ognizant Communication Corporation



Oncology Research, Volume 14, pp. 469-474
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

The Ability of New Sugar-Modified Derivatives of Antitumor Anthracycline, Daunorubicin, to Stimulate NAD(P)H Oxidation in Different Cellular Oxidoreductase Systems: NADH Dehydrogenase, NADPH Cytochrome P450 Reductase, and Xanthine Oxidase

Jolanta Pawlowska,1 Waldemar Priebe,2 Mark J. I. Paine,3 C. Roland Wolf,3 Edward Borowski,1 and Jolanta Tarasiuk1,4

1Department of Pharmaceutical Technology and Biochemistry, Technical University of Gda?sk, Narutowicza St 11/12, 80-952 Gda?sk, Poland
2M.D. Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030
3Cancer Research UK Molecular Pharmacology Unit, Biomedical Research Centre, Ninewells Hospital and Medical School, Dundee, UK
4Department of Biochemistry, University of Szczecin, Szczecin, Poland

Numerous data indicate that cellular oxidoreductases may be responsible for the cardiotoxic effects of antitumor anthracycline drugs as a consequence of the mediation by these agents of one-electron transfer from reduced nucleotides to atmospheric oxygen. This process is catalyzed primarily by NADH dehydrogenase, NADPH cytochrome P450 reductase, and xanthine oxidase and leads to the formation of reactive oxygen species (ROS). In this work the data on the ability of new amino sugar derivatives of daunorubicin to stimulate NAD(P)H oxidation in the above oxidoreductase systems are presented. They represent analogues of daunorubicin in which the amino sugar nitrogen is bounded to an unsubsituted, or amino- or nitro-substituted benzyl group. It was found that the ability of examined sugar-modified derivatives of daunorubicin to stimulate NAD(P)H oxidation differs considerably depending on the subsituent in the phenyl ring. It was also determined that this ability was not identical in the three enzymatic systems studied, showing that these derivatives have different affinities for the enzymes examined. More similarities were observed in their interaction with NADH dehydrogenase and NADPH cytochrome P450 reductase than with xanthine oxidase.

Key words: Daunorubicin; Sugar-modified derivatives; Reactive oxygen species formation; NAD(P)H oxidation; NADH dehydrogenase; NADPH cytochrome P450 reductase; Xanthine oxidase

Address correspondence to Jolanta Tarasiuk, Department of Biochemistry, University of Szczecin, 3a Felczaka St, 71-412 Szczecin, Poland. Tel/fax: (48) (91) 444-15-50; E-mail: tarasiuk@univ.szczecin.pl

Oncology Research, Volume 14, pp. 475-482
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Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

PAR1-Type Thrombin Receptor Stimulates Migration and Matrix Adhesion of Human Colon Carcinoma Cells by a PKCe-Dependent Mechanism

Ilonka Heider,1 Beate Schulze,1 Elke Oswald,1 Peter Henklein,2 Johannes Scheele,1 and Roland Kaufmann1

1Department of General and Visceral Surgery, Medical Faculty at the Friedrich Schiller University Jena, Erlanger Allee 101, D-07747, Germany
2Institute of Biochemistry, Charité, Humboldt University of Berlin, Monbijoustr. 2, D-10117 Berlin, Germany

The proteinase-activated receptor1 (PAR1) was characterized as a functional receptor for thrombin in cells from different tumor entities. In colon carcinoma, its function has to be defined. In this study we demonstrate that the PAR1-selective agonist peptide TFLLRN induced activation of protein kinase C isoenzymes a and e in human HT-29 colon carcinoma cells expressing PAR1 endogeneously. On the cellular level, TFLLRN and thrombin prompted HT-29 cell migration and matrix adhesion by a PKCe-dependent mechanism as concluded because of the inhibition of PAR1-mediated effects by the PKC inhibitor bisindolylmaleimide I and the PKCe translocation inhibitory peptide EAVSLKPT but not by the PKC inhibitor Gö 6976. In addition, blockade of PAR1 by RWJ 56110, a selective PAR1 antagonist, fully abolished the effect of thrombin on HT-29 cell migration and adhesion. Therefore, PAR1 seems to be the responsible receptor for thrombin-induced migration and adhesion of human colon carcinoma cells including PKCe as an essential signal transducer.

Key words: Proteinase-activated receptors; PAR1; Colon carcinoma; Protein kinase C

Address correspondence to Dr. Roland Kaufmann, Research Lab., Department of General and Visceral Surgery, Research Center Lobeda, Medical Faculty at the Friedrich Schiller University Jena, Erlanger Allee 101, 07747 Jena, Germany. Tel: +49-9325870; Fax: +49-9325872; E-mail: roland.kaufmann@med.uni-jena.de

Oncology Research, Volume 14, pp. 483-489
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Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Nicotinamide Relaxes Vascular Smooth Muscle by Inhibiting Myosin Light Chain Kinase-Dependent Signaling Pathways: Implications for Anticancer Efficacy

Mark W. Ruddock and David G. Hirst

Radiation Science Group, School of Biomedical Sciences, University of Ulster, Cromore Road, Coleraine, County Londonderry, Northern Ireland, BT52 1SA

Nicotinamide has been shown to be an effective tumor oxygenator in preclinical studies and is part of a promising clinical protocol for the treatment of cancer of the larynx. It has been known for some time that nicotinamide sensitizes tumors, at least in part, by modulating vascular smooth muscle contraction; however, the cellular target within the smooth muscle cell has yet to be identified. Our previous studies have eliminated targets within several agonist and antagonist signaling pathways in smooth muscle, suggesting that it must act at a point close to the contractile machinery of the cell. The present study investigated the effect of nicotinamide on four key steps responsible for force generation via actin/myosin interaction in the smooth muscle cell: calcium binding to calmodulin, calcium-calmodulin binding to smooth muscle myosin light chain kinase (MLCK) inhibitor peptide 480-501 (smMLCIP), modulation of MLCK-dependent signaling, and MLCK-induced phosphorylation of the regulatory myosin light chain, MLC20. Nicotinamide abolished the phosphorylation of MLC20, but had no significant effect on the other endpoints tested. We conclude that the vasorelaxant effects of nicotinamide are mediated mainly through inhibition of MLC20 phosphorylation, and that this could be a promising target for the development of novel tumor oxygenators to enhance radio- and chemotherapy.

Key words: Nicotinamide; Myosin light chain kinase; Myosin light chains; Tumor radiosensitizer; Vascular smooth muscle

Address correspondence to David G. Hirst at his present address: School of Pharmacy, Medical Biology 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. 491-499
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Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

The Differential Proteome Profile of Stomach Cancer: Identification of the Biomarker Candidates

Joung S. Jang, Hee Y. Cho, Young J. Lee, Woo S. Ha, and Hwal W. Kim

Department of Internal Medicine, Department of Surgery, Department of Pathology, GyeongSang National University College of Medicine, and GyeongSang Institute of Health Sciences, Jinju, South Korea

By comparative proteome analysis we searched for characteristic alterations of human stomach adenocarcinoma tissue and paired surrounding normal tissue. Selected differential protein spots were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) and database searching. We identified protein alterations in 18 stomach cancer tissues compared with normal controls, comprising elevated levels of eight proteins, including 14-3-3 zeta, calcyclin, keratin, apolipoprotein A-1 precursor, proteasome activator complex subunit, nucleoside diphosphate kinase, nicotinamide N-methyltransferase, and pyridoxal kinase. Five proteins (CA11, prohibitin, peroxiredoxin 4, serum amyloid P component, and NADH-ubiquinone oxidoreductase 23 kDa subunit) were decreased. These data are valuable for identification of differentially expressed proteins involved in stomach cancer carcinogenesis, providing biomarker candidates to develop diagnostic and therapeutic tools.

Key words: Stomach cancer; Proteome; Biomarker

Address correspondence to Joung Soon Jang, M.D., Department of Internal Medicine, GyeongSang National University Hospital, 90 Chiram-dong Jinju, 660-702 South Korea. Tel: +82-55-750-8054; Fax: +82-55-758-9122; E-mail: jsjang@nongae.gsnu.ac.kr

Oncology Research, Volume 14, pp. 501-511
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Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

Proteolysis of Human Plasma Reveals the Presence of Complexes Formed by Endogenous Heparin and Peptides That Stimulate Angiogenesis

Stefania Pacini,1 Marco Ruggiero,2 Fabiola Cecchi,2 Benedetta Peruzzi,1 and Simonetta Vannucchi2

1Department of Anatomy, Histology and Forensic Medicine, and 2Department of Experimental Pathology and Oncology, University of Firenze, Italy

Human endogenous plasma heparin associates with proteins that mask its anticoagulant activity. This association persists after exhaustive proteolysis of plasma, and resulting peptide/heparin complexes have no anticoagulant activity. Looking for functions other than inhibition of coagulation, we considered that commercial preparations of heparin from bovine or porcine sources show alternative effects on angiogenesis, either stimulating or inhibiting the process. However, the effects of endogenous human heparin on angiogenesis are unknown. In this study, the fraction of plasma containing endogenous heparin was prepared by means of exhaustive proteolysis, either in the presence or in the absence of 35S-labeled heparin. Plasma from healthy donors was digested and the supernatant was precipitated with 66% ethanol, dialyzed, and submitted to basic and acidic ion-exchange chromatography. 35S-Labeled heparin as well as endogenous heparin bound plasmatic peptides, forming acidic, basic, and neutral complexes. Binding of peptides, eluting from both resins, impaired migration of heparin on cellulose acetate electrophoresis. Endogenous neutral complexes (i.e., those formed by human endogenous plasma heparin and peptides) were tested for angiogenic activity in chick embryo chorioallantoic membrane assay. Bovine heparin induced a moderate angiogenic response. Neutral complexes of human endogenous plasma heparin and basic plasma peptides induced a very strong angiogenic response. Treatment of neutral complexes with nitrous acid, which degrades heparin, abolished the angiogenic effect, thus demonstrating that it was due to the presence of heparin. These results demonstrate that proteolysis of human plasma generates angiogenic peptide/heparin complexes.

Key words: Glycosaminoglycans; Peptides; Heparin; Human plasma; Proteolysis; Angiogenesis

Address correspondence to Prof. Simonetta Vannucchi, Dipartimento di Patologia e Oncologia Sperimentali, viale Morgagni 50, I-50134 Firenze, Italy. Fax: +39 055 4282333; E-mail: svannucchi@unifi.it

Oncology Research, Volume 14, pp. 513-527
0965-0407/04 $20.00 + .00
Copyright © 2004 Cognizant Comm. Corp.
Printed in the USA. All rights reserved.

In Vivo Molecular Pharmacology and Antitumor Activity of the Targeted Akt Inhibitor PX-316

Emmanuelle J. Meuillet,1 Nathan Ihle,1 Amanda F. Baker,1 Jaime M. Gard,1 Chelsea Stamper,1 Ryan Williams,1 Amy Coon,1 Daruka Mahadevan,1 Benjamin L. George,1 Lynn Kirkpatrick,2 and Garth Powis1

1Arizona Cancer Center, University of Arizona, Tucson, AZ 85724
2ProlX Pharmaceuticals, Tucson, AZ 85701

Akt, a serine/threonine kinase that promotes cell survival, is activated by binding of its pleckstrin homology (PH) domain to membrane phosphatidylinositol (PtdIns)-3-phosphates formed by PtdIns-3-kinase. D-3-Deoxy-phosphatidyl-myo-inositols that cannot be phosphorylated on the 3-position of the myo-inositol group are inhibitors of the Akt PH domain. The most active compound is D-3-deoxy-phosphatidyl-myo-inositol 1-[(R)-2-methoxy-3-octadecyloxypropyl hydrogen phosphate] (PX-316). PX-316 administered intraperitoneally to mice at 150 mg/kg inhibits Akt activation in HT-29 human tumor xenografts up to 78% at 10 h with recovery to 34% at 48 h. Phosphorylation of GSK-3b, a downstream target of Akt, is also inhibited. There is no decrease in PtdIns(3,4,5)-trisphosphate levels by PX-316, showing it is not an inhibitor of PtdIns-3-K in vivo. Gene expression profiling of HT-29 tumor xenografts shows many similarities between the effects of PX-316 and the PtdIns-3-K inhibitor wortmannin, with downregulation of several ribosomal-related genes, while PX-316 uniquely increases the expression of a group of mitochondrial-related genes. PX-316 has antitumor activity against early human MCF-7 breast cancer and HT-29 colon cancer xenografts in mice. PX-316 formulated in 20% hydroxypropyl-b-cyclodextrin for intravenous administration is well tolerated in mice and rats with no hemolysis and no hematological toxicity. Thus, PX-316 is the lead compound of a new class of potential agents that inhibit Akt survival signaling.

Keywords: PX-316; D-3-Deoxy-phosphatidyl-myo-inositol ether lipid; Akt; Antitumor activity; Pharmacodynamics

Address correspondence to Dr. Garth Powis, Arizona Cancer Center, University of Arizona, 1515 N Campbell Avenue, Tucson AZ 85724 5024. Tel: (520) 626 6408; Fax: (520) 626 4848; E-mail: gpowis@azcc.arizona.edu