Papers - Fujii, Hideki

Antimetastatic activities of synthetic Arg-Gly-Asp-Ser (RGDS) and Arg-Leu-Asp-Ser (RLDS) peptide analogues and their inhibitory mechanisms.

H Fujii, H Komazawa, H Mori, M Kojima, I Itoh, J Murata, I Azuma, I Saiki

Biological & pharmaceutical bulletin 18 ( 12 ) 1681 - 8 1995.12

Research paper (scientific journal), Joint Work, Lead author, Last author, Corresponding author, Accepted,  ISSN  0918-6158

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We have investigated the inhibitory effect of the N-terminal modified Arg-Gly-Asp-Ser (RGDS) analogues, AcDRGDS and AcDRLDS, on tumor cell adhesion to the components of extracellular matrix and basement membrane, and also tested the antimetastatic effect of their conjugates with trimesic acid, Ar(DRGDS)3 and Ar(DRLDS)3. AcDRGDS significantly inhibited tumor cell adhesion to fibronectin, vitronectin and RGDS substrates, but not to CS1 substrate which is a ligand for the alpha 4 beta 1 tumor surface integrin receptor. In contrast, AcDRLDS variant peptide significantly inhibited tumor cell adhesion to laminin, in addition to RGDS-mediated adhesion to fibronectin and vitronectin. AcDRLDS also inhibited tumor cell adhesion to CS1 as well as the RGDS sequence within the fibronectin molecule in a concentration-dependent manner, although the inhibitory effect was less than that of the CS1 (EILDV) peptide. Ar(DRLDS)3 inhibited the laminin- and fibronectin-mediated invasion and migration of tumor cells, whereas Ar(DRGDS)3 selectively inhibited fibronectin-mediated invasion and migration. Ar(DRGDS)3 and Ar(DRLDS)3 were much more effective in inhibiting experimental lung or liver metastases of various types of murine and human tumors than the original RGDS-containing peptides or Ar(COONa)3. Multiple administrations of Ar(DRGDS)3 or Ar(DRLDS)3 potently inhibited spontaneous lung metastasis produced by intra-footpad injection of B16-BL6 cells without affecting the primary tumor size at the time of surgical excision, as compared with RGDS peptide or untreated control. Thus, Ar(DRGDS)3 and Ar(DRLDS)3 substantially increased the exhibiting any antimetastatic effect of the peptides without direct cytotoxicity.

Role of the heparin-binding domain of chimeric peptides derived from fibronectin in cell spreading and motility.

J Yoneda, I Saiki, Y Igarashi, H Kobayashi, H Fujii, Y Ishizaki, F Kimizuka, I Kato, I Azuma

Experimental cell research 217 ( 1 ) 169 - 79 1995.03

Research paper (scientific journal), Joint Work, Lead author, Last author, Corresponding author, Accepted,  ISSN  0014-4827

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Cellular responses to fibronectin (FN) are likely to have a complex molecular basis involving the interactions between multiple functional domains of FN and specific cell surface molecules. We have utilized several types of recombinant FN fragments and their chimeric fragments to examine the regulatory mechanisms of the spreading and chemotactic migration of HT1080 human fibrosarcoma cells on FN. The CH-271 fusion fragment, in which the cell-binding domain (C-274) of FN is adjacent to the heparin-binding domain (H-271), promoted cell spreading more efficiently than C-274, H-271, or their mixture (C-274 + H-271) over a 60-min incubation. The CH-271 variants containing various modules in the heparin-binding domain (CHV peptide) showed different promotion of cell migration, spreading, and vinculin accumulation at focal adhesion, respectively. The preincubation of the cells with heparitinase I resulted in significant inhibition of chemotactic migration to FN and its fragments containing the III13 and/or III14 modules of the heparin-binding domain. Additionally, migration to CH-271 was inhibited by the presence of the RGDS peptide in a concentration-dependent fashion. Thus, the spread and migration responses of HT1080 cells onto FN fusion peptides require the adjacent coexistence of cell- and heparin-binding domains and are mediated by the interactions between cell surface heparan sulfate and the heparin-binding domain, in concert with the interaction between cell surface integrin and the cell-binding domain. In conclusion, our present study demonstrated that fusion peptides of fibronectin can efficiently induce two signals from the cell-binding and heparin-binding domains required for the completion of cell spreading, the formation of focal contact, and motility at the early stage of the culture, suggesting that the III13 or III14 modules within the heparin-binding domain are responsible for the initiation of different cellular responses.

Combination of anti-cell adhesive synthetic Arg-Gly-Asp-Ser analogue and anticancer drug doxorubicin heightens their original antimetastatic activities.

H Komazawa, H Fujii, M Kojima, H Mori, M Ono, I Itoh, I Azuma, I Saiki

Oncology research 7 ( 7-8 ) 341 - 51 1995

Research paper (scientific journal), Joint Work, Lead author, Last author, Corresponding author, Accepted,  ISSN  0965-0407

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A new compound containing the cell-adhesive Arg-Gly-Asp-Ser (RGDS) peptide was synthesized, i.e. tetrahydrofurantetracarboxylic acid (THFTCA)-RGDS conjugate [THFTCA- (RGDS)3, FC-243], and the inhibitory effect of FC-243 on lung metastasis of B16-BL6 melanoma in mice was examined in combination with or without the anticancer agent doxorubicin (DOX). FC-243 showed an inhibitory effect on lung metastasis of melanoma cells in a dose-dependent manner. A mixture of THFTCA and RGDS peptide or THFTCA alone did not show any inhibitory effect on experimental lung metastasis as compared with FC-243 on a molar basis. RGDS peptide, however, required a higher dose to obtain a sufficient antimetastatic effect. Intermittent IV administration of FC-243 after the inoculation of B16-BL6 cells caused significant inhibition of spontaneous lung metastasis as compared with multiple administration of RGDS or untreated control. The in vitro tumor invasion study showed that FC-243 as well as RGDS+THFTCA on a molar basis resulted in similar inhibition of the invasion of B16-BL6 cells into reconstituted basement membrane Matrigel. Combined treatment with FC-243 and DOX significantly inhibited lung metastasis of melanoma as compared with either treatment alone or the untreated control. Administrations of FC-243 and DOX in combination substantially prolonged the survival time of mice. These results demonstrate that combination therapy of the anti-cell adhesive FC-243 and the anticancer agent DOX, i.e. antiadhesion therapy and chemotherapy, is a new approach that offers enhanced inhibitory effects on tumor metastasis and invasion.

Inhibitory effect of recombinant fibronectin polypeptides on the adhesion of liver-metastatic lymphoma cells to hepatic sinusoidal endothelial cells and tumor invasion.

J Yoneda, I Saiki, H Kobayashi, H Fujii, Y Ishizaki, I Kato, M Kiso, A Hasegawa, I Azuma

Japanese journal of cancer research : Gann 85 ( 7 ) 723 - 34 1994.07

Research paper (scientific journal), Joint Work, Lead author, Last author, Corresponding author, Accepted,  ISSN  0910-5050

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We have investigated the inhibitory mechanism of the initial arrest of L5178Y-ML25 lymphoma cells in a target organ (liver) by using recombinant fibronectin fragments with cell- and/or heparin-binding domains (C-274, H-271 or the fusion fragment CH-271). Pretreatment of hepatic sinusoidal endothelial (HSE) cell monolayers with lymphoma cells or their conditioned medium for 4 to 6 h resulted in the enhancement of lymphoma cell adhesion to HSE cell monolayer. The increased tumor adhesiveness was completely abolished by preincubation of the conditioned medium with anti interleukin-1 beta monoclonal antibody (mAb). Synthetic sialyl Le(x) (SLe(x)) as a ligand for endothelial cell leukocyte adhesion molecule-1 (ELAM-1) adhesion receptor and anti ELAM-1 mAb blocked the conditioned medium-induced enhancement of tumor-endothelial cell interaction, while pretreatment of the activated HSE cell monolayer with anti vascular cell adhesion molecule-1 (VCAM-1) mAb did not affect the enhanced tumor cell adhesion. These results indicate that tumor cell interaction with the stimulated HSE cells is mediated by ELAM-1 molecules on HSE cells. However, the expression of SLe(x) and SLe(a) on the tumor surface was not observed by flow cytometric analysis. ELAM-1-mediated enhancement of tumor cell adhesion to HSE monolayer was also inhibited in a concentration-dependent manner by CH-271 fusion polypeptide or the sulfated chitin derivative sulfated carboxymethyl-chitin, which can bind to the heparin-binding domain of CH-271. In addition, CH-271 inhibited not only tumor-endothelium interaction but also tumor cell invasion into reconstituted basement membrane Matrigel in vitro.

Inhibition of tumor cell invasion by ubenimex (bestatin) in vitro.

Saiki I, Murata J, Watanabe K, Fujii H, Abe F, Azuma I

Jpn. J. Cancer Res. 80 ( 9 ) 873 - 878 1989.09

Research paper (scientific journal), Joint Work, Lead author, Last author, Corresponding author, Accepted

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