Non-Invasive Diagnosis Of Early-Stage Lung Cancer Via Targeted High-Throughput Dna Methylation Sequencing Of Circulating Tumor Dna (Ctdna)

Current state-of-the-art lung cancer early screening involves using low-dose CT scan to identify lung nodules smaller than 3cm in diameter. However, it’s still a clinical dilemma to differentiate between malignant and benign nodules. We took the approach of methylation profiling by high-throughput bisulfite DNA sequencing in tissue samples to identify specific methylation signatures. We learned methylation patterns that differentiate malignant vs. benign lesions from tissue samples by in-depth data mining, and then used pattern matching to classify plasma samples. Given the usual low amount of ctDNA in plasma, we also developed an ultra-sensitive library preparation method to perform targeted bisulfite DNA sequencing from as low as 1ng of cfDNA or 1mL of plasma. From a training set of 88 tissue samples, which includes 60 malignant specimens with different subtypes (invasive adenomas, IA; minimally invasive adenomas, MIA; atypical adenomatous hyperplasia, AAH; adenocarcinoma in situ, AIS; carcinoid; lymphoepithelioma-like carcinoma of the lung, LELC; squamous carcinoma, SC; as well as 28 benign specimens in different categories including hematomas, granuloma, tuberculosis, inflammatory pseudotumor (IPT), sclerosing hemangiomas (SHL) and infections, we were able to achieve a sensitivity of 95% for identification of malignant lesions, with a specificity of 78.6%. From an independent validation set of 45 plasma samples, we achieved a sensitivity of 94.7% and a specificity of 85.7% for differentiating patients with malignant tumor from patients with benign lesions. Specifically, our assay is demonstrated to be highly sensitive towards early-stage lung cancer detection, with a sensitivity of 93.3% in a total of 15 patients with stage IA/B lung cancer. In summary, we have developed a highly sensitive blood-based non-invasive diagnostic assay for identification of early stage lung cancer, which can aid clinical decisions for patients with a CT scan positive for lung nodules. This approach can also be extended to non-invasive early screening for various cancer types. Citation Format: Xuyu Cai, Yangbin Gao, Hui Shen, Peter Laird, Jian-bing Fan, Weihong Xu, Wenhua Liang, Jianxing He. Non-invasive diagnosis of early-stage lung cancer via targeted high-throughput DNA methylation sequencing of circulating tumor DNA (ctDNA) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5383. doi:10.1158/1538-7445.AM2017-5383