Introduction: Phase I and II trials are currently underway using immune checkpoint inhibitors targeted at programmed cell death protein 1 (PD-1) as an immune therapy for glioblastoma patients. Despite this, no consensus exists as to the best assay to determine whether or not a patient’s tumor has significant expression of programmed death-ligand 1 (PD-L1), indicating that this is a clinically relevant mechanism of immune suppression in patients. Previous reports have disagreed over the true prevalence of PD-L1 expression in brain tumor patients, with multiple different antibodies and staining conditions in use at various centers.
Methods: We have previously published an effective method of staining for PD-L1 in glioblastoma tissues using Abcam’s EPR11661(2) antibody, but the 28-8 clone, currently being used for immunohistochemistry (IHC) in clinical trials, has not been stringently validated in glioma patients. HEK 293 cells were grown in culture and either transfected with a PD-L1 expression plasmid or left untreated. These cells were then analyzed for PD-L1 expression using Western blot with the 28-8 antibody. Placenta control slides were stained by IHC for PD-L1 using both EPR1161(2) and the 28-8 PD-L1 antibodies. Patient tumor samples were then stained using the 28-8 antibody.
Results: 28-8 was able to detect PD-L1 expression in HEK 293 cells transfected with the PD-L1 plasmid. IHC using either EPR1161(2) or 28-8 antibodies showed equivalent positive staining on placenta positive control slides. Our review of the slides revealed 1 patient with an atypical pilocytic astrocytoma having near complete 3+ staining, 1 patient with a glioblastoma with 2+ staining, 3 malignant glioma patients with minimal 1+ staining and another 4 malignant glioma patients demonstrating no PD-L1 positivity, consistent with our previously reported data.
Conclusions: Immunohistochemistry using antibody 28-8 appears to be equivalent to EPR1161(2) in its ability to detect PD-L1 expression in brain tumor patients.
Patient Care: By validating strategies to determine whether tumors express PD-L1, we will be able to better risk-stratify patients into positive and negative cohorts to guide immune therapy going forward.
Learning Objectives: By the conclusion of this session, attendees should be able to:
1) Identify PD-L1 positivity based on immunohistochemistry
2) Describe the importance of risk stratification using expression of immunosuppressive markers in trials for immune therapeutics
3) Accurately anticipate the expected percentage of malignant glioma patients who will express PD-L1
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Nduom EK, Wei J, Yaghi NK, Huang N, Kong LY, Gabrusiewicz K, Ling X, Zhou S, Ivan C, Chen JQ, Burks JK, Fuller GN, Calin GA, Conrad CA, Creasy C, Ritthipichai K, Radvanyi L, Heimberger AB. PD-L1 expression and prognostic impact in glioblastoma. Neuro Oncol. 2016 Feb;18(2):195-205. doi: 10.1093/neuonc/nov172. Epub 2015 Aug 30.