisms of Action

Amanda L. Dunne,1 Carmel Mothersill,2 Tracy Robson,1 George D. Wilson,3 and David G. Hirst1

1School of Biomedical Sciences, University of Ulster, Cromore Road, Coleraine, BT52 1AS, UK
2Radiation Science Centre, Dublin Institute of Technology, Dublin 1, 3Gray Cancer Institute, Northwood, Middx., HA6 2JR UK

The radiation-modifying action of docetaxel in experimental systems is well established. Docetaxel is also an increasingly important drug for the treatment of cancer in concurrent radiotherapy protocols. However, the mechanisms of docetaxel radiosensitization are not fully understood. We have investigated the magnitude and mechanisms of docetaxel radiosensitization in vitro in four human colorectal cancer cell lines (SW480, SW707, SW48, and HT29) with widely differing radiosensitivities. Cell survival curves were generated for a range of docetaxel concentrations (5-20 nM) alone and for X-rays (1-5 Gy) ± 10 or 20 nM docetaxel (for 24 h before irradiation). Cell cycle distributions and apoptotic frequencies were measured during the treatments. Sensitivity to docetaxel alone was similar in all cell lines and could be attributed to massive induction of apoptosis (60-80% by 24 h). Radiosensitivity varied widely; the surviving fractions at 2 Gy in the most resistant (HT29) and most sensitive (SW28) lines were 0.81 and 0.13, respectively. Exposure to 10 nM docetaxel induced a progressive accumulation of SW480, SW707, and SW48 cells in G2/M. After 24 h, 55-70% of the cells were in G2/M. It is likely, therefore, that accumulation in this radiosensitive phase of the cell cycle contributes significantly to radiosensitization by the drug.

Key words: Docetaxel; Radiosensitization; Apoptosis; G2/M block; Colon cancer

Address correspondence to Prof. David G. Hirst, School of Pharmacy, Medical Bilogy Centre, Queen's University Belfast, 97 Lisburn Road, Belfast BT9 7BL, UK. Tel: 028 9097 2027 ; Fax: 028 9024 7794; E-mail: D.Hirst@qub.ac.uk

Oncology Research, Volume 14, pp. 455-468
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

PEG Prodrugs of 6-Mercaptopurine for Parenteral Administration Using Benzyl Elimination of Thiols

Yun H. Choe, Richard B. Greenwald, Charles D. Conover, Hong Zhao, Clifford B. Longley, Shuiyun Guan, Qiuxia Zhao, and Jing Xia

Enzon Pharmaceuticals, Inc., 20 Kingsbridge Road, Piscataway, NJ 08854

6-Mercaptopurine (6-MP) is an orally administered, water-insoluble purine analog that is effective against acute lymphatic leukemia. Oral absorption of 6-MP, however, is quite erratic, with only 16-50% of the administered dose reaching the blood. In this report, water-soluble parenterally administered poly(ethylene glycol) (PEG) prodrugs of 6-MP were synthesized using several chemical approaches that enabled the protection of the thiol group through a modification of the benzyl elimination (BE) system. In our earlier work on antimetabolites, it was found that branching of the PEG allowed greater loading of the active drug. This approach was also utilized within this work to give multiloaded systems. The resulting conjugates were stable in pH 7.4 PBS buffer as well as in rat plasma for extended periods. However, these conjugates did act as prodrugs in vivo and a number of PEG-6-MP constructs had significant (P < 0.05) activity in murine leukemia, as well as certain solid tumors, compared with unconjugated 6-MP in a solubilizing vehicle. The fact that some PEG-6-MP conjugates were stable during in vitro plasma dissociation assays, but demonstrated in vivo anticancer activity, suggests extravascular cleavage of the linking group. This work demonstrates that PEG conjugation is an effective means of solubilizing 6-MP for parenteral administration.

Key words: 6-Mercaptopurine; Poly(ethylene glycol); Prodrug; Parenteral administration; Benzyl elimination

Address correspondence to Charles Conover, Ph.D., Department of Organic & Medicinal Chemistry, Enzon Pharmaceuticals, Inc., 20 Kingsbridge Road, Piscataway, NJ 08854. Tel: (732) 980-4958; Fax: (732) 885-2950; E-mail: charles.conover@enzon.com.