Abstract Background The standard neoadjuvant therapy for resectable locally advanced esophageal squamous cell carcinoma (ESCC) in Japan is DCF (docetaxel [DTX] + cisplatin [CDDP] + 5-fluorouracil [5-FU]). In addition, neoadjuvant immune checkpoint inhibitors have shown a survival benefit for metastatic ESCC and several other cancers. We previously reported that neoadjuvant CF (CDDP + 5-FU) + nivolumab (Nivo), DCF + Nivo, and FLOT (5-FU + l-leucovorin [l-LV] + oxaliplatin [L-OHP] + DTX) + Nivo are tolerable and show promising short-term efficacy for resectable locally advanced ESCC. However, the long-term efficacy of these neoadjuvant treatments remains unclear. Methods JCOG1804E (FRONTiER) is a multi-cohort phase I study designed to evaluate the safety and efficacy of Nivo combined with neoadjuvant CF (cohorts A and B [adding run-in Nivo]) or DCF (cohorts C and D [adding run-in Nivo]) or neoadjuvant FLOT (cohort E) for resectable locally advanced ESCC. The eligibility criteria were histologically proven ESCC staged as cT1N1-3M0 or cT2-3N0-3M0 (8th UICC TNM classification), age 20-75 years, performance status (PS) ≤1, and no prior cancer treatment. Secondary efficacy endpoints were pathological complete response (pCR), progression-free survival (PFS), and overall survival (OS) for each treatment. Results Thirty-seven patients were enrolled (median age [range]: 62 [31-75] years; PS 0/1: 32/5; clinical stage I/II/III/IVA: 7/9/20/1; PD-L1 TPS <1%/≥1%/unknown: 10/26/1). In CF-based treatment cohorts (n=13), median follow-up time (range), pCR, and 3-year PFS were 42.3 (20.9-50.1) months, 7.7%, and 66.6%, respectively. The respective values for the DCF-based treatment cohorts (n=12) were 32.2 (8.0-39.8) months, 33.3%, and 83.3% and for the FLOT-based treatment cohort (n=12) were 16.2 (5.3-20.6) months, 41.7%, and 91.7%. In the overall population (n=37), median follow-up time was 31.9 (5.3-50.1) months and 3-year OS was 82.4%. Conclusion Neoadjuvant CF or DCF or neoadjuvant FLOT + Nivo followed by surgery for resectable locally advanced ESCC showed promising short- and long-term efficacy. (NCT03914443)