Abstract 2260: Classification of patients with lung cancer and benign lung disease via assessment of DNA methylation of SHOX2 and PTGER4 in plasma
Authors: Schlegel Anne , Hasinger Oliver , Esche Selina , Martini Melanie , König Thomas , Weiss Gunter 
Published: July 2016

Abstract
Introduction:
Annual screening for lung cancer (LC) with low-dose computed tomography (LDCT) is recommended in the US for specific high risk patients. Despite this, LC patients are frequently symptomatic at time of presentation and are often diagnosed with advanced stage disease associated with poor prognosis. At the same time the substantial rate of false positive findings of LDCT raised controversy about the risk profile of the groups eligible for LDCT screening. Non-invasive diagnostic tools may contribute to further risk stratification for screening candidates or patients with findings in LDCT. Aberrant DNA methylation detected in cell free-floating DNA has been shown to be clinically useful. Specifically, a DNA methylation panel of the genes SHOX2 and PTGER4 has been evaluated in three independent case-control studies comprising a total of 330 plasma specimens from LC patients and healthy individuals with promising results (AUC = 91 to 95%). Here, we report on further evaluation of the methylation marker panel in patients with malignant (LC) or benign lung disease (BLD).
Material and Methods:
For this case-control study 116 plasma specimen were collected from patients in Bulgaria and the US. The majority of patients were either diagnosed with LC (30 patients) or a BLD (50 patients). The remaining 36 subjects were healthy controls. A triplex real-time PCR assay for methylated DNA of SHOX2 and PTGER4 and the methylation independent ACTB control assay was developed. Total DNA was extracted from 3.5ml plasma samples and bisulfite converted utilizing a commercially available kit. After purification DNA was assayed in PCR triplicates. Cycle threshold values were aggregated utilizing a predefined algorithm. Receiver operating characteristic (ROC) curve and the area under the ROC curve (AUC) were analysed.
Results:
The marker panel showed significant discriminatory power to distinguish LC cases from the remaining subjects (AUC = 0.84), including BLD patients (AUC = 0.83) and healthy subjects (AUC = 0.86). At a fixed specificity level of 95% the sensitivity for LC was 50%, while for a fixed 90% sensitivity the observed specificity was 53%. BLD and healthy subjects were similar with respect to the methylation markers (AUC = 0.55, p-value = 0.45). The clinically important group of COPD patients (18 cases) was significantly different from the LC group (AUC = 0.76, p-value &lt; 0.01).
Conclusions:
Combination of DNA methylation markers SHOX2 and PTGER4 can be used to distinguish lung cancer patients from healthy subjects as well as patients with non-malignant diseases with high sensitivity at a reasonable false positive rate. The marker panel will be developed into a blood-based lung cancer diagnostic tool that may ultimately show clinical utility in combination with current imaging techniques to improve lung cancer risk stratification.
Citation Format: Anne Schlegel, Oliver Hasinger, Selina Esche, Melanie Martini, Thomas König, Gunter Weiss. Classification of patients with lung cancer and benign lung disease via assessment of DNA methylation of SHOX2 and PTGER4 in plasma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2260.