Peritoneal recurrence of gastric cancer patients is a serious problem. Recently, the CD44 molecule was reported to be a marker of cancer stem cells in gastric cancer. In this study, we examined the prognostic significance of CD44 mRNA levels in magnetically separated CD45-negative EpCAM-positive (CD45‑EpCAM+) cells, in the peritoneal washes collected from gastric cancer patients with TNM stage II and III. A total of 147 gastric cancer patients with stage II (n=75) and III (n=72) were included. All patients were negative by peritoneal cytology. Peritoneal washes of the Douglas pouch were collected and used for pathological cytology and molecular diagnosis. Prior to molecular diagnosis, CD45‑EpCAM+ cells were separated from peritoneal washings by an auto-magnetic-activated cell separation system. CD44 and CEA mRNA levels of the CD45‑EpCAM+ fraction were detected by real-time RT-PCR. The CD44 mRNA and CEA mRNA levels in the peritoneal washes showed a significant correlation with tumor size and stage. In patients with stage III, peritoneal recurrence-free survival rates (PRFS) and overall survival rates (OS) of CD44 mRNA+ or CEA mRNA+ patients were significantly worse than those of marker genenegative patients. In stage II patients, PRFS and OS of CD44 mRNA+ patients were significantly worse than those of marker gene-negative patients. In the Cox regression hazard model analysis, the presence of CD44 mRNA in peritoneal washes was found to be an independent prognostic factor for PRFS and OS in stage II and III. In contrast, CEA mRNA levels of these samples showed a prognostic value only in stage III. Our results suggest that CD44 mRNA of magnetically separated CD45‑EpCAM+ cell fraction of peritoneal washes is a useful genetic marker for predicting high-risk individuals among gastric cancer patients with stage II and III.

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