Validation of PD-L1 clone 22C3 immunohistochemical stain on two Ventana DISCOVERY autostainer models: detailed protocols, test performance characteristics, and interobserver reliability analyses

ABSTRACT

Immunohistochemical (IHC) stain for PD-L1 as a biomarker for immunotherapy is recommended in non-small cell lung cancer (NSCLC). Under the FDA, the selection of patients for pembrolizumab requires companion diagnostic testing using the Dako Agilent PD-L1 IHC 22C3 pharmDx kit performed on the Dako Autostainer Link 48 platform. However, because it is not widely available, there is need for cross-platform validation. Existing studies provide incomplete protocol detail. In our study, 73 lung tumors were stained using the FDA-approved test (‘gold standard’). The same blocks were stained using two different models of the Ventana DISCOVERY platform (ULTRA, n = 73 and XT, n = 70) using different parameters, and interpreted by three pathologists. The ULTRA group met College of American Pathologists (CAP) validation criteria (concordance 91.8%) while the XT group did not (concordance 67.1%). Using tumor proportion score (TPS) ≥1% and TPS ≥50% as cut-offs, the ULTRA protocol had higher sensitivity (97.8% and 91.7%) than XT (73.3% and 60.9%) and similar specificity (ULTRA 88.9% and 100%, XT 88% and 100%). Discordance between ULTRA and XT was 27%, and in all these cases ULTRA was concordant with gold standard. Interobserver reliability was substantial for ULTRA and almost perfect for XT, providing evidence that staining rather than observer variability accounts for XT’s inferior performance. Cross-validation of the clinically used 22C3 anti PD-L1 antibody test with substantial interobserver agreement is possible on the commonly used the Ventana DISCOVERY ULTRA automated instrument, while the validation failed on the XT. Cautious attention to detail must be paid when choosing cross-validation parameters.

KEYWORDS:

• Validation
• immunohistochemistry
• Ventana Discovery Ultra
• Ventana Discovery XT
• interobserver
• PD-L1
• pembrolizumab
• Keytruda
• Dako
• cross-platform
• harmonization
• lung cancer
• immunotherapy
• 22C3

Acknowledgments

The authors would like to thank Yuhe Xia, MS (NYU Langone Health, Division of Biostatistics, Department of Population Health, New York, NY, USA) for her help with statistics. The authors would also like to acknowledge Briana Zeck for her efforts.

Disclosure statement

Authors have no conflicts of interest.

Supplementary material

Supplemental data for this article can be accessed here.

Additional information

Funding

The NYULH Center for Biospecimen Research and Development, Histology and ImmunohistochemistryLaboratory (RRID:SCR_018304) is supported in part by the Laura and Isaac Perlmutter Cancer Center Support Grant: NIH/NCI P30CA016087 and the National Institutes of Health S10 Grants; NIH/ORIP S10OD01058 and S10018338.