Abstract 2311: Cell-free DNA from bronchoalveolar lavage fluid (BALF) for the identification of lung cancer: A new medium of liquid biopsy
Authors: Jiang Junhong , Mu Chuanyong , Zhao Jun , Zeng Daxiong , Wang Changguo , Ye Junyi , Xu Jiayue , Su Jing , Han-Zhang Han , Li Bingsi , Li Haiyan , Zhang Zhihong 
Published: September 2020

Abstract
Introduction: LDCT, the gold standard for lung cancer screening, introduces excessive false positives. Differentiating malignant lung tumors from solitary pulmonary nodules is still a great challenge. cfDNA-based plasma testing has been developed for the purpose of screening and diagnosis. However, due to the limited amount of ctDNA present in early stage lung cancer patients, the sensitivity of such test is limited. Bronchoalveolar lavage (BAL) is a technique for sampling the epithelial lining fluid of the respiratory tract. Analysis of the returned fluid is used for the diagnosis of infections, interstitial lung diseases and sarcoidosis. Limited evidences support the use of BAL fluid (BALF) for lung cancer diagnosis. This study interrogates the potential of using BALF for lung cancer diagnosis.
Methods: 52 patients with solid pulmonary nodules (≤2cm in diameter) from the First Affiliated Hospital of Soochow University were enrolled, including 45 patients with solitary pulmonary nodule at initial diagnosis and 7 patients with small pulmonary nodule at advanced stage. Among the 45 patients, 30 patients underwent curative surgery, 3 patients had confirmed benign nodules by TTNA or follow-up and the remaining 12 with uncertain nodules at present. Tissue and BALF samples were obtained for mutation profiling using a panel consisting of 168 lung cancer-related genes. BALF samples with sufficient remaining DNA after somatic mutation profiling were subjected to DNA methylation profiling.
Results: Of the 30 patients underwent surgery, 24 were confirmed to have malignant nodules and 6 were confirmed to have benign nodules. First, we performed targeted sequencing on matched tissue and BALF samples from patients with malignant tumors (N=31, including patients with metastatic or relapsed disease). 93.5% (29/31) and 64.5% (20/31) patients had mutations detected from their tissue and BALF samples, respectively, resulting in a concordance of 71%. 83 and 59 mutations were detected from tissue and BALF samples, respectively. Using tissue samples as references, BALF samples revealed 20 new mutations and missed 44 mutations. Next, we evaluated the performance of BALF cfDNA mutation profiling in differentiating malignant from benign pulmonary nodules(≤2cm in diameter). 24 patients with malignant nodules and 9 patients with confirmed benign nodules were included for this analysis. BALF cfDNA mutation profiling resulted in a sensitivity of 63%, specificity of 100%, positive predictive value of 100% and negative predictive value of 50%. A majority of patients also underwent tracheoscopy, bronchial brushing and/or BALF cytology analysis with sensitivities of 12%, 9% and 0%, respectively. Targeted bisulfite sequencing was performed on 20 BALF samples (8 benign and 12 stage IA patients) to profile DNA methylation status. Of the 8 benign samples, only 1 patient with pulmonary tuberculosis exhibited a cancerous signal. Of the 12 stage IA samples, 10 exhibited cancerous signals, achieving 83% sensitivity and 87.5% specificity.
Conclusions: BALF cfDNA profiling demonstrates good performance in distinguishing small malignant tumors (≤2cm in diameter) from benign pulmonary nodules, thus having the potential to serve as a diagnostic tool. Targeted bisulfite sequencing shows potential as a supplemental diagnostic method.
Citation Format: Junhong Jiang, Chuanyong Mu, Jun Zhao, Daxiong Zeng, Changguo Wang, Junyi Ye, Jiayue Xu, Jing Su, Han Han-Zhang, Bingsi Li, Haiyan Li, Zhihong Zhang. Cell-free DNA from bronchoalveolar lavage fluid (BALF) for the identification of lung cancer: A new medium of liquid biopsy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2311.