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

Abstract

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.

DNA Methylation Cancer Biomarkers: Translation to the Clinic

“…While cytology analysis is typically performed on bronchoalveolar lavage (Carvalho et al, 2017) there is also opportunity to use the lavage fluid for liquid biopsy. Methylated SHOX2 and RASSF1A gene promoters were detected in lavage fluid from 322 patients with a sensitivity of 81% for lung cancer detection (Zhang et al, 2017). Other studies have also revealed high sensitivity for lung cancer detection using bronchoalveolar lavage fluid, with 75% and 78% for PCDHGA12 (Jeong et al, 2018) and SHOX2 (Dietrich et al, 2012) methylated DNA, respectively.…”

SEPT9 Gene Methylation as a Noninvasive Marker for Hepatocellular Carcinoma

“…DNA hypermethylation is an important event in carcinogenesis that can be detected in plasma-derived cell-free DNA, which makes it an ideal and useful biomarker for early detection and staging of cancer. Many studies have demonstrated that DNA methylation analysis of specific genes can be used for the early diagnosis of cancer, such as plasma mSEPT9 in colorectal cancer [ 19 ], bronchoalveolar lavage fluid SHOX2 and RASSF1A in lung cancer, [ 20 ], and urine GSTP1 in prostate cancer [ 21 ]. However, there is no FDA-approved circulating epigenetic biomarker that has been shown to be useful to diagnose HCC.…”

Precision Detection Technology: Equipping Precision Oncology with Wings

“…Additionally, detection of ctDNA methylation can be used for early tumor diagnosis and assessment of prognosis, as it can reflect tumor load changes [ 43 , 44 ]. The most widely used lung cancer-specific gene methylation detection kit is the human SHOX2 and RASSF1A gene methylation DNA detection kit [ 45 ], which could assist in the early diagnosis of lung cancer and in the differentiation between benign and malignant pulmonary nodules. Additionally, together with a cytological test, it can effectively improve the sensitivity and specificity of lung cancer detection.…”