Cisplatin-induced ototoxicity (CIO) is caused by cisplatin accumulation in the inner ear cochlea, which is mediated by organic cation transporter 2 (OCT2). Proton pump inhibitors, including lansoprazole (LPZ), ameliorated cisplatin-induced nephrotoxicity via inhibiting OCT2. In the present study, we investigated the protective effect of LPZ against CIO using zebrafish and real-world data. Using zebrafish, we compared the effect of LPZ on CIO through in vivo fluorescence imaging of the hair cells stained with fluorescence dyes. Cisplatin treatment to zebrafish significantly decreased the fluorescence intensities (FI) of hair cells (approximately 50% of those of control zebrafish). Co-treatment of LPZ or knockout (KO) of oct2 significantly suppressed the reduction of FI by cisplatin (approximately 75% of those of control zebrafish). The protective effect of LPZ were not significantly different between wild type and oct2-KO zebrafish. Using electronic medical records in Mie University Hospital, we validated the preventive effect of LPZ against CIO in 289 patients who received cisplatin contained chemotherapy. The rate of co-administrated LPZ in patients without ototoxicity was significantly higher than that of patients with ototoxicity (34% vs 7%). These results suggest that LPZ should suppress CIO through the inhibition of OCT2.