PURPOSE Four programmed death ligand 1 (PD-L1) immunohistochemistry assays (28-8, 22C3, SP263, and SP142) have been approved for use by the US Food and Drug Administration (FDA). Analytical concordance between these assays has been evaluated in multiple studies. This systematic review included studies that investigated the analytical concordance of immunohistochemistry assays utilizing two or more PD-L1 antibodies from FDA-approved diagnostics for evaluation of PD-L1 expression on tumor or immune cells across a range of tumor types and algorithms. METHODS Literature searches were conducted in MEDLINE (via PubMed) and EMBASE to identify studies published between January 1, 2010, and March 31, 2019, that evaluated analytical concordance between two or more assays based on antibodies from FDA-approved assays. Proceedings of key oncology and pathology congresses that took place between January 2016 and March 2019 were searched for abstracts of studies evaluating PD-L1 assay concordance. RESULTS A total of 42 studies across a range of tumor types met the selection criteria. Concordance between 28-8-, 22C3-, and SP263-based assays in lung cancer, urothelial carcinoma, and squamous cell carcinoma of the head and neck was high when used to assess PD-L1 expression on tumor cells (TCs). SP142-based assays had overall low concordance with other approved assays when used to assess PD-L1 expression on TCs. Analytical concordance for assessment of PD-L1 expression on immune cells was variable and generally lower than for PD-L1 expression on TCs. CONCLUSION A large body of evidence supports the potential interchangeability of 28-8-, 22C3-, and SP263-based assays for the assessment of PD-L1 expression on TCs in lung cancer. Further studies are required in tumor types for which less evidence is available.

Abstract Background: Programmed death-1/programmed death ligand 1 (PD-[L]1) inhibitors are approved for use in a range of cancers. PD-L1 expression in the tumor microenvironment, assessed with an FDA-approved PD-L1 immunohistochemistry (IHC) diagnostic assay such as the Dako PD-L1 IHC 28-8 and 22C3 pharmDx or Ventana PD-L1 SP142 and SP263 assays, is associated with improved PD-(L)1 inhibitor treatment outcomes in some tumor types, including breast cancer (BC). In March 2019, the FDA approved atezolizumab + nab-paclitaxel for the treatment of patients with advanced triple-negative BC and immune cell (IC) PD-L1 expression ≥ 1% using the SP142 assay. Here, we investigated test utilization, test turnaround time (TAT), PD-L1 expression prevalence by assay and biopsy location, and analytical concordance between assays in real-world BC samples. Design: The study included samples from patients with BC that were tested for PD-L1 expression between Oct 2015 and Sep 2019 at NeoGenomics Laboratories, a US national reference laboratory. Patient characteristics from Symphony Healthcare Solutions were matched to PD-L1 test results using unique identifiers. Test volume and TAT were assessed for the 28-8, 22C3, SP142, and SP263 assays. PD-L1 expression was determined by trained pathologists using the 28-8, 22C3, or SP142 assays. Results for the 28-8 assay for the entire study period and for the 22C3 assay until Dec 2018 were reported as the percentage of tumor cells (% TC) with PD-L1 expression. From Jan 2019 onwards, 22C3 assay results were reported as a combined positive score (CPS). All SP142 assay results were reported as the percentage of ICs (% IC) with PD-L1 expression. Analytical concordance between assays was assessed in patients with matched samples (biopsies from the same site and collected on the same date). BioStat Solutions performed statistical analyses. Results: 2955 PD-L1 tests were performed on samples from 2508 patients with BC. The volume of PD-L1 tests on BC samples increased > 100-fold over the study period. Mean TAT was < 5 days for all 4 assays pooled. Table 1 shows PD-L1 expression prevalence in patients with a 28-8, 22C3, or SP142 test result. Median PD-L1 expression did not differ between primary tumors and metastatic sites. In matched samples, overall percentage agreement (OPA) between the 28-8 (TC ≥ 1%) and 22C3 (CPS ≥ 1) assays was 94%, and OPA between the 22C3 (CPS ≥ 1) and SP142 (IC ≥ 1%) assays was 64% (Table 2). Analytical concordance between the 28-8 and 22C3 assays for % TC scoring in matched samples from 27 patients was high (Kendall’s tau = 0.997 [95% CI, 0.883-1.000]). Conclusion: Mean PD-L1 test TAT for BC samples remained < 5 days across all tests despite a large increase in test volume over the study period. Prevalence of PD-L1 expression ≥ 1(%) was higher with the CPS and % IC algorithms than the % TC algorithm, although differences could be due to multiple confounding factors. Despite a small sample size, analytical concordance between the 28-8 and 22C3 assays in matched samples was high. These data provide real-world context for the PD-L1 testing landscape in BC. Table 1. Prevalence of PD-L1 expression in patients with BCPD-L1 expressiona28-8 and 22C3b22C3cSP142% TC, n (%)CPS, n (%)% IC, n (%)(N = 608)(N = 609)(N = 1080)<1(%)390 (64)253 (42)367 (34)≥1(%)218 (36)356 (58)713 (66)All patients had a single test result or ≥ 2 identical results. aThe CPS algorithm is reported on a scale of 0-100, not as a percentage; bSamples tested with the 22C3 assay between Q4 2015 and Q4 2018 were scored using the % TC algorithm; cSamples tested with the 22C3 assay between Q1 2019 and Q4 2019 were scored using the CPS algorithm. Table 2. Agreement between assays on matched samples from patients with BCAgreement between 28-8 (TC ≥ 1%) and 22C3 (CPS ≥ 1) (N = 18)a28-8 as reference22C3 as referenceOPA (n/N)94 (17/18)PPA (n/N)100 (6/6)86 (6/7)NPA (n/N)92 (11/12)100 (11/11)Agreement between 22C3 (CPS ≥ 1) and SP142 (IC ≥ 1%) (N = 33)b22C3 as referenceSP142 as referenceOPA (n/N)64 (21/33)PPA (n/N)86 (12/14)55 (12/22)NPA (n/N)47 (9/19)82 (9/11)aData are presented for 28-8 and 22C3 tests on matched samples with 22C3 tests performed between Q1 2019 and Q4 2019; bData are presented for 22C3 and SP142 tests on matched samples with 22C3 tests performed between Q1 2019 and Q4 2019. N, total number of samples; n, number of samples with the same results with the 2 tests; NPA, negative percentage agreement; PPA, positive percentage agreement. Citation Format: Shreya Mitra, Emily A. Prince, James Pratt, James Novotny, Jr, Vladislav Chizhevsky, Josette William Ragheb, David Huron. Real-world PD-L1 test utilization and analytical concordance of the PD-L1 IHC 28-8 and 22C3 assays in patients with breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS4-18.

Abstract Background: Programmed death-1/programmed death ligand 1 (PD-1/PD-L1) inhibitors are approved in a range of tumor types, including non-small cell lung cancer, with PD-L1 immunohistochemistry (IHC) diagnostic assays approved to inform treatment in some settings. There is evidence that PD-L1 expression can vary between primary tumors and metastatic sites, but the relationship remains unclear. In this real-world study, we compared PD-L1 expression between matched primary tumor and metastatic site biopsies in patients with lung cancer. Methods: NeoGenomics Laboratories Inc (Fort Myers, FL), a US national reference laboratory, provided results for PD-L1 tests performed on samples from 21,224 patients with lung cancer between Oct 2015 and Mar 2018. Test results were linked to clinical characteristics provided by Symphony Healthcare Solutions using unique identifiers. PD-L1 tests were performed using the Dako PD-L1 IHC 28-8 or 22C3 pharmDx assays according to the manufacturers' protocols at the time. The percentage of tumor cells (TCs) expressing PD-L1 was determined by trained pathologists. Patients were included in the analysis if they had matched biopsies from a primary lung tumor and a metastatic site that were collected in any order within a 3-month period, and if both samples were tested with the same PD-L1 assay ≤ 3 months apart. Patients were excluded if they received treatment between biopsies or had > 2 biopsies. Statistical analysis was performed by BioStat Solutions Inc. Results: In total, 121 patients had matched primary and metastatic biopsy samples, with sites biopsied in any order; a subgroup of 59 patients had their second biopsy obtained after the PD-L1 test result for the first biopsy was reported. Matched biopsy pairs showed modest concordance (Kendall's tau 0.43 [95% CI, 0.33–0.54]; Spearman's correlation 0.56 [95% CI, 0.42–0.67]). Overall percentage agreement was 69–80% (Cohen's kappa 0.34–0.53) across a range of PD-L1 expression cutoffs (1%, 5%, 10%, 25%, and 50% of TCs). Identical PD-L1 expression was observed in 26% of matched biopsy pairs; 44% of sample pairs had a < 5% difference and 35% of sample pairs had a > 20% difference in PD-L1 expression scores between primary and metastatic sites. PD-L1 expression in primary tumor and metastatic sites was heterogeneous, with no clear trends across biopsy sites. In the subgroup of 59 patients whose second biopsy was obtained after the test result for their first biopsy was reported, 50% of patients (15/30) with PD-L1 expression on < 1% of TCs in their first biopsy had PD-L1 expression on ≥ 1% of TCs in their second biopsy. Conclusion: This real-world study suggests that agreement of PD-L1 expression between matched primary and metastatic biopsy sites is low, further highlighting PD-L1 expression heterogeneity in lung cancer. Variation in PD-L1 expression between biopsy sites may affect treatment decisions relating to PD-1/PD-L1 inhibitors. Citation Format: Emily A. Prince, Vladislav Chizhevsky, Josette William Ragheb, James L. Pratt, Dimple Pandya, David Huron. Comparison of PD-L1 expression in primary and metastatic lung cancer biopsies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2004.

AimsProgrammed death-1/programmed death ligand 1 (PD-1/PD-L1) inhibitor therapy is accompanied by companion or complementary PD-L1 testing in some tumour types. We investigated utilisation of the Dako PD-L1 IHC 28-8 and 22C3 pharmDx assays and the Ventana PD-L1 (SP142) assay and evaluated concordance between the 28-8 and 22C3 assays in a real-world cohort of patients tested at a single US national reference laboratory.MethodsNeoGenomics Laboratories performed PD-L1 testing on tumour samples between October 2015 and March 2018. PD-L1 test results were matched with patient characteristics using unique identifiers. Concordance between the 28-8 and 22C3 assays was evaluated in matched tumour samples. Data were evaluated across multiple tumour types and in subgroups of patients with lung cancer, melanoma, squamous cell carcinoma of the head and neck, and urothelial carcinoma.Results62 180 individual PD-L1 tests were conducted on samples from 55 652 patients. PD-L1 test volume increased ~10-fold over the period evaluated. Test failure rates were typically low, and test turnaround time (TAT) ranged between 2 and 4 days. Concordance between the 28-8 and 22C3 assays was strong in the overall population and across tumour type subgroups (Kendall’s tau correlations of 0.94 and 0.92–0.98, respectively).ConclusionsTest failure rates for PD-L1 tests were low and TAT remained reasonable despite marked increases in test volume. Concordance was high between the 28-8 and 22C3 assays across a range of tumour types and biopsy locations. These findings add to the literature showing high concordance between the 28-8 and 22C3 assays.