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Gastric cancer is a disease with high heterogeneity, and this heterogeneity may result in an uneven distribution of subclones with varied genetic properties at disease locations (spatial heterogeneity) or temporal changes in subclonal composition (temporal heterogeneity). We present the case of a 69-year-old woman with metastatic gastric cancer who presented for axillary lymph node enlargement and underwent axillary lymphadenectomy. Pathological evidence showed human epidermal growth factor receptor 2 (HER2)(3+). Abdominal computed tomography revealed a mass in the gastric body, gastroscopic biopsy showed HER2(3+). After tumor shrinkage by preoperative translational chemotherapy (oxaliplatin, calcium folate, fluorouracil) and targeted therapy (trastuzumab), she had laparoscopic-assisted total gastrectomy. However, HER2 immunohistochemistry was found to be diffusely negative in the surgically removed tissue, and there was no evidence of HER2 amplification in the whole exon sequencing either. After 10 months of trastuzumab treatment, her disease progressed. Although trastuzumab treatment was initially beneficial, the residual HER2-negative subclones may cause tumor recurrence and metastasis due to temporal heterogeneity, as shown in this case. [ABSTRACT FROM AUTHOR]