CLDN18/CLDN18.2 IHC assay comparison (SP455, 43-14A, EPR19202) and co-prevalence expression with other biomarkers in gastric carcinoma.

4089 Background: Despite recent advances in treatment, gastric cancer (GC) remains the fourth leading cause of cancer-related mortality worldwide. Claudin 18.2 (CLDN18.2) has emerged as a promising therapeutic target as its expression is mostly restricted to the gastric epithelium and persists in a significant fraction of GC and other upper gastrointestinal tumors (e.g. pancreatic ductal adenocarcinoma). The goal of this study was to compare the analytical performance of multiple CLDN18/CLDN18.2 IHC assays and understand the prevalence of CLDN18.2 with other GC-relevant biomarkers (PD-L1, HER2) and the tumor immune microenvironment. Methods: Cell lines (n=11) and primary resection specimens from patients with GC (n=91) were studied. To compare the analytical performance of three CLDN18/CLDN18.2 IHC assays, FFPE samples were stained with pan-CLDN18 (43-14A) and CLDN18.2-specific (SP455 and EPR19202) clones. Staining results were validated using orthogonal methods i.e., flow cytometry and mass spectrometry. The relationship of CLDN18.2 expression with PD-L1 (SP263) and HER2 (HercepTest) as well as T cell contexture (CD3/CD8/PanCK; CD3/PD-L1/Ki67) were explored by IHC and chromogenic multiplex IHC. Association of these markers with clinicopathological features was also investigated. Results: Comparison of the three CLDN18/CLDN18.2 IHC assays showed similar analytical performance across cell lines and GC samples. Using orthogonal methods, we determined that the limit of detection and linear range was comparable amongst assays. In GC, samples classified as CLDN18/CLDN18.2 positive at a cutoff of 75% with 2-3+ intensity included 22% (SP455), 22% (43-14A), and 15% (EPR19202) of samples. Classification at other CLDN18/CLDN18.2 cutoffs were also explored and will be presented. Only one case (1%) showed significantly discrepant staining patterns between pan-CLDN18 and CLDN18.2-specific assays (H score 235 vs. 105, respectively). CLDN18.2 positivity did not correlate with HER2 expression, PD-L1 expression, T cell densities (cells/mm2), or any of the tested clinicopathological characteristics i.e., tumor grade, histologic subtype, or TNM stage. Conclusions: We demonstrated a concordant analytical performance between the tested CLDN18/CLDN18.2 IHC assays in GC. Their implementation in the clinical setting might help identify patient candidates that could benefit from treatment with CLDN18.2 targeted therapies. Additional research is needed to confirm similar analytical performance between the three IHC assays in other indications.