Partner of Sld five 3: a potential prognostic biomarker for colorectal cancer

The study population comprised 137 consecutive patients (79 males and 58 females) who were examined and treated at Yuhuangding Hospital between January 2008 and December 2012 for colorectal cancer. All cases underwent complete resection in this study. Details of the clinical and demographic information, prognostic factors, and disease progression were collected prospectively. Of the 137 patients, 46, 54, 22, and 15 had stage I, II, III, and IV tumors, respectively. Forty-two patients were administered postoperiative adjuvant chemotherapy every three weeks for six months (Oxaliplatin 130"mg/m² d1 + Capecitabine 1000 mg/m² d1-d14). The study protocol was approved by the institutional review board of Yuhuangding Hospital and the study was conducted according to the principles of the Declaration of Helsinki. All patients provided written informed consent.

Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was used to determine the PSF3 expression level. Briefly, total RNA was extracted with Trizol reagent (Invitrogen, Grand Island, NY, USA) and dissolved in water according to the manufacturer"s instructions. Relative complementary DNA (cDNA) quantitation for PSF3 and an internal reference gene (β-actin) was done using a fluorescence-based, real-time detection method. The sequences of the primer used were as follows: PSF3 forward 5′-TGACAGTCCCGAGAATGCAGA-3′ and reverse 5′-TGCCTACCAGGGCTGAAGTG-3′; β-actin (internal reference gene) forward 5′-TGGCACCCAGCACAATGAA-3′, reverse 5′-CTAAGTCATAGTCCGCCTAGAAGCA-3′. The PCR mixture consisted 1200"nmol/l primer, 200"nmol/l probe, 200 nmol/l each of deoxyadenosine triphosphate, deoxycytidine triphosphate, deoxyguanosine triphosphate, deoxythymidine triphosphate, 3.5 mmol/l MgCl₂, and × 1 Taqman Universal PCR Master mix to a final volume of 20 μl (all reagents were from PE Applied Biosystems, Foster City, CA). Cycling conditions were 95"C for 35"s and 60"C for 30s, followed by 40"cycles at 95"C for 15"s and 60"C for 1"min. Relative gene expression levels are expressed as ratios (differences between the Ct values) between two absolute measurements (PSF3/β-actin).

The follow-up period was defined as the interval between the date of operation and the date of the patient"s death or the last visit. The follow-up time ranged from 7 months to 73 months (median, 41 months). All patients were followed until May 2014 with a follow-up rate of 100%. Disease-free survival was measured from the day of surgery to the day of the first evidence of tumor recurrence or metastasis. Overall survival was measured from the day of surgery to the day of death.

Associations between PSF3 expression in tumor tissue and clinicopathological features were determined using the χ²-test. Survival was examined using the Kaplan-Meier method, and the significance of the difference was evaluated by a log-rank test. A Cox regression analysis was carried out to assess independent prognostic factors for disease-free survival and overall survival in colorectal cancer. All statistical calculations were performed using SPSS software (SPSS 17.0, Chicago, IL, USA) and P <0.05 was considered statistically significant.

The mRNA expression level of PSF3 were determined in 137 colorectal cancer and the adjacent normal tissues by qRT-PCR. Median mRNA expression levels were 1.35 × 10^-3 (range 2.88 × 10^-4 to 3.16 × 10^-2) for tumor tissues and 2.94 × 10^-4 (range 5.48 × 10^-5 to 1.27 × 10^-3) for adjacent normal tissues, and the differences were statistically significant (P < 0.05). To evaluate the role of PSF3 in colorectal cancer, we investigated whether PSF3 expression was associated with any of clinicopathological variables in the 137 enrolled cases of colorectal cancer. By adopting cut-off value according to median PSF3 expression level, we found that PSF3 expression was significantly associated with tumor size, depth of wall invasion, lymph node metastasis, TNM stage, tumor differentiation, and five-year survival. No significant relationship was noted between PSF3 expression and age, gender, distant metastasis, and adjuvant chemotherapy (Table"1).

Using the data collected from 137 patients, we evaluated their prognosis and its relationship to the expression of PSF3. The disease-free survival in patients with low PSF3 levels (39.5 × 7.2 months) was significantly longer than that in patients with high levels (28.6 × 6.4 months) (P = 0.007; Figure 1A). Univariate analysis initially included age, gender, tumor size, depth of wall invasion, lymph node metastasis, distant metastasis, tumor differentiation, TNM stage, adjuvant chemotherapy, and PSF3 expression level for disease-free survival analysis. The tumor size, lymph node metastasis, distant metastasis, TNM stage, tumor differentiation, adjuvant chemotherapy, and PSF3 expression level were associated with disease-free survival and were introduced into the multivariate analysis (Table"2). In the multivariate analysis, late TNM stage, poor differentiation, and high PSF3 level were shown to have a statistically independent prognostic value with respect to disease-free survival (Table"2). In addition, we also examined the overall survival of PSF3 low level and PSF3 high level groups and found a statistically difference between the two groups by using the log-rank test (P = 0.003). The median survival time of patients with low PSF3 levels (59.7 × 13.8 months) was significantly longer than that of patients with high PSF3 levels (47.2 × 11.4 months) (Figure 1B). A univariate analysis indicated that among the clinicopathological factors, tumor size, lymph node metastasis, distant metastasis, tumor differentiation, TNM stage, adjuvant chemotherapy, and PSF3 expression level were correlated with the outcome (Table"3). Further assessment using the Cox multivariate analysis indicated that distant metastasis, poor differentiation, late TNM stage, and high PSF3 expression were statistically significant predictors for poor overall survival (Table"3).

In the current study, we further analyzed the association of PSF3 expression in early stage (stage I and II) colorectal cancer. Among the early stage cases, 46 and 54 patients were classified as high PSF3 level and low PSF3 level, respectively. A survival analysis that included only early stage patients revealed that the overall survival for the low PSF3 expression group was longer than that for the high PSF3 expression group. The log-rank test showed that the intergroup difference was statistically significant (P < 0.001; Figure 1C).