Preanalytical Determinants of Patient Eligibility for Immune Checkpoint Inhibitors Revealed by Quantitative Programmed Cell Death Ligand-1 (PD-L1) Analysis.

PURPOSE: The assessment of programmed cell death ligand-1 (PD-L1) protein expression is a predictive biomarker for subsequent immune checkpoint inhibitor-based therapies. The purpose of this study was to granularly and quantitatively define the impact of preanalytical variables such as cold ischemia time (CIT) and tissue block and slide aging on PD-L1 expression evaluation via immunohistochemistry.

MATERIALS AND

METHODS

Using tonsillar tissue as a standardized reference model, we systematically evaluated the effects of CIT, tissue block aging, tissue slide aging, and fixation strategy on PD-L1 immunohistochemistry expression. PD-L1 staining (clone SP263) was quantified by digital pathology in distinct tissue compartments, including lymphoreticular cells, the crypt epithelium, and the superficial epithelium.

RESULTS

Both increasing CIT and tissue block aging were associated with a rapid and significant reduction in PD-L1 expression, with a relative loss exceeding 52% after 30 hours and 49.2% after 60 months of storage, respectively. Tissue slide aging presented the most pronounced effect: PD-L1 expression declined by 60.1% (surgical samples) and 60.6% (endoscopic biopsies) within 60 days. Cold formalin fixation yielded higher baseline PD-L1 expression and slowed antigen decay compared with standard formalin fixation but did not prevent progressive PD-L1 loss, especially in biopsy samples.

CONCLUSIONS

Using tonsillar tissue as a standardized PD-L1 reference model, we showed that selected preanalytical variables are associated with measurable and progressive loss of PD-L1 immunoreactivity, particularly in lymphoreticular cells. These findings support increased awareness of tissue handling, archival storage, and unstained slide storage as potential sources of analytical variability in PD-L1 assessment.