Immune Cell Infiltration in Primary and Recurrent Glioblastoma, IDH‐Wild Type: Validation of an Immunohistochemistry‐Based Scoring System for Research and Clinical Practice

Significant heterogeneity in the immune microenvironment of Glioblastoma, IDH‐wildtype (GBM), has been reported, necessitating a standardized approach to evaluate immune infiltration in the context of emerging immunotherapies. To address this, we developed and validated a standardized immunohistochemistry‐based scoring system for quantifying immune cell infiltration in formalin‐fixed, paraffin‐embedded (FFPE) tissue. Paired primary and recurrent GBM specimens from 20 adult patients were labeled for CD3, CD8, CD45, CD68, and PD‐1, and scored across six anatomical regions, including intra‐ and peritumoral, meningeal, and normal brain areas. The scoring system demonstrated excellent interrater (ICC = 0.932) and intrarater (ICC = 0.953) reliability. CD68+ and CD45+ cells were most numerous across all regions. CD3+ and CD8+ cells concentrated more in the perivascular area rather than within the parenchyma. No significant differences in immune infiltration were observed between primary and recurrent GBM. Cox proportional‐hazards analysis showed worse survival with higher CD8+ and CD45+ infiltration in primary GBM, and higher CD45+ and CD68+ infiltration in recurrent GBM. In conclusion, we propose a feasible, cost‐efficient, and robust method to assess immune infiltration on FFPE material, enabling standardized comparison of inflammation, with applications for ongoing clinical trials.