DNA Methylation Analysis of the SHOX2 and RASSF1A Panel in Bronchoalveolar Lavage Fluid for Lung Cancer Diagnosis

Introduction: Currently the majority of lung cancer patients are diagnosed as advanced diseases for no sensitive and specific biomarkers exist, noninvasive biomarkers with high sensitivity and specificity are urgently needed in lung cancer diagnosis.Bronchoscopy is a standard procedure of the diagnostic work-up of patients with suspected lung cancer despite of the limited diagnostic accuracy.Besides, epigenetic changes through DNA methylation play an important role in tumorigenesis.Thus, we examined the aberrant methylation of the SHOX2 and RASSF1A in bronchoalveolar lavage fluid (BALF) in comparing with conventional cytology examination and serum CEA in order to evaluate the new diagnostic method.Patients and Methods: BALF and serum samples were collected from 322 patients at the time of diagnosis, 284 of them were pathologically confirmed lung cancer, 35 were benign lung diseases and 3 were malignancies in other systems.For all of the 322 patients, the methylation status of the SHOX2 and RASSF1A gene were detected by a new RT-PCR platform and then confirmed by sanger sequencing.Serum CEA were detected using electrochemiluminescence immunoassay.Results: Profiling data showed the consistency of RT-PCR and sanger sequencing in detecting the methylation of the SHOX2 and RASSF1A.Besides, the combination of SHOX2 and RASSF1A methylation in BALF yielded a diagnostic sensitivity of 81.0% and specificity of 97.4%.When compared with established cytology examination (sensitivity: 68.3%, specificity: 97.4%) and serum biomarker carcinoembryonic antigen (CEA) (sensitivity: 30.6%, specificity: 100.0%), the SHOX2 and RASSF1A methylation panel showed the highest diagnostic efficiency.Notably, the combination of cytology and the SHOX2 and RASSF1A methylation panel could significantly improve the diagnostic efficacy. Conclusion:The methylation analysis of the SHOX2 and RASSF1A panel in BALF with RT-PCR achieved a satisfactory sensitivity and specificity in lung cancer diagnosis, especially in an early stage.It could be used as a promising noninvasive biomarker for auxiliary diagnosis of lung cancer.