We previously demonstrated that increase of procathepsin L secretion by human melanoma cells strongly increased their tumourigenicity and switched their phenotype from low to highly metastatic. Thus, we herein analysed whether it was possible to inhibit procathepsin L secretion using anti-cathepsin L ScFv. For this purpose, we produced different forms of fusion cathepsin L in prokaryotic or eukaryotic expression systems. An anti-cathepsin L monoclonal antibody (mAb), named 3D8, was isolated from mice immunized with purified procathepsin L-His. This 3D8 mAb interacted with an epitope localized on the 156—197 amino acid sequence of cathepsin L and recognized recombinant or native forms of cathepsin L synthesized by human melanoma cells. An active anti-cathepsin L ScFv was generated and characterized from 3D8 mAb heavy and light variable chains. Then, human melanoma cells were transiently co-transfected with 3D8 ScFv and cathepsin L cDNAs. Data demonstrated that increase of 3D8 ScFv expression in human melanoma cells totally inhibited procathepsin L secretion and induced accumulation of intracellular procathepsin L. Our results constitute the first demonstration that anti-cathepsin L ScFv could be used in human melanoma cells to inhibit procathepsin L secretion. This ScFv represents a new molecular tool to explore cell therapy of human melanomas.
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October 2002
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Research Article|
October 01 2002
Cloning and characterization of anti-cathepsin L single chain variable fragment whose expression inhibits procathepsin L secretion in human melanoma cells Available to Purchase
Nathalie GUILLAUME-ROUSSELET;
Nathalie GUILLAUME-ROUSSELET
Immunochimie des Régulations Cellulaires et des Interactions Virales, INSERM U.354, Centre INSERM, Hôpital Saint-Antoine, 75012 Paris, France
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Didier JEAN;
Didier JEAN
Immunochimie des Régulations Cellulaires et des Interactions Virales, INSERM U.354, Centre INSERM, Hôpital Saint-Antoine, 75012 Paris, France
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Raymond FRADE
Raymond FRADE
1
Immunochimie des Régulations Cellulaires et des Interactions Virales, INSERM U.354, Centre INSERM, Hôpital Saint-Antoine, 75012 Paris, France
1To whom correspondence should be addressed, at INSERM U.354, Bâtiment G8, Génopole d'Evry, RN.7, 91 030 Evry, France (e-mail [email protected]).
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Publisher: Portland Press Ltd
Received:
March 01 2002
Revision Received:
June 12 2002
Accepted:
June 17 2002
Accepted Manuscript online:
September 24 2002
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2002
2002
Biochem J (2002) 367 (1): 219–227.
Article history
Received:
March 01 2002
Revision Received:
June 12 2002
Accepted:
June 17 2002
Accepted Manuscript online:
September 24 2002
Citation
Nathalie GUILLAUME-ROUSSELET, Didier JEAN, Raymond FRADE; Cloning and characterization of anti-cathepsin L single chain variable fragment whose expression inhibits procathepsin L secretion in human melanoma cells. Biochem J 1 October 2002; 367 (1): 219–227. doi: https://doi.org/10.1042/bj20020350
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