Recent developments in the genomic characterization of tumors have contributed to novel therapeutic approaches, and some molecular-targeted therapies based on the genetic profiles of tumors have improved patient survival. Human epidermal growth factor 2 (HER2) is a member of the HER family containing four receptor tyrosine kinases. Recently, we have identified novel mutations in the transmembrane domain of HER2 in lung adenocarcinomas. In this study, we clarified that these mutations could be oncogenic alterations of lung cancer. We also investigated the efficacy of afatinib as a HER2-targeted therapy in these HER2 altered lung cancer cells. In addition, we clarified a novel HER2 binding protein cytokeratin 19 (KRT19), and investigated the impact of KRT19 and HER2 interactions in signal transduction pathways to decode their possible roles in oncogenesis.