Erythropoietin and Erythropoietin Receptor Expression in Early Stage Non-Small Cell Lung Cancer: Prognostic Significance.
Authors: Amin Khalid , Haroon Zishan A. , Kim Seok J. , Li Sufeng , Rabbani Zahid N. , Vollmer Robin T. , Wang Xiaofei F. , Kelley Michael J. , Arcasoy Murat O. 
Published: April 2020

Abstract
Erythropoietin (EPO) and its cell surface receptor EPOR have been shown to be expressed in many different types of cancer such as breast, prostate and head and neck cancer. The objectives of this study were to determine the frequency of EPO and EPOR expression in non-small cell lung cancer (NSCLC) and to characterize the association of EPO and EPOR expression with clinical and histological features of early stage NSCLC and its outcome. Seventy three patients with primary NSCLC from the Durham Veterans Affairs Hospital who underwent primary resection of their tumors were included in the study after providing informed consent in accordance with a research protocol approved by the Institutional Review Board at Durham Veterans Administration Medical Center. The median follow up period was 3.8 years. At the time of last follow-up 24 patients (33%) had recurred and 38 patients (52%) had died. Histologically, 38 primary tumors (52%) were squamous cell carcinomas, 27 (37%) were adenocarcinoma (ADC) and 8 (11%) were other types of NSCLC. Thirty-three patients (45%) had T1 lesions, 37 (51%) had T2 and 3 patients (4%) had T3 tumors. Nodal stage was N0 in 59 patients (81%) and N1 in 14 patients (19%). Fifty-six patients (77%) had pathologic stage I disease and 17 patients (23%) has stage II disease. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded tumors from each patient using monoclonal antibodies against EPO and EPOR. The expression was determined using a semi-quantitative scoring scale taking into consideration both the intensity and the percentage of tumor cells staining positive. Weak expression in ≤ 5% of tumor cells was considered negative. EPO was expressed in 60 of the 73 tumors (82%) and frequently exhibited a focal pattern of staining. EPOR was expressed in 60 tumors (82%) with predominantly cytoplasmic localization except for several cases where membrane staining of EPOR was also observed. Both EPO and EPOR were expressed in 55 (75%) tumors. We also characterized the expression of hypoxia-inducible factor-1a (HIF1a), the hypoxia regulated protein CA-IX, and to determine the Ki-67 proliferation index and microvessel density (MVD) using CD31 immunostaining. Patients with adenocarcinoma had lower EPOR expression in comparison to patients with other histologic subtypes (Wilcoxon test, P=0.03). EPOR expression was not associated significantly with other clinical or histological parameters or outcome. Patients with early nodal stage (N0) and overall pathologic stage (stage I) disease had significantly higher EPO expression (Wilcoxon test, P=0.01 and P=0.02, respectively). High MVD levels were associated with high EPO expression (Wilcoxon test, P=0.01) but EPO was not associated with any other histologic variables. Interestingly, higher EPO expression was associated with significantly better recurrence-free survival (Logrank test, P=0.006) and overall survival (P=0.008). Cox proportional hazard regression analysis revealed that high EPO expression was associated with better survival (P&lt;0.05). In multivariate analysis, EPO was not an independent prognostic factor. In conclusion, these data in our cohort of patients demonstrate that
- EPO and EPOR are expressed with high frequency in primary NSCLC with the majority of tumors co-expressing both EPO and EPOR and - High EPO expression in tumor cells is associated with better recurrence-free and overall survival.