The role of trefoil factor 3 (intestinal) (TFF3) continues to be analyzed in numerous cancers, such as breast and gastrointestinal cancer, and has been associated with poor prognosis. tumors (95% CI: 13.8C16.6) (P=0.183). The median overall survival was 53.9 months in TFF3-positive tumors (95% CI: Non-applicable) vs. 44.4 months in TFF3-negative cases (95% CI: 30.5C58.3) (P=0.36). TFF3 negativity was significantly associated with higher tumor grade (P=0.05). Based on our results, further studies are required in order to elucidate whether survival PF-04929113 PF-04929113 and chemosensitivity are affected by TFF3 expression in ovarian cancer. in 2011, as being most predictive of ovarian carcinoma subtype in both the archival and the tumor bank cohorts (2). The selection of TFF3 was based on comprehensive gene expression profiling data (2,6). To the best of our knowledge, data on TFF3 in ovarian cancer are scarce. Therefore, the expression of the TFF3 in 91 ovarian cancer patients was investigated to determine its effect on prognosis and platinum sensitivity in ovarian cancer. Patients and methods Patients and treatment The study included patients with primary EOC treated between 1995 and 2008 at the Department of Obstetrics and Gynecology of Goethe University (Frankfurt, Germany). A total of 91 patients were retrospectively analyzed. Formalin-fixed, paraffin-embedded (FFPE) tissue samples were obtained from the Department of Pathology. The patient characteristics are listed in Table I. Clinical and pathological factors were evaluated by reviewing medical charts and pathology records. The Local Research Ethics Committee approved studies of human tissue and the samples were processed anonymously. Clinical outcome was assessed from the date of surgery to the date of death or until the end of 2009. Only patients with histologically proven EOC were included. The majority of the patients had advanced-stage disease (FIGO IIICIV). All the patients received primary surgery followed by platinum- and taxane-based chemotherapy. Table I. Patients characteristics. Tissue samples and immunohistochemistry The tissue samples were processed as previously described (7). Routine histopathological sections stained with hematoxylin and eosin were Tmem34 used for primary diagnosis and second reviewing (M.O.). Diagnosis and grading were performed according to the current World Health Organization criteria (8,9). Following mounting on Superfrost Plus slides (Fisherbrand, Fisher Scientific, Hampton, NH, USA), 2-m paraffin sections were dewaxed in xylene and rehydrated with graduated ethanol treatment. For antigen retrieval, the sections were incubated for 20 min in a microwave oven (800 W) using citrate buffer (10 mM; pH 6.0). The monoclonal anti-TFF3 antibody (ab57752, Lot GR71649-1; Abcam, Cambridge, UK,) was used at a dilution of 1 1:100. Incubation with the antibody for 1 h at room temperature was performed. For negative controls, the primary antibody was omitted. For secondary antibody incubation, the Dako REAL Detection System Alkaline Phosphatase/RED (REF K5005, Lot 20023341; Dako, Glostrup, Denmark) was applied, according to the manufacturer’s instructions. The sections were counterstained with hematoxylin (Gill no. 3, Lot 060M4356; Sigma-Aldrich, St. Louis, Missouri, USA). Expression levels for cytoplasmic TFF3 were scored semi-quantitatively based on the product of staining intensity (SI) and percentage of positive cells (PP) using the immunoreactive score (IRS) as follows (10): IRS=SIxPP. SI was assigned as 0, negative; 1, weak; 2, moderate; or 3, strong. PP was defined as 0, negative; 1, <10%; 2, 11C50%; 3, 51C80%; or 4, >80% positive cells. All assessments were performed blinded with respect to clinical patient data. Statistical analysis For statistical analysis a cut-off value was defined according to the IRS, i.e., scores of 0C3 (negative and low) were collectively defined as a low score, whereas scores 3 were defined as PF-04929113 a high TFF3 expression score. The Chi-square and Fisher’s exact tests were used to assess the associations between TFF3 expression of tumors and clinicopathological parameters. For those patients with available follow-up data, Kaplan-Meier curves were constructed and the log-rank test was used to determine the univariate significance of the variables. Cox regression analysis was performed to determine hazard ratios. All reported P-values were two-sided and P-values of 0.05 were considered to indicate statistically significant differences. All analyses were performed using the SPSS software package, version 22.0 (IBM SPSS, Armonk, NY, USA). Results Patient characteristics A total of 91 patients were included in this study. All the patients underwent primary debulking surgery including hysterectomy, bilateral salpingo-oophorectomy, infragastric omentectomy, appendectomy (mucinous subtype), systematic pelvic and para-aortic lymphadenectomy and, in distinct cases, peritonectomy and resection of affected tissues with intended resection of all visible tumors. In the majority of the patients (n=62; 68%) optimal debulking, i.e., achieving an optimal postoperative residual.