Flot-2 Expression Correlates with EGFR Levels and Poor Prognosis in Surgically Resected Non-Small Cell Lung Cancer

Abstract

We previously reported that expression of Flotillin 2 (Flot-2), a protein isolated from caveolae/lipid raft domains, increased significantly in nasopharyngeal carcinoma (NPC) compared with normal tissues. Signal transduction through epidermal growth factor receptors (EGFR) and Flot-2 play an important role in cancer development, but their precise role in lung cancer has not been investigated. In this study, we have investigated the correlation between the expression of Flot-2 and EGFR, which increase significantly in non-small cell lung cancer (NSCLC) patients (n=352) compared with non-cancer tissues. Additionally, patients with advanced stages of NSCLC had higher positive expression of Flot-2 and EGFR than patients with early stages. NSCLC patients with increased expression of Flot-2 and EGFR had significantly less overall survival rates than patients with less expression of Flot-2 and EGFR. Taken together, our data suggest that increased expression of Flot-2 and EGFR in NSCLC patients is inversely proportional to the disease prognosis and that increased expression of Flot-2 associated with increased EGFR may serve as a biomarker to predict poor disease prognosis.

Fig 1. Expression of Flot-2 protein in lung SCC cells, lung ADC cells and control of non-cancerous lung tissues were detected by IHC using specific antibody as described in the section of materials and methods.

Strong positive staining of Flot-2 protein was found on cell membranes of lung SCC and lung ADC cells (Fig 1A and 1B, 20×, IHC, DAB staining). Negative staining of Flot-2 was showed in non-cancerous lung tissue (Fig 1C, 20×, IHC, DAB staining). Negative control showed no Flot-2 staining in lung ADC cells (Fig 1D, 20×, IHC, DAB staining).

Fig 2. Expression of EGFR protein in lung SCC cells, lung ADC cells and the control of non-cancerous lung tissue were detected by IHC using specific antibody as described in the section of materials and methods.

Strong positive staining of EGFR protein was found in cell membranes and cytoplasm of lung SCC and lung ADC cells (Fig 2A and 2B, 20×, IHC, DAB staining). Negative staining of EGFR was showed in non-cancerous lung tissue (Fig 2C, 20×, IHC, DAB staining). Negative control showed no EGFR staining in the lung SCC cells (Fig 2D, 20×, IHC, DAB staining).

Fig 3. Expression of Flot-2 and EGFR proteins in lung SCC and lung ADC compared to the control of non-cancerous lung tissues.

Results showed that there were significant differences between the groups which were statistically evaluated by chi-square test.

Fig 4. Kaplan-Meier cures for overall survival of lung SCC and ADC patients with expression of Flot-2 and EGFR.

Kaplan-Meier analysis was used to plot the overall survival curves of 159 cases of lung SCC and 193 cases of lung ADC patients with differential expression of Flot-2, EGFR, and combined expression of either of these two proteins, which statistical significance was assessed by log-rank test. (A) Lung SCC patients with positive expression of Flot-2 protein showed worse overall survival rates compared to patients with negative Flot-2 (P = 0.043, two sided). (B) Positive expression of EGFR had no significantly correlation with overall survival rates of lung SCC patients (P> 0.05, two sided). (C) Kaplan-Meier curves showed lung SCC patients with positive expression with either of Flot-2 and EGFR proteins had worse overall survival rates than these with all negative staining of two proteins above (P = 0.02, two sided). (D) Lung ADC patients with positive expression of Flot-2 had worse overall survival rates than that with negative one (P = 0.007, two sided, respectively). (E) Lung ADC patients with positive expression of EGFR had worse overall survival rates than that with negative one (P = 0.033, two sided, respectively). (F) Lung ADC patients with positive expression with either of Flot-2 and EGFR proteins showed worse overall survival rates compared with all negative staining of two proteins above (P = 0.005, two sided).