Background: Results from the phase III Prep-02/JSAP-05 trial demonstrated the efficacy and safety of gemcitabine plus S-1 as neoadjuvant chemotherapy for resectable pancreatic cancer. However, consensus on neoadjuvant chemotherapy for pancreatic cancer has not yet been established, and the effects of neoadjuvant therapy on clinical features of the tumor and improvement in prognosis have not been fully investigated. We aimed to investigate the effect of neoadjuvant chemotherapy on resectable pancreatic cancer. Methods: Between 2013 and 2021, 291 patients who underwent curative resection for resectable pancreatic cancer at our hospital were enrolled. The patients were categorized into groups of those who received neoadjuvant chemotherapy (NAC group: n = 91) and those who received up-front surgery (US group: n = 200). Prognostic predictors were examined in (i) all patients, (ii) US group, and (iii) NAC group. Statistical significance was considered at p < 0.05. Results: The median follow-up periods were 20.7 and 33.2 months in the NAC and US groups, respectively. In the NAC group, 84 of the 91 patients (94.5%) received gemcitabine-based chemotherapy, and three (3.3%) received chemoradiotherapy. There was no difference in overall survival (OS) and recurrence free survival (RFS) between the two groups. In the background, the NAC group had lower preoperative CA19-9 levels, lower lymph node ratio (LNR), and smaller tumor size than in the US group. During multivariable analysis of OS, preoperative CA19-9 levels, LNR, modified Glasgow prognostic score (mGPS), resection margin, and tumor size were identified as possible prognostic predictors in all patients and in the US group, whereas LNR and mGPS were identified in the NAC group. Conclusions: The prognostic significance of preoperative CA19-9 level, tumor size, and LNR were counteracted in the NAC group. These findings may indicate the effect of neoadjuvant chemotherapy for resectable pancreatic cancer.