Decade‐Resolved Proteomic Profiling of Gastric Cancer FFPE Archives: Evaluating Storage‐Associated Shifts and Signal Stability Over 50 Years

ABSTRACT

Archival formalin‐fixed, paraffin‐embedded (FFPE) tissues are invaluable for retrospective clinical research, yet the impact of multi‐decade storage on quantitative proteomic fidelity remains poorly understood. We conducted a systematic, decade‐resolved proteomic assessment using data‐independent acquisition (DIA) LC–MS/MS to evaluate FFPE specimens spanning 50 years. This study included 33 gastric cancer cases collected at approximately 10‐year intervals from 1972 to 2022. From each case, tumor tissue, matched non‐tumorous mucosa, and muscle layer were macrodissected and processed simultaneously using a standardized workflow to minimize technical bias. Longitudinal analysis across five decades demonstrated that a substantial portion of the detectable proteome remains quantitatively stable. While storage‐ and archival‐era‐associated factors contributed to measurable variance, tissue‐specific biological signatures were the dominant drivers of proteomic profiles. The muscle layer exhibited the highest stability, serving as a structural reference for evaluating archival ageing. Differential expression analysis between tumor and nontumorous mucosa identified robust pathological signatures that persisted even in 50‐year‐old specimens. Although we observed era‐specific variations in protein detectability, a core set of differentially expressed proteins and their associated functional pathways—including cell cycle and metabolic remodeling—were consistently preserved across the evaluable year groups. Notably, the treatment‐naïve 1972 cohort provided high‐depth biological insights comparable to contemporary samples. Our findings establish that long‐term archival storage does not preclude biologically interpretable proteomic profiles, although storage‐ and archival‐era‐associated shifts should be explicitly accounted for in analysis and interpretation.