Early diagnosis of bladder cancer through the detection of urinary tyrosine-phosphorylated proteins

Abstract

Background: A noninvasive, highly sensitive and specific urine test is needed for bladder cancer (BC) diagnosis and surveillance in addition to the invasive cystoscopy. We previously described the diagnostic effectiveness of urinary tyrosine-phosphorylated proteins (UPY) and a new assay (UPY-A) for their measurement in a pilot study. The aim of this work was to evaluate the performances of the UPY-A using an independent cohort of 262 subjects.

Methods: Urinary tyrosine-phosphorylated proteins were measured by UPY-A test. The area under ROC curve, cutoff, sensitivity, specificity and predictive values of UPY-A were determined. The association of UPY levels with tumour staging, grading, recurrence and progression risk was analysed by Kruskal-Wallis and Wilcoxon's test. To test the probability to be a case if positive at the UPY-A, a logistic test adjusted for possible confounding factor was used.

Results: Results showed a significant difference of UPY levels between patients with BC vs healthy controls. For the best cutoff value, 261.26 Standard Units (SU), the sensitivity of the assay was 80.43% and the specificity was 78.82%. A statistically significant difference was found in the levels of UPY at different BC stages and grades between Ta and T1 and with different risk of recurrence and progression. A statistically significant increased risk for BC at UPY-A ⩾261.26 SU was observed.

Conclusions: The present study supplies important information on the diagnostic characteristics of UPY-A revealing remarkable performances for early stages and allowing its potential use for different applications encompassing the screening of high-risk subjects, primary diagnosis and posttreatment surveillance.

Figure 1

Urinary tyrosine-phosphorylated protein levels in urine samples. (A) Analysis of urinary UPY levels in samples of healthy subjects (n=170) and BC patients (n=92) using the UPY-A. Healthy subject (control (CTRL)) mean levels: 157.9±114.5 SU; BC mean levels: 434.8±258.4 SU. Significance of the differences: P=1.71 × 10⁻²³ by Wilcoxon's rank-sum test. The solid lines indicate the mean values; the dotted line indicates the best cutoff value. (B) ROC curve of total UPY levels adjusted for age, smoking status and gender. (C) Estimated risks of BC: results of the regression model including UPY-A, age, smoking status and gender as predictors.

Figure 2

Urinary tyrosine-phosphorylated protein levels in subjects with different BC stages and grades. (A) Distribution of UPY levels in subjects with different BC stages. Control (CTRL), n=170, mean 157.9±114.5 SU; carcinoma in situ (CIS), n=4, mean 400.8±114.8 SU; Ta, n=53, mean 356.3±181.0 SU; T1, n=20, mean 540.9±346.6 SU; and T2–3, n=15, mean 579.5±290.6 SU. Urinary tyrosine-phosphorylated protein levels are significantly different in groups of BC patients with different BC stages (Kruskal–Wallis test, P=8.10 × 10⁻²²). (B) Distribution of UPY levels in subjects with different BC grades. Control, n=170, mean 157.9±114.5 SU; G1, n=29, mean 357.6±217.9 SU; G2, n=29, mean 361.7±122.9 SU; and G3, n=30, mean 584.5±336.4 SU. Urinary tyrosine-phosphorylated protein levels are significantly different in groups of BC patients with different BC grades (Kruskal–Wallis test, P=6.99 × 10⁻²²). The solid lines indicate the mean values; the dotted line indicates the best cutoff value.

Figure 3

Urinary tyrosine-phosphorylated protein levels in BC subjects classified according to the WHO 2004 classification of BC. High grade, n=43, mean 528.06±297.19 SU; low grade, n=43, mean 354.22±191.16 SU. Urinary tyrosine-phosphorylated protein levels are significantly different in groups of BC patients with different BC grade (Kruskal–Wallis test, P=0.0005).

Figure 4

Urinary tyrosine-phosphorylated protein levels in relation to classification of recurrence and progression risks. (A) Analysis of UPY levels in relation to classification of recurrence risks of BC patients (Kruskal–Wallis test, P=0.002). (B) Analysis of UPY levels in relation to classification of progression risk of BC patients (Kruskal–Wallis test, P=0.001).

Figure 5

Urinary tyrosine-phosphorylated protein levels in subjects with different age ranges. (A) Distribution of UPY levels in control (CTRL) and BC patients with age ranges: ⩽55 years old (CTRL, n=53, mean 134.3±123.0 SU; BC, n=11, mean 326.8±164.9 SU), 56–65 years old (CTRL, n=72, mean 169.2±114.0 SU; BC, n=25, mean 437.2±354.5 SU), 66–75 years old (CTRL, n=28, mean 168.5±108.0 SU; BC, n=37, mean 431.2±187.1 SU) and >75 years old (CTRL, n=15, mean 168.2±103.4 SU; BC, n=19, mean 501.0±270.6 SU). Urinary tyrosine-phosphorylated protein levels are significantly increased in BC patients in all age groups (Wilcoxon's rank-sum test). The solid lines indicate the mean values. (B) Distribution of patients with different BC stages: data are expressed as percentage in different age groups. (C) Distribution of patients with different BC grades: data are expressed as percentage in different age groups.