Whole-genome sequencing reveals the molecular implications of the stepwise progression of lung adenocarcinoma

Article https://doi.org/10.1038/s41467-023-43732-y Whole-genome sequencing reveals the molecular implications of the stepwise progression of lung adenocarcinoma Received: 31 January 2023 Check for updates 1234567890():,; 1234567890():,; Accepted: 17 November 2023 Yasuhiko Haga1,12, Yoshitaka Sakamoto1,12, Keiko Kajiya1,12, Hitomi Kawai 2, Miho Oka1,3, Noriko Motoi 4,5, Masayuki Shirasawa6,7, Masaya Yotsukura8, Shun-Ichi Watanabe8, Miyuki Arai1, Junko Zenkoh1, Kouya Shiraishi 7,9, Masahide Seki 1, Akinori Kanai 1, Yuichi Shiraishi 10, Yasushi Yatabe 4, Daisuke Matsubara2, Yutaka Suzuki 1 , Masayuki Noguchi2,11, Takashi Kohno 7 & Ayako Suzuki 1 The mechanism underlying the development of tumors, particularly at early stages, still remains mostly elusive. Here, we report whole-genome long and short read sequencing analysis of 76 lung cancers, focusing on very early-stage lung adenocarcinomas such as adenocarcinoma in situ (AIS) and minimally invasive adenocarcinoma. The obtained data is further integrated with bulk and spatial transcriptomic data and epigenomic data. These analyses reveal key events in lung carcinogenesis. Minimal somatic mutations in pivotal driver mutations and essential proliferative factors are the only detectable somatic mutations in the very early-stage of AIS. These initial events are followed by copy number changes and global DNA hypomethylation. Particularly, drastic changes are initiated at the later AIS stage, i.e., in Noguchi type B tumors, wherein cancer cells are exposed to the surrounding microenvironment. This study sheds light on the pathogenesis of lung adenocarcinoma from integrated pathological and molecular viewpoints. Lung cancer is a major cause of death worldwide. An increasing number of patients are diagnosed with lung adenocarcinoma every year, rendering it the most frequently detected subtype of non-small cell lung cancer (NSCLC). In 1995, Noguchi et al. reported the “non-invasive” adenocarcinomas of the lung. They are small-sized adenocarcinomas (≤2 cm in diameter) that have classified into two groups1,2: replacement and non-replacement adenocarcinomas. The former was further subclassified into three subtypes (types A, B, and C). Patients with types A and B adenocarcinomas (pure-lepidic adenocarcinomas) have highly favorable prognosis and are considered very-early-stage adenocarcinomas, known as adenocarcinoma in situ (AIS). Thereafter, the prognosis of these patients rapidly deteriorates, with the 5-year survival rate 1 Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8561, Japan. 2Department of Diagnostic Pathology, Faculty of Medicine, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan. 3Ono Pharmaceutical Co., Ltd., Ibaraki, Japan. 4Department of Pathology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 5Department of Pathology, Saitama Cancer Center, 780 Komuro, Ina, Kita-Adachi-gun, Saitama 362-0806, Japan. 6Department of Thoracic Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 7Division of Genome Biology, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 8Department of Thoracic Surgery, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 9Department of Clinical Genomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 10Division of Genome Analysis Platform Development, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. 11Clinical Cancer Research Division, Shonan Research Institute of Innovative Medicine, Shonan Kamakura General Hospital, 1370-1 Okamoto, Kamakura, Kanagawa 247-8533, Japan. 12These authors contributed e-mail: ; ; equally: Yasuhiko Haga, Yoshitaka Sakamoto, Keiko Kajiya. Nature Communications | (2023)14:8375 1 Article of type C adenocarcinoma with fibroblastic proliferation foci being 75%. In the current World Health Organization (WHO) histological classification3, lung adenocarcinomas (≤3 cm in diameter) showing pure-lepidic growth are classified collectively as AIS. AIS corresponds to Noguchi types A and B to some extent and show a 100% 5-year patient survival rate. However, to ensure the accurate diagnosis of AIS, WHO has introduced a new type of adenocarcinoma called minimally invasive adenocarcinoma (MIA). These adenocarcinomas have a small invasion area (≤5 mm) and a likely have an extremely favorable prognosis. In both pathological criterion, AIS is defined as a pure-lepidic, patterned tumor with a diameter of ≤3 cm that shows no relevant invasion. AIS shows lepidic growth, mimicking non-tumorous lung alveoli. It does not exhibit invasive features, such as having non-lepidic histological subtypes, causing necrosis, and leading to vascular and lymphatic invasions. MIA also has a diameter of ≤3 cm, but it is characterized by its small invasion area (≤5 mm). It is considered to eventually progress to the overtly invasive adenocarcinoma (≥3 cm). Tumor can also develop into more malignant and distinct histological subtypes, such as papillary, acinar, micropapillary, and solid subtypes, other than the lepidic subtype. Although patients with an invasive lung adenocarcinoma shows a poor prognosis, those with AIS and MIA can be successfully treated by performing surgical resection4. Therefore, to understand additional and essential molecular events that lead to the development of invasive adenocarcinoma, it is crucial to characterize its molecular progression from being a pre-invasive (AIS or MIA) to overtly invasive adenocarcinoma. Recently, several researchers worldwide, The Cancer Genome Atlas5,6, and International Cancer Genome Consortium (ICGC)7,8 have attempted to sequence the lung adenocarcinoma genome. In particular, the Pan-Cancer Analysis of Whole Genomes9 consortium of ICGC conducted the whole-genome sequencing (WGS) analysis of lung adenocarcinomas, covering single nucleotide variants (SNVs) and large-scale structural variants (SVs). However, early-stage cancers are not sufficiently represented in these datasets, as the analyzed cases mainly include cancers of advanced stages with invasive characteristics. Recently, some studies have reported genomic and epigenomic features and their heterogeneity in early lung adenocarcinoma genomes10–12. However, these studies only performed short read sequencing in limited regions of the genome (i.e., the exome) or focused on AIS parts within overall advanced tumors. Thus, a precise and comprehensive characterization of SVs and other complicated genomic aberrations has not been achieved so far. Notably, long read sequencing could not be performed for lung adenocarcinoma genomes until very recently, as the meager amount of required starting materials hampered intensive analyses of small-tumor specimens. (...truncated)