J Cancer 2020; 11(20):5867-5879. doi:10.7150/jca.47318
Co-overexpression of AXL and c-ABL predicts a poor prognosis in esophageal adenocarcinoma and promotes cancer cell survival
1. Department of Surgery, Vanderbilt University Medical Center, Nashville, TN, USA.
2. Vanderbilt Center for Quantitative Sciences, Department of Biostatistics, Vanderbilt University Medical Center, Nashville, TN, USA.
3. Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA.
4. Department of Surgery, University of Miami Miller School of Medicine, Miami, FL, USA.
5. Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, TN, USA.
Hong J, Abid F, Phillips S, Salaria SN, Revetta FL, Peng D, Washington MK, El-Rifai W, Belkhiri A. Co-overexpression of AXL and c-ABL predicts a poor prognosis in esophageal adenocarcinoma and promotes cancer cell survival. J Cancer 2020; 11(20):5867-5879. doi:10.7150/jca.47318. Available from https://www.jcancer.org/v11p5867.htm
Background: Esophageal adenocarcinoma (EAC) is highly aggressive and characterized by poor prognosis. AXL expression has been linked to Barrett's tumorigenesis and resistance to chemotherapy, which is associated with c-ABL intracellular localization. However, the molecular and functional relationship between AXL and c-ABL and the clinical significance of the co-expression of these proteins in EAC remain unclear.
Methods: We used immunohistochemical analysis (IHC) on tissue microarrays containing human EAC samples (n=53) and normal esophageal tissues (n=11) in combination with corresponding deidentified clinicopathological information to evaluate the expression and the prognostic significance of AXL and c-ABL in EAC. The data were statistically analyzed using Kruskal-Wallis, the chi-square, the Fisher's exact, and Pearson tests. The Kaplan-Meier method and Cox proportional hazards regression model were used to evaluate cancer patient survival. We used a serum deprivation EAC cell model to investigate the pro-survival function of AXL and c-ABL using cell viability, apoptosis, and lactate dehydrogenase activity assays. We performed in vitro assays, including Western blotting, quantitative real-time PCR, and translational chromatin immunoprecipitation (TrIP-Chip) to study the molecular relationship between AXL and c-ABL in EAC cells.
Results: IHC analysis revealed that AXL and c-ABL were overexpressed in 55% and 66% of EAC samples, respectively, as compared to normal tissues. Co-overexpression of the two proteins was observed in 49% of EAC samples. The chi-square test indicated a significant association between AXL and c-ABL expression in the EAC samples (χ2 = 6.873, p = 0.032), and the expression of these proteins was significantly associated with EAC patient age (p < 0.001), tumor stage (p < 0.01), and lymph node status (p < 0.001). AXL and c-ABL protein expression data analysis exhibited an identical clinicopathological association profile. Additionally, we found a significant association between expression of AXL (χ2 = 16.7, p = 0.002) or c-ABL (χ2 = 13.4, p = 0.001) and survival of EAC patients. The Cox proportional hazards model and log rank test predicted a significant increase in mortality of patients with high expression of AXL [hazard ratio (HR): 2.86, 95% confidence interval (CI): 1.53 - 5.34, p = 0.003] or c-ABL [HR: 3.29, 95% CI: 1.35 - 8.03, p = 0.001] as compared to those patients with low expression of AXL or c-ABL proteins. Molecular investigations indicated that AXL positively regulates c-ABL protein expression through increased cap-dependent protein translation involving phosphorylation of EIF4E in EAC cells. Next, we investigated the functional relationship between AXL and c-ABL in EAC cells. We demonstrated that the pro-survival activity of AXL requires c-ABL expression in response to serum deprivation.
Conclusion: This study highlights the importance of the co-overexpression of AXL and c-ABL proteins as a valuable prognostic biomarker and targeting these proteins could be an effective therapeutic approach in EAC or other solid tumors expressing high levels of AXL and c-ABL proteins.
Keywords: Esophageal adenocarcinoma, Prognosis, Cell survival, AXL, c-ABL, Necrosis