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Clinical Chemistry 51: 1432-1442, 2005. First published June 16, 2005; 10.1373/clinchem.2005.049692
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(Clinical Chemistry. 2005;51:1432-1442.)
© 2005 American Association for Clinical Chemistry, Inc.

Proteomics and Protein Markers

Human Kallikrein 4: Quantitative Study in Tissues and Evidence for Its Secretion into Biological Fluids

Christina V. Obiezu1,2, Shannon J.C. Shan1,2, Antoninus Soosaipillai1, Liu-Ying Luo1,2, Linda Grass1, Georgia Sotiropoulou3, Constantina D. Petraki4, Panos A. Papanastasiou4, Michael A. Levesque1 and Eleftherios P. Diamandis1,2,a

1 Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada.
2 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.
3 Department of Pharmacy, School of Health Sciences, University of Patras, Rion, Greece.
4 Departments of Pathology and Urology, Evangelismos Hospital, Athens, Greece.

aAddress correspondence to this author at: Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, 600 University Ave., Toronto, Ontario, Canada M5G 1X5. Fax 416-586-8628; e-mail ediamandis{at}mtsinai.on.ca.

Background: Human kallikrein 4 (hK4) is a proteolytic enzyme belonging to the tissue kallikrein family of serine proteases. Previous tissue expression studies have demonstrated highest KLK4 mRNA expression in prostatic tissue, but there has been only limited evidence for the presence of hK4 protein in prostate and other tissues and in corresponding biological secretions.

Methods: To investigate the concentrations of hK4 in tissues and biological fluids, we developed a new hK4-specific sandwich-type immunoassay using a monoclonal antibody as the capture reagent.

Results: The assay has a detection limit of 0.02 µg/L and <0.1% cross-reactivity toward any of the other 14 human kallikreins. Twelve of 40 tissue extracts prepared from various human tissues contained detectable hK4 concentrations (0.68–7143 ng/g of total protein), with healthy prostate tissue containing the highest amount of hK4. Examination of 16 malignant and 18 benign prostate tissues revealed no significant differences in hK4 protein content, and the tissues contained a wide range of values (benign, <0.02 to 801 ng/g; malignant, <0.02 to 824 ng/g). Among the biological fluids tested, seminal plasma and urine contained widely varying amounts of hK4; concentrations in 54 urine samples were <0.02 to 2.6 µg/L, whereas concentrations in 58 seminal plasma samples were 0.2–202 µg/L. Affinity purification of hK4 from seminal plasma and subsequent mass spectrometry demonstrated the secreted nature of hK4 in seminal plasma.

Conclusions: hK4 is found primarily in prostate tissue and is secreted in seminal plasma. Its value as a novel prostatic biomarker needs to be defined further.




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