MMP-9 activity in urine from patients with various tumors, as measured by a novel MMP activity assay using modified urokinase as a substrate.
Matrix metalloproteinases (MMPs) play an important role in many pathologic processes, but their activities are difficult to determine since no simple specific and/or chromogenic substrate exists. We have developed a novel MMP activity assay using a modified urokinase as a substrate. Protein engineering enabled the plasmin activation site in this urokinase to be substituted by a specific activation site recognized by MMPs. In this way the MMP activity can be monitored via urokinase activity as measured by a simple chromogenic assay. The assay was made specific for MMP-9 by a capture step using MMP-9-specific antibodies that do not interfere with MMP-activity. This assay monitors the amount of active enzyme as well as the latent, but potentially active proform. Using this assay the levels of MMP-9 were investigated in urine from patients with various kinds of carcinoma. High levels of both active and latent MMP-9 were detected in urine from patients with carcinoma of the bladder, whereas hardly any activity was observed in urine from healthy controls. MMP-9 in urine was present in its intact form. Surprisingly, MMP-9 was also increased in the urine of patients with nonurogenital carcinoma. Therefore, measurement of urinary MMP-9 activity levels may be a convenient diagnostic tool for various types of carcinoma.
Gaubius Laboratory, TNO-PG, Leiden, The Netherlands. R.Hanemaaijer@PG.TNO.NL
R
R
Hanemaaijer
C F
CF
Sier
H
H
Visser
L
L
Scholte
N
N
van Lent
K
K
Toet
K
K
Hoekman
J H
JH
Verheijen
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000652:urine*; D001943:Breast Neoplasms / Q000652:urine; D017364:Collagenases / Q000652:urine*; D004938:Esophageal Neoplasms / Q000652:urine; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000652:urine; D008297:Male; D020780:Matrix Metalloproteinase 9; D008875:Middle Aged; D009369:Neoplasms / Q000201:enzymology / Q000628:therapy / Q000652:urine*; D010051:Ovarian Neoplasms / Q000652:urine; D012016:Reference Values; D013379:Substrate Specificity; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D014408:Biomarkers, Tumor; D014568:Urokinase-Type Plasminogen Activator; D017364:Collagenases; D020780:Matrix Metalloproteinase 9