HCC is the fifth most common cancer and the third most common cause of cancer-related deaths worldwide. It has a high prevalence in Asia, including Korea, due to endemic hepatitis B virus infection. numerous clinical trials using cytotoxic chemotherapy or targeted agents have failed, except sorafenib . While intrahepatic recurrence is most common, lung metastases are the most common extrahepatic spread and account for up to 40% of metastatic HCC . survival of patients with metastatic HCC is usually less than 1 year . Recently, prolonged median survival durations have been reported from 6.4 months to 40.7 months following metastasectomies . However, despite advances in therapeutic strategies, extrahepatic metastasis is still a major impediment to better prognosis in HCC patients.
In this study, we employed whole-genome sequencing to profile somatic mutations and structural variations in primary HCC (HBV+) and their matched metachronous lung metastases. Several recent studies on HCCs of different etiologies (HBV, HCV, and alcohol) using whole-genome and exome sequencing have not only confirmed previously known mutations of β-catenin and TP53 but also identified novel genetic alterations in genes that are involved in epigenetic regulation, such as ARID1A, ARID1B, ARID2, MLL, and MLL3 . Wnt/β-catenin, p53 signaling, cell cycle, and chromatin remodeling have recently emerged as dominant cancer pathways in primary HCC . To the best of our knowledge, this study is the first to examine the genomic profiles of matched pairs of primary and metastatic HCC by whole-genome sequencing.
In summary, comparison of structural variations between primary and metastatic tumors shows very similar and largely overlapped mutated segments (Figure 1). In addition, CNVs in primary and metastatic pairs were found to be closely related, and 38.88-78.49% of SNVs detected in primary HCC tumors were also detected in lung metastases. These preservations in genomic profiles from liver primary tumors to metachronous lung metastases indicate that the genomic features during tumorigenesis may be retained during metastasis.
Despite similarities between primary and metastatic pairs, a few somatic mutations were detected only in metastatic tumors. Multiple mutations of zinc finger (ZNF) genes were detected in the metastasis samples, and mutated genes that are involved in apoptosis and transcription regulations were enriched in metastases (Additional file 1: Table S10B). The ZNF family represents a large group of proteins involved in various aspects of transcriptional regulation . It has been reported that there were almost twice as many ZNF mutated genes in the HBV-positive hepatitis . In our study, a total of 5 ZNF, namely ZNF257, ZNF682, ZNF404, ZNF514, and ZNF142, were mutated in lung metastases from HCC (Additional file 1: Table S10B). Among these, ZNF257 and ZNF682 were previously reported to be upregulated in lung and bone metastases, relative to the primary breast adenocarcinoma, and were speculated to be putative metastasis genes . In addition, ZNF mutations were found in head and neck squamous cell carcinoma (HNSCC); therefore, the genetic changes to these transcriptional regulators have been implicated in the development of HNSCC . Nevertheless, the functional role of ZNFs in HCC tumorigenesis is yet to be defined.
In addition, mutations in known cancer gene suppressor TP53 were also detected in our study samples, including a nonsense mutation (E343*) in the regulator of TAT binding and a missense mutation (N131D) in the DNA-binding domain, both were previously reported by COSMIC (http://cancer.sanger.ac.uk/cancergenome/projects/cosmic/) (Additional file 1: Table S3).
Notably, a frameshift insertion in WNT5A and an in-frame deletion in DAAM2 were detected only in lung metastases; both WNT5A and DAAM2 are involved in the Wnt signaling pathway. Recent genome sequencing studies have revealed that the WNT signaling pathway is the most frequently altered oncogenic pathway in primary HCC . In addition to the high prevalence of CTNNB1 mutations, genetic alterations of other components, such AXIN1, AXIN2, and APC genes that encode proteins containing the destruction box, also occur at a lower frequency. Wnt5a, which is up-regulated in poorly differentiated and highly motile mesenchymal-like HCC cells, has been suggested to play a role in tumor progression by inducing epithelial mesenchymal transition . In line with this, Wnt5a up-regulation was significantly shown to enhance migration, proliferation, and invasiveness in pancreatic cancer cells in vitro. Importantly, Wnt signaling has been implicated in the activation of HCC-initiating cells [30, 31]. While Wnt5a has been described as a tumor promoter in melanoma, gastric, pancreatic, and prostate cancers, it was suggested to be a tumor suppressor in HCC, neuroblastoma, leukemia, colon, and thyroid cancers . The functional significance of Wnt5 aberrations in tumorigenesis of lung metastases from HCC primary tumors should be evaluated in subsequent studies.
Synthetically, primary tumor harbored more alterations located in Wnt, cell cycle, JAK-STAT pathways (Figure 5), which plays an important role in encoding protein molecular emphasizing the cell proliferation, apoptosis and inflammation, as well as focal adhesion pathway. Focal adhesion pathway has been implicated in the cell migration and initiated metastasis . For metastasis, specific alterations arose in tight junction and focal adhesion pathways (Figure 6), which enhanced the cell migration mechanism.
In this study, for the first time, the whole-genome sequencing profiles of primary HCC and paired lung metastases were compared. We found similar genomic alterations between them, with a few mutations that were found specifically in lung metastases, which may explain the clinical observation that both primary and metastatic tumors are usually sensitive or resistant to the same systemic treatments.