A new assay for measurement of the liberated domain I of the urokinase receptor in plasma improves the prediction of survival in colorectal cancer.
BACKGROUND
The liberated domain I of the urokinase plasminogen activator receptor [uPAR(I)] is a significant prognostic marker in lung and ovarian cancer, although the uPAR(I) concentration is below the limit of quantification (LOQ) in a substantial proportion of patient samples (Lung Cancer 2005;48:349-55; Clin Cancer Res 2008;14:5785-93; APMIS 2009;117:755-61). This study was undertaken to design an immunoassay with improved functional sensitivity for measuring uPAR(I) and to evaluate the prognostic value of uPAR(I) for colorectal cancer (CRC) patients.


METHODS
Surface plasmon resonance analysis identified 2 monoclonal antibodies, R3 and R20, that simultaneously bind to the liberated uPAR(I) but not to intact uPAR. We used R3 for capture and Eu-labeled R20 for detection in designing a 2-site sandwich time-resolved fluorescence immunoassay (TR-FIA 4) for measuring liberated uPAR(I). TR-FIA 4 was validated for use with citrated plasma. The prognostic value of the uPAR(I) concentration was evaluated in 298 CRC patients. The Cox proportional hazards model was used for the uni- and multivariate survival analyses.


RESULTS
The LOQ was 0.65 pmol/L. Liberated uPAR(I) was measurable in all patient samples with TR-FIA 4. In the multivariate analysis that included sex, age, tumor stage, tumor localization, and adjuvant treatment, the uPAR(I) concentration measured with TR-FIA 4 (hazard ratio, 1.72; 95% CI, 1.15-2.57; P = 0.009), as well as the concentration of intact soluble uPAR plus the cleaved uPAR fragment containing domains II and III, tumor stage, and age were independent predictors of prognosis.


CONCLUSIONS
TR-FIA 4 has a functional sensitivity improved 4-fold over that of the previous uPAR(I) assay. The uPAR(I) concentration measured with TR-FIA 4 is an independent predictor of prognosis in CRC patients.
Thurison
Tine
T
Finsen Laboratory, Copenhagen University Hospital, Copenhagen, Denmark.
Lomholt
Anne F
AF
Rasch
Morten G
MG
Lund
Ida K
IK
Nielsen
Hans J
HJ
Christensen
Ib J
IJ
Høyer-Hansen
Gunilla
G
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000097:blood*; D015179:Colorectal Neoplasms / Q000097:blood / Q000175:diagnosis* / Q000401:mortality; D005260:Female; D006801:Humans; D007118:Immunoassay; D053208:Kaplan-Meier Estimate; D008297:Male; D008875:Middle Aged; D011379:Prognosis; D017434:Protein Structure, Tertiary; D055293:Receptors, Urokinase Plasminogen Activator / Q000097:blood*; D012680:Sensitivity and Specificity
D014408:Biomarkers, Tumor; D055293:Receptors, Urokinase Plasminogen Activator