Cisplatin-based combination chemotherapy is the foundation for treatment of advanced bladder cancer (BlCa), but many patients develop chemoresistance mediated by increased Akt and ERK phosphorylation. However, the mechanism by which cisplatin induces this increase has not been elucidated. Among six patient-derived xenograft (PDX) models of BlCa, we observed that the cisplatin-resistant BL0269 express high epidermal growth factor receptor, ErbB2/HER2 and ErbB3/HER3. Cisplatin treatment transiently increased phospho-ErbB3 (Y1328), phospho-ERK (T202/Y204) and phospho-Akt (S473), and analysis of radical cystectomy tissues from patients with BlCa showed correlation between ErbB3 and ERK phosphorylation, likely due to the activation of ERK via the ErbB3 pathway. In vitro analysis revealed a role for the ErbB3 ligand heregulin1-β1 (HRG1/NRG1), which is higher in chemoresistant lines compared to cisplatin-sensitive cells. Additionally, cisplatin treatment, both in PDX and cell models, increased HRG1 levels. The monoclonal antibody seribantumab, that obstructs ErbB3 ligand-binding, suppressed HRG1-induced ErbB3, Akt and ERK phosphorylation. Seribantumab also prevented tumor growth in both the chemosensitive BL0440 and chemoresistant BL0269 models. Our data demonstrate that cisplatin-associated increases in Akt and ERK phosphorylation is mediated by an elevation in HRG1, suggesting that inhibition of ErbB3 phosphorylation may be a useful therapeutic strategy in BlCa with high phospho-ErbB3 and HRG1 levels.