Human kallikrein 13 involvement in extracellular matrix degradation.
The human kallikrein family is a group of 15 serine protease genes clustered on chromosome 19q13.4 and shares a high degree of homology. These proteolytic enzymes have diverse physiological functions in many different tissues. Growing evidence suggests that many kallikreins are differentially expressed in cancer and may play a role in metastasis. Human kallikrein gene 13 (KLK13) is a member of this family and codes for a trypsin-like, secreted serine protease (hK13) that is overexpressed in ovarian cancer patients. The aim of this study was to determine if hK13 can degrade extracellular matrix components. Recombinant hK13 was produced in yeast and purified using cation exchange and reverse-phase chromatography. The protein was used as an immunogen to generate mouse monoclonal antibodies. Enzymatic activity of hK13 was verified by using synthetic tri-peptide fluorogenic substrates and gelatin zymography. Active hK13 was incubated with biotinylated extracellular matrix (ECM) proteins and degradation was evaluated by Western blot analysis. hK13-secreting cancer cell lines were treated in a chemotaxis invasion chamber that was coated with various ECM proteins, to determine if hK13 plays a role in tumor cell migration and invasion. Assay with the synthetic substrates and zymography have shown that recombinant hK13 was enzymatically active. The Western blot results showed that hK13 was able to cleave the major components of the extracellular matrix. In the chemotaxis invasion chamber experiment, it was found that ovarian cancer cell lines that secreted hK13 and were treated with an hK13 neutralizing antibody migrated less than untreated cells. Human kallikrein13 may play a role in tissue remodeling and/or tumor invasion and metastasis. Targeting hK13 activity with neutralizing antibodies may have therapeutic applications.
Kapadia
Carl
C
Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada M5G 1X5.
Ghosh
Manik C
MC
Grass
Linda
L
Diamandis
Eleftherios P
EP
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D005109:Extracellular Matrix / Q000737:chemistry*; D016326:Extracellular Matrix Proteins / Q000737:chemistry*; D005260:Female; D006801:Humans; D007610:Kallikreins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D009369:Neoplasms / Q000201:enzymology* / Q000276:immunology*
D000911:Antibodies, Monoclonal; D016326:Extracellular Matrix Proteins; C470778:KLK13 protein, human; D007610:Kallikreins