OLA1 is highly expressed in gastric cancer tissues
To detect the expression level of OLA1 mRNA in gastric cancer, we first downloaded two cohorts of GC datasets (GSE118916, GSE79973) from GEO database. Bioinformatic software QOE3.1 was employed to analyze the expression level of OLA1 mRNA in GC tissues and adjacent non-tumor tissues. As shown in Figure 1A, OLA1 mRNA level was significantly higher in GC tissues compared with that in adjacent non-tumor tissues (P<0.01, P<0.05, respectively) (Figure 1A). Then, qRT-PCR assay was used to confirm these results in 30 GC tissues and paired adjacent non-tumor tissues. Our results showed that OLA1 mRNA expression was significantly higher in GC tissues than that in adjacent non-tumor tissues (Figure 1B, P<0.01).
To further evaluate the expression pattern of OLA1 protein in GC tissues, immunohistochemical staining was conducted in GC tissue microarray including 334 GC tissues and corresponding 30 non-tumor tissues. OLA1 protein was detected in both gastric cancer tissues and adjacent non-tumor tissues. High expression of OLA1 protein was detected in 81.1% (252/334) primary GC tissues compared with 33.3% (10/30) in adjacent non-tumor tissues (P<0.01) (Figure 2A-C). These results suggested that OLA1 was highly expressed in GC tissues at both mRNA and protein level.
Association between OLA1 protein expression and clinicopathological parameters of gastric cancer.
In order to detect the clinical significance of OLA1 in GC, the relationships between the expression of OLA1 and clinicopathological parameters were analyzed. High or low expression rates of OLA1 protein in GC with respect to several standard clinicopathological features are presented in Table 1. It was showed that OLA1 protein overexpression was significantly related to tumor size, lymph node metastasis, TNM stage. The frequency of OLA1 positivity was significantly higher in patients with a tumor size ≥5cm compared with patients with a tumor size <5cm (81.9%, 122/149 vs 70.3%, 130/185, P=0.0146). Furthermore, the expression of OLA1 was higher in gastric cancer specimens with lymph node metastasis compared with those without lymph node metastasis (79.9%, 187/234 vs 65.0%, 65/100, P=0.0037). The proportion of OLA1 positive samples was lower in early TNM stage (I+II) than those in advanced stage (III+IV) (52.9%, 62/117 vs 87.6%, 190/217, P<0.001). There was no significant correlation between OLA1 protein expression and the other clinicopathological parameters, such as gender, age at surgery and histological type.
OLA1 predicts inferior prognosis of gastric cancer patients
Kaplan-Meier survival analysis showed that the mean survival time of patients with high expression of OLA1 in GC tissues was 31.34±1.73 months, which was significantly shorter compared with patients in the low OLA1 expression group, 42.54±2.95 months (P=0.002) (Figue 2D). Then, Multivariate Cox regression analysis showed that high expression of OLA1 was an independent inferior prognostic factor for GC patients (hazard ratio, 0.573; 95% confidence interval, 0.376-0.872; P=0.009), as well as other clinicopathological variables (lymph node metastasis and TNM stage) (Table 2).
OLA1 expression is positively correlated with Snail in gastric cancer tissues
Previous study reported that OLA1 contributed to epithelial-mesenchymal transition (EMT) in lung cancer by modulating Snail, which is a main EMT activating transcriptional factor. Upregulation of Snail plays a vital role in the initiation and development of GC. Thus, we aim to uncover the relationship between OLA1 and Snail expression in GC by immunohistochemical staining. We found that high Snail expression was detected in 65.6%(219/334) GC tissues. Furthermore, Spearman’s rank correlation analysis indicated a significantly positive correlation between expression of OLA1 and Snail in GC tissues (r=0.334, P<0.001) (Figure 3A-B, Table 3) .
Combining OLA1 expression with Snail reveals improved prognostic accuracy for GC patients
To analyze the prognostic value of combining OLA1 and Snail in GC, we divided the patients into four groups, OLA1 high expression/Snail high expression (OLA+/Snail+), OLA1 high expression/Snail low expression (OLA1+/Snail-), OLA1 low expression/Snail high expression (OLA1-/Snail+), OLA1 low expression/ Snail low expression (OLA1-/Snail-). The results revealed that group OLA1+/Snail+ had the worst prognosis, while the most favourable prognosis was seen in group OLA1-/Snail- (P<0.001) (Figure 3C). These data showed that the combination of OLA1 elevation and Snail increase in GC tissues seems to be predictive of the poorest prognosis.