Comparative analyses of tumor immune microenvironment using paired samples from primary non-small cell lung cancer and brain metastases

Abstract The efficacy of immune checkpoint inhibitors against brain metastases (BMs) from non-small cell lung cancer (NSCLC) often appears limited, potentially due to differences in the tumor immune microenvironment (TIME) between primary tumors (PTs) and BMs. We aimed to characterize the TIME of paired PTs and BMs and to investigate the impact of metastatic timing (synchronous BMs [sBMs] vs. metachronous BMs [mBMs]). We retrospectively analyzed 14 paired samples (7 sBMs, 7 mBMs) using immunohistochemistry for CD163, T-cell subsets (CD3, CD8, programmed cell death 1 [PD-1], forkhead box protein 3 [FoxP3]), programmed cell death ligand 1 (PD-L1), and peripheral lymph node addressin (PNAd). The densities of T-cell subsets and proportions of CD163-positive areas were quantitatively evaluated in intratumoral and stromal compartments using digital pathology. PD-L1 expression was assessed as the tumor proportion score. Compared to PTs, BMs exhibited a more immunosuppressive TIME, with a significantly larger CD163-positive area, lower densities of intratumoral T cells (CD3+, CD8+, PD-1+), and complete absence of PNAd + tumor-associated high-endothelial venules. PD-L1 expression was well-conserved between sites. Critically, TIMEs differed by metastatic timing. PTs of the sBM group showed higher intratumoral and stromal FoxP3/CD3 and lower stromal CD8/CD3 compared to the mBM group. Subsequently, sBMs displayed a distinct TIME with higher intratumoral CD8/CD3 and PD-1/CD3 than mBMs. BMs from NSCLC thus appear to harbor a distinct, immunosuppressive TIME. The timing of metastasis appears fundamentally associated with this TIME profile, with sBM groups showing a unique immunosuppressive signature originating from the PT. These findings provide a rationale for stratifying patients with BMs to guide personalized immunotherapy.