title: Cathepsin D specifically cleaves the chemokines macrophage inflammatory protein-1 alpha, macrophage inflammatory protein-1 beta, and SLC that are expressed in human breast cancer.
abstract: Cathepsin D (Cath-D) expression in human primary breast cancer has been associated with a poor prognosis. In search of a better understanding of the Cath-D substrates possibly involved in cancer invasiveness and metastasis, we investigated the potential interactions between this protease and chemokines. Here we report that purified Cath-D, as well as culture supernatants from the human breast carcinoma cell lines MCF-7 and T47D, selectively degrade macrophage inflammatory protein (MIP)-1 alpha (CCL3), MIP-1 beta (CCL4), and SLC (CCL21). Proteolysis was totally blocked by the protease inhibitor pepstatin A, and specificity of Cath-D cleavage was demonstrated using a large chemokine panel. Whereas MIP-1 alpha and MIP-1 beta degradation was rapid and complete, cleavage of SLC was slow and not complete. Mass spectrometry analysis showed that Cath-D cleaves the Leu(58) to Trp(59) bond of SLC producing two functionally inactive fragments. Analysis of Cath-D proteolysis of a series of monocyte chemoattractant protein-3/MIP-1 beta hybrids indicated that processing of MIP-1 beta might start by cleaving off amino acids located in the C-terminal domain. In situ hybridization studies revealed MIP-1 alpha, MIP-1 beta, and Cath-D gene expression mainly in the stromal compartment of breast cancers whereas SLC transcripts were found in endothelial cells of capillaries and venules within the neoplastic tissues. Cath-D production in the breast carcinoma cell lines MCF-7 and T47D, as assessed by enzyme-linked immunosorbent assay of culture supernatants and cell lysates, was not affected by stimulation with chemokines such as interleukin-8 (CXCL8), SDF-1 (CXCL12), and SLC. These data suggest that inactivation of chemokines by Cath-D possibly influences regulatory mechanisms in the tumoral extracellular microenvironment that in turn may affect the generation of the antitumoral immune response, the migration of cancer cells, or both processes.
authors.affiliation: Theodor-Kocher Institute, University of Bern, Bern, Switzerland.
authors.forename: Marlene
authors.initials: M
authors.lastname: Wolf
authors.forename: Ian
authors.initials: I
authors.lastname: Clark-Lewis
authors.forename: Caroline
authors.initials: C
authors.lastname: Buri
authors.forename: Hanno
authors.initials: H
authors.lastname: Langen
authors.forename: Maddalena
authors.initials: M
authors.lastname: Lis
authors.forename: Luca
authors.initials: L
authors.lastname: Mazzucchelli
mesh_terms: D000595:Amino Acid Sequence; D001943:Breast Neoplasms / Q000201:enzymology / Q000235:genetics / Q000276:immunology / Q000473:pathology*; D002402:Cathepsin D / Q000235:genetics / Q000378:metabolism*; D054421:Chemokine CCL21; D054405:Chemokine CCL3; D054407:Chemokine CCL4; D019742:Chemokines, CC / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic / Q000276:immunology; D006801:Humans; D007700:Kinetics; D019402:Macrophage Inflammatory Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
chemical_list: C515478:CCL21 protein, human; D054421:Chemokine CCL21; D054405:Chemokine CCL3; D054407:Chemokine CCL4; D019742:Chemokines, CC; D019402:Macrophage Inflammatory Proteins; D002402:Cathepsin D