PD-L1 Expression in Melanoma: A Quantitative Immunohistochemical Antibody Comparison

Abstract

Purpose: PD-L1 expression in the pretreatment tumor microenvironment enriches for response to anti-PD-1/PD-L1 therapies. The purpose of this study was to quantitatively compare the performance of five monoclonal anti-PD-L1 antibodies used in recent landmark publications.Experimental Design: PD-L1 IHC was performed on 34 formalin-fixed paraffin-embedded archival melanoma samples using the 5H1, SP142, 28-8, 22C3, and SP263 clones. The percentage of total cells (including melanocytes and immune cells) demonstrating cell surface PD-L1 staining, as well as intensity measurements/H-scores, were assessed for each melanoma specimen using a computer-assisted platform. Staining properties were compared between antibodies.Results: Strong correlations were observed between the percentage of PD-L1(+) cells across all clones studied (R² = 0.81-0.96). When present, discordant results were attributable to geographic heterogeneity of the melanoma tissue section rather than differences in PD-L1 antibody staining characteristics. PD-L1 intensity/H-scores strongly correlated with percentage of PD-L1(+) cells (R² > 0.78, all clones).Conclusions: The 5H1, SP142, 28-8, 22C3, and SP263 clones all demonstrated similar performance characteristics when used in a standardized IHC assay on melanoma specimens. Reported differences in PD-L1 IHC assays using these antibodies are thus most likely due to assay characteristics beyond the antibody itself. Our findings also argue against the inclusion of an intensity/H-score in chromogenic PD-L1 IHC assays. Clin Cancer Res; 23(16); 4938-44. ©2017 AACR.

Figure 1.. Representative photomicrographs of PD-L1 immunohistochemistry performed on a cutaneous melanoma metastasis using clones 5H1, SP142, 28-8, SP263, and 22C3.

All five antibodies showed similar regional patterns of PD-L1 expression by tumor cells, macrophages, and lymphocytes. A degree of geographic heterogeneity between the different tissue sections is evident. Qualitative differences in staining with regard to cytoplasmic or membranous staining or non-specific background staining were not observed. Original magnification, 200x, all fields.

Figure 2.. Comparison of anti-PD-L1 antibody clones in melanoma showing percentage of total PD-L1 (+) cells scored by automated image analysis.

(A) Distribution of PD-L1 expression across antibodies for all melanoma specimens. (B) Representative plot showing percentage of total cells in 34 archival melanoma specimens demonstrating membranous PD-L1 staining with clones SP142 and 28-8. Each dot represents a single specimen. A strong correlation was observed, R²=0.938. (C) Pearson’s correlation coefficients for comparisons of percentage of total cells staining for PD-L1 between all studied clones.

Figure 3.. Comparison of percentage of total PD-L1(+) cells and intensity/H-score using automated image analysis.

Strong correlations were observed between the percentage of cells demonstrating membranous PD-L1 expression and the intensity/H-score across all clones studied, arguing against the inclusion of intensity measurements as an independently scored parameter in PD-L1 IHC assays.