Urinary protein profiling by liquid chromatography/tandem mass spectrometry: ADAM28 is overexpressed in bladder transitional cell carcinoma

Abstract

Bladder cancer is the most common urological cancer with higher incidence rate in the endemic areas of Blackfoot disease (BFD) in southern Taiwan. The aim of this study was to utilize the proteomic approach to establish urinary protein patterns of bladder cancer. The experimental results showed that most patients with bladder cancer had proteinuria or albuminuria. The urine arsenic concentrations of bladder cancer patients in BFD areas were significantly higher than those patients from non-BFD areas. In the proteomic analysis, the urinary proteome was identified by nano-high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (nano-HPLC/ESI-MS/MS) followed by peptide fragmentation pattern analysis. We categorized 2782 unique proteins of which 89 proteins were identified with at least three unique matching peptide sequences. Among these 89 proteins, thirteen of them were not found in the control group and may represent proteins specific for bladder cancer. In this study, three proteins, SPINK5, ADAM28 and PTP1, were also confirmed by Western blotting and showed significant differential expression compared with the control group. ADAM28 may be used as a possible biomarker of bladder cancer.

Developing proteomic biomarkers for bladder cancer: towards clinical application

“…Urinary concentrations of ADAM28 were also evaluated with ELISAs, which confirmed the increased abundance of ADAM28 in urine samples from patients with bladder cancer. 53 This finding was confirmed elsewhere using liquid chromatography and/or electrospray ionization tandem mass spectro metry to analyse urine samples from patients with bladder cancer. 54 In another study, pooled urine samples from 27 patients with bladder cancer were analysed using LC-MS/MS, which led to the identification of a total of 146 proteins.…”

“…51 This finding is in general agreement with the results of other studies in this area. 53,54 Increased urinary levels of disintegrin and metalloproteinase domain-containing protein 28 (ADAM28) have been detected using LC-MS/MS in two independent investigations. 52,53 In one study, analysis of pooled urine samples from 16 patients with bladder cancer and 18 individuals without the disease revealed 17 differentially expressed proteins, including four proteins uniquely identified in urine samples from patients without bladder cancer and 13 solely detected in urine samples from patients with bladder cancer.…”

“…53,54 Increased urinary levels of disintegrin and metalloproteinase domain-containing protein 28 (ADAM28) have been detected using LC-MS/MS in two independent investigations. 52,53 In one study, analysis of pooled urine samples from 16 patients with bladder cancer and 18 individuals without the disease revealed 17 differentially expressed proteins, including four proteins uniquely identified in urine samples from patients without bladder cancer and 13 solely detected in urine samples from patients with bladder cancer. 53 Among the 13 proteins identified only in patients with bladder cancer, serine protease inhibitor Kazal-type 5, ADAM28 and tyrosine-protein phosphatase non-receptor type 1 were selected for validation by western blotting of individual urine samples, although no justification was given for this selection.…”

“…53 Among the 13 proteins identified only in patients with bladder cancer, serine protease inhibitor Kazal-type 5, ADAM28 and tyrosine-protein phosphatase non-receptor type 1 were selected for validation by western blotting of individual urine samples, although no justification was given for this selection. 53 Findings from western blot analysis confirmed the proteomic results in all cases. Urinary concentrations of ADAM28 were also evaluated with ELISAs, which confirmed the increased abundance of ADAM28 in urine samples from patients with bladder cancer.…”

Changes in urine proteome accompanying diabetic nephropathy progression

“…The time interval between sample collection and preceding urination was at least 2 hours, but no more than 4 hours. Urine samples were stored at room temperature for less than 1 hour and processed using a modified standard protocol [5,6]. Briefly, 4 ml of 1 M HEPES (pH 7.2) buffer was added to 46 ml of urine in a Falcon tube to a final volume of 50 ml, vortexed for 2 minutes, and centrifuged at 3000 rcf for 10 minutes to remove cellular debris.…”