Background: Relationships between cancer onset, progression, and inflammation have been reported; yet, the significance of inflammatory markers before and after treatment with nanoliposomal irinotecan and fluorouracil with folic acid (NFF) is unclear. Objectives: We aimed to (1) investigate whether worsening inflammation is associated with disease progression and whether it can be used as a biomarker by comparing inflammatory markers before and after treatment with NFF and (2) verify which is a superior biomarker. Design: This was a preplanned analysis of a retrospective cohort of the NAPOLEON-2 study, a multicenter observational study. Patients from 20 institutions were enrolled. Those with unresectable pancreatic cancer who received NFF as a second-line or later-line treatment between June 1, 2020, and May 31, 2021, were included. Methods: The primary endpoint was overall survival, and the main secondary endpoint was progression-free survival. The following inflammatory markers were assessed: neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio, Prognostic Nutrition Index (PNI), C-reactive protein/albumin ratio (CAR), Glasgow prognostic score (GPS), Prognostic Index (PI), and Komori score. Results: A total of 161 patients were enrolled. The CAR, NLR, GPS (1 vs 2), PI (1 vs 2), PNI, and Komori score were useful pre-NFF biomarkers (hazard ratios (HRs): 2.59, 2.28, 2.76/3.35, 2.16/18.89, 2.13, and 3.39, respectively). The CAR, NLR, GPS (compared with a score of 1/2), and PI (compared with a score of 1/2) were useful post-NFF biomarkers (HRs: 2.63, 1.88, 2.54/3.09, 2.30/5.39, respectively). In terms of the receiver operating characteristic curve and Akaike information criterion, the CAR, GPS, and PI showed favorable trends, and overall, CAR was a useful biomarker. Conclusion: Inflammatory markers can be used as biomarkers both before and after NFF treatment, and CAR may be useful. We hope these findings will contribute to the selection of appropriate treatment options.