Novel prognostic protein markers of resectable pancreatic cancer identified by coupled shotgun and targeted proteomics using formalin-fixed paraffin-embedded tissues.
Pancreatic cancer is among the most lethal malignancies worldwide. We aimed to identify novel prognostic markers by applying mass spectrometry (MS)-based proteomic analysis to formalin-fixed paraffin-embedded (FFPE) tissues. Resectable, node positive pancreatic ductal adenocarcinoma (PDAC) with poor (n = 4) and better (n = 4) outcomes, based on survival duration, with essentially the same clinicopathological backgrounds, and noncancerous pancreatic ducts (n = 5) were analyzed. Cancerous and noncancerous cells collected from FFPE tissue sections by laser microdissection (LMD) were processed for liquid chromatography (LC)-tandem MS (MS/MS). Candidate proteins were identified by semiquantitative comparison and then analyzed quantitatively using selected reaction monitoring (SRM)-based MS. To confirm the associations between candidate proteins and outcomes, we immunohistochemically analyzed a cohort of 87 cases. In result, totally 1,229 proteins were identified and 170 were selected as candidate proteins for SRM-based targeted proteomics. Fourteen proteins overexpressed in cancerous as compared to noncancerous tissue showed different expressions in the poor and better outcome groups. Among these proteins, we found that three novel proteins ECH1, OLFM4 and STML2 were overexpressed in poor group than in better group, and that one known protein GTR1 was expressed reciprocally. Kaplan-Meier analysis showed high expressions of all four proteins to correlate with significantly worse overall survival (p < 0.05). In conclusion, we identified four proteins as candidates of prognostic marker of PDAC. The combination of shotgun proteomics verified by SRM and validated by immunohistochemistry resulted in the prognostic marker discovery that will contribute the understanding of PDAC biology and therapeutic development.
Division of Gastroenterological Surgery, Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Miyagi, Japan.
Tatsuyuki
T
Takadate
Tohru
T
Onogawa
Tetsuya
T
Fukuda
Fuyuhiko
F
Motoi
Takashi
T
Suzuki
Kiyonaga
K
Fujii
Makoto
M
Kihara
Sayaka
S
Mikami
Yasuhiko
Y
Bando
Shimpei
S
Maeda
Kazuyuki
K
Ishida
Takashi
T
Minowa
Nobutaka
N
Hanagata
Hideo
H
Ohtsuka
Yu
Y
Katayose
Shinichi
S
Egawa
Toshihide
T
Nishimura
Michiaki
M
Unno
D000368:Aged; D014408:Biomarkers, Tumor / Q000032:analysis*; D021441:Carcinoma, Pancreatic Ductal / Q000737:chemistry* / Q000401:mortality; D002853:Chromatography, Liquid; D005260:Female; D005557:Formaldehyde; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D009363:Neoplasm Proteins / Q000032:analysis*; D010190:Pancreatic Neoplasms / Q000737:chemistry* / Q000401:mortality; D016612:Paraffin Embedding; D011379:Prognosis; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry
D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D005557:Formaldehyde