We report a case of panitumumab-resistant rectal cancer with HER2 gene amplification detected by CancerPlex®. A 51- year-old man was diagnosed with an obstructive rectal cancer having lung and adrenal metastases. He underwent the Hartmann 's operation, and KRAS mutations were not detected. After the surgery, 3 courses of CapeOx plus bevacizumab were administered as first-line chemotherapy; however, the lung and adrenal metastases progressed. Subsequently, 24 courses of IRIS/panitumumab was administered as second-line chemotherapy, and the metastases slowly progressed. Six courses of regorafenib were administered as third-line chemotherapy followed by a course of TAS-102 as fourth-line chemotherapy. Subsequently, a left femoral head metastasis and cerebellar metastases were detected. The patient received best supportive care including palliative femoral head replacement and stereotactic irradiation for the cerebellar metastases, and he died of cancer 3 years 5 months after the primary surgery. The comprehensive genomic analysis focusing on 413 cancer-related genes with CancerPlex®revealed that EGFR, BRAF, KRAS, NRAS, and HRAS had no mutations; however, ERBB2 amplification was detected. Furthermore, immunohistochemical staining revealed overexpression of HER2 protein in both the primary and bone metastatictumor. HER2 and EGFR independently promote the RAS-RAF-MAPK pathway. In the present case, the efficacy of anti-EGFR therapy may be attenuated because of ERBB2 amplification in the metastatic tumor.