# Oncogenic EFNA4 Amplification Promotes Lung Adenocarcinoma Lymph Node Metastasis
###### Abstract  
Lymph nodes metastases are common in patients with lung cancer. Additionally, those patients are often at a higher risk for death from lung tumor than those with tumor-free lymph nodes. Somatic DNA alterations are key drivers of cancer, and copy number alterations (CNAs) are major types of DNA alteration that promote lung cancer progression. Here, we performed genome-wide DNA copy number analysis, and identified a novel lung-cancer-metastasis-related gene, _EFNA4_. The _EFNA4_ genome locus was significantly amplified, and _EFNA4_ mRNA expression was significantly up-regulated in lung cancer compared with normal lung tissue, and also in lung cancer with lymph node metastases compared with lung cancer without metastasis. _EFNA4_ encodes Ephrin A4, which is the ligand for Eph receptors. The function of EFNA4 in human lung cancer remains largely unknown. Through cell line experiments we showed that _EFNA4_ overexpression contributes to lung tumor cells growth, migration and adhesion. Conversely, _EFNA4_ knockdown or knockout led to the growth suppression of cells and tumor xenografts in mice. Lung cancer patients with _EFNA4_ overexpression have poor prognosis. Together, by elucidating a new layer of the role of EFNA4 in tumor proliferation and migration, our study demonstrates a better understanding of the function of the significantly amplified and overexpressed gene _EFNA4_ in lung tumor metastasis, and suggests EFNA4 as a potential target in metastatic lung cancer therapy.  
EFNA4; lung cancer; lymph node metastasis; copy number alterations 2022144226. 14226.  
outcome of patients with lung cancer [8]. However, limited therapeutic effect on patients with distant metastasis leads to poor prognosis. Metastasis of lung cancer can occur, even when the primary tumor is still small. Therefore, patients with advanced LUAD have a poor prognosis, with an average 5-year survival rate of less than 20% [9; 10; 11; 12]. Further research on new therapeutic targets for LUAD is essential. The development of microarray and high-throughput sequencing technology has greatly promoted bioinformatics, which use of computers for processing the biological data, for example, form the Cancer Genome Atlas (TCGA). By mining these cancer genome data, new cancer-related genes can be effectively identified [13; 14; 15].  
_EFNA4_ is an important member of the ephrins family. Ephrins and ephrin receptors (Ephs) are membrane-binding proteins, forming a bidirectional and interactive signaling system [16; 17; 18]. Ephrins are membrane-bound proteins, divided into two families, class A and class B. The Ephrin A members are anchored to the membrane by glycosyl-phosphatidylinositol (GPI) whereas Ephrin B members possess a transmembrane segment. At the same time, their receptors (Ephs) were classified as EphA or EphB subfamily according to their preferential binding to Ephrin A or Ephrin B [19]. Previous studies have shown that members of the ephrin family play an important role in the pattern of nervous system, vascular system assembly and angiogenesis [20; 21; 22; 23; 24]. EFNA4 has also been reported to be involved in the proliferation of oral squamous cell carcinoma, hepatocellular carcinoma and gastric cancer [25; 26; 27]. However, the functions and mechanisms of _EFNA4_ in lung cancer are still not clear.  
We reported here the results of a systematic cancer genome analysis, and _EFNA4_ locus amplification was identified as one of the somatic DNA alterations that exerted a potent lung cancer driving function. Moreover, we found that _EFNA4_ overexpression is linked to lymph node metastasis in LUAD. Together, these results identified a novel oncogenic driving function of _EFNA4_ in lung cancer metastasis, and implicate _EFNA4_ as a potential target in lung cancer therapy.