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Objective. This study aimed to investigate the efficacy and safety of anti-PD-1 immunotherapy combined with gemcitabine chemotherapy in multiline treatment of advanced pancreatic cancer. Methods. A retrospective analysis was performed on the clinical data of 32 patients with advanced pancreatic cancer treated with sintilimab regimen from January 2019 to December 2021 in our hospital. All patients were followed up until death or April 2022, in the form of outpatient, in-hospital review, or telephone follow-up. Follow-up content included routine blood, liver and kidney functions, tumor markers, plain or enhanced abdominal CT, and abdominal MRI examinations. Clinical efficacy was evaluated according to mRECIST criteria, and the severity of adverse effects was evaluated according to American Institute for Cancer Research (AICR) Standard Term for Adverse Events, Version 5.0. Results. During treatment, the dosage of sintilimab was halved in 2 patients due to adverse reactions. All patients were treated with sintilimab for 1~10 times, with an average of 6 ± 4 times. The total response rate (ORR) and disease control rate (DCR) were 6.25% and 12.50% and 25.00% and 37.50%, respectively, after 1 and 3 months of treatment. The mean follow-up time of 32 patients was 1-12 months, and the median follow-up time was 4 ± 3 months. By the end point of follow-up, a total of 25 patients died, and the median progression-free survival (PFS) was 3.8 (95% CI (1.85-5.63)) months. The median overall survival (OS) was 5.1 months (95% CI (3.63~7.68). After treatment, the levels of tumor markers CA125, CEA and CA199 were partly decreased compared with those before treatment (all P < 0.001). After treatment, the blood routine indexes d-dimer, CRP (C-reactive protein), NLR (neutral granulocyte to lymphocyte ratio), and MLR (monocyte to lymphocyte ratio) decreased compared with those before treatment. In 32 patients with advanced pancreatic cancer, the adverse reactions with an incidence more than 10% included fatigue, rash, hypothyroidism, hyperuricemia, and renal insufficiency. Only 2 patients showed grade 3 fatigue symptom, and all the others showed no adverse reactions of grades 3~5. In this study, all patients' adverse reactions were relieved after symptomatic treatment. Conclusion. Gemcitabine chemotherapy in multiline treatment of advanced pancreatic cancer with sintilimab can achieve certain clinical benefits without serious adverse reactions. [ABSTRACT FROM AUTHOR]