PD-L1 Expression by RNA-Sequencing in Non-Small Cell Lung Cancer: Concordance with Immunohistochemistry and Associations with Pembrolizumab Treatment Outcomes

SIMPLE SUMMARY: We compared RNA next-generation sequencing (RNA-seq) to standard immunohistochemistry (IHC) for PD-L1 expression measurement and associations with pembrolizumab immunotherapy outcomes in NSCLC patient tumors. RNA-seq and IHC PD-L1 score interpretation agreed for 80% of patients, and an RNA-seq “high” cutoff that accurately separated IHC high versus low or negative expression was identified. However, RNA-seq could not discern PD-L1 IHC from negative expression due to the limited sensitivity of IHC as a reference test. High PD-L1 expression by RNA-seq alone and in combination with IHC high or low status was associated with better pembrolizumab outcomes in NSCLC patients than IHC alone. ABSTRACT: Programmed cell death ligand (PD-L1) expression by immunohistochemistry (IHC) lacks sensitivity for pembrolizumab immunotherapy selection in non-small cell lung cancer (NSCLC), particularly for tumors with low expression. We retrospectively evaluated transcriptomic PD-L1 by mRNA next-generation sequencing (RNA-seq). In an unselected NSCLC patient cohort (n = 3168) tested during standard care (2017–2021), PD-L1 IHC and RNA-seq demonstrated moderate concordance, with 80% agreement overall. Most discordant cases were either low or negative for PD-L1 expression by IHC but high by RNA-seq. RNA-seq accurately discriminated PD-L1 IHC high from low tumors by receiver operator curve (ROC) analysis but could not distinguish PD-L1 IHC low from negative tumors. In a separate pembrolizumab monotherapy cohort (n = 102), NSCLC tumors classified as PD-L1 high versus not high by RNA-seq had significantly improved response, progression-free survival, and overall survival as an individual measure and in combination with IHC high or low status. PD-L1 IHC status (high or low) trended toward but had no significant associations with improved outcomes. Conventional PD-L1 IHC testing has inherent limitations, making it an imperfect reference standard for evaluating novel testing technologies. RNA-seq offers an objective PD-L1 measure that could represent a complementary method to IHC to improve NSCLC patient selection for immunotherapy.