P11.17.B PD-L1 DETECTION IN CEREBROSPINAL FLUID OF LEPTOMENINGEAL METASTASIS BASED ON THINPREP CYTOLOGY

Abstract BACKGROUND Leptomeningeal metastasis (LM) involves tumor cell dissemination within the leptomeninges and subarachnoid space. Intrathecal anti-programmed death 1 immunotherapy has been explored clinically for LM. Programmed cell death ligand 1 (PD-L1) is currently the only approved immunotherapy marker. This study aims to detect PD-L1 expression in cerebrospinal fluid (CSF) from LM patients based on ThinPrep cytology method. MATERIAL AND METHODS Cell lines (lung adenocarcinoma: NCI-H358, A549; breast cancer: MDA-MB-231, MCF7; gastric adenocarcinoma: SGC-7901, AGS) with confirmed high/low PD-L1 expression were processed via ThinPrep liquid-based cytology and cell-block sections. CSF tumor cells from LM patients underwent ThinPrep slide preparation. Immunocytochemical analysis with PD-L1 antibodies (Ventana SP263 and Dako 22C3) was performed. Five independent observers assessed PD-L1 expression using the tumor proportion score (TPS), defined as the percentage of viable tumor cells showing membrane staining among ≥100 evaluable cells (positive threshold: TPS ≥1%) with absolute cell counts recorded for specimens containing <100 tumor cells, and inter-assay concordance was determined using Cohen’s κ statistics. RESULTS Cell lines with high PD-L1 gene expression (NCI-H358, MDA-MB-231, SGC-7901) demonstrated strong membranous PD-L1 protein expression (TPS range: 94-100%) by both 22C3 and SP263 assays. In contrast, low-expressing lines (A549, MCF7, AGS) showed minimal staining (TPS <1%) with either antibody. PD-L1 expression showed excellent concordance between ThinPrep slides and cell-block sections. In clinical specimens, analysis of 88 CSF samples from 44 LM patients (37 lung adenocarcinomas, 5 breast cancers, and 2 gastric adenocarcinomas) revealed PD-L1 positivity rates of 45% (20/44) with 22C3 and 52% (23/44) with SP263, demonstrating a high inter-assay concordance (Cohen κ=0.86). Among 44 slides with <100 tumor cells, PD-L1 expression was as follows: 27 within TPS <1% (14 for 22C3 and 13 for SP263), 15 within TPS 1-49% (7 for 22C3 and 8 for SP263), and 2 within TPS ≥50% (1 for 22C3 and 1 for SP263). All CSF specimens from breast cancer and gastric adenocarcinoma patients showed negative PD-L1 expression. Among lung adenocarcinoma cases, the assays demonstrated Cohen κ of 0.84, with positivity rates of 54% (20/37) for 22C3 and 62% (23/37) for SP263. CONCLUSION Immunocytochemical analysis of PD-L1 expression in CSF specimens from LM patients using ThinPrep cytology demonstrated feasibility, showing good concordance between the Ventana SP263 and Dako 22C3 assays.