Abstract
IntroductionPatient suitability to anti–programmed death ligand 1 (PD-L1) immune checkpoint inhibition is key to the treatment of NSCLC. We present, applied to PD-L1 testing: a comprehensive cross-validation of two immunohistochemistry (IHC) clones; our descriptive experience in diagnostic reflex testing; the concordance of IHC to in situ RNA (RNA-ISH); and application of digital pathology. MethodsEight hundred thirteen NSCLC tumor samples collected from 564 diagnostic samples were analyzed prospectively, and 249 diagnostic samples analyzed retrospectively in tissue microarray format. Validated methods for IHC and RNA-ISH were tested in tissue microarrays and full sections and the QuPath system were used for digital pathology analysis. ResultsAntibody concordance of clones SP263 and 22C3 validation was 97% to 98% in squamous cell carcinoma and adenocarcinomas, respectively. Clinical NSCLC cases were reported as PD-L1–negative (48%), 1% to 49% (23%), and more than 50% (29%), with differences associated to tissue-type and EGFR status. Comparison of IHC and RNA-ISH was highly concordant in both subgroups. Comparison of digital assessment versus manual assessment was highly concordant. Discrepancies were mostly around the 1% clinical threshold. Challenging IHC interpretation included 1) calculating the total tumor cell denominator and the nature of PD-L1 expressing cell aggregates in cytology samples; 2) peritumoral expression of positive immune cells; 3) calculation of positive tumor percentages around clinical thresholds; and 4) relevance of the 100 malignant cell rule. ConclusionsSample type and EGFR status dictate differences in the expected percentage of PD-L1 expression. Analysis of PD-L1 is challenging, and interpretative guidelines are discussed. PD-L1 evaluations by RNA-ISH and digital pathology appear reliable, particularly in adenocarcinomas.
Highlights
Patient suitability to anti–programmed death ligand 1 (PD-L1) immune checkpoint inhibition is key to the treatment of NSCLC
Clinical NSCLC cases were reported as PD-L1–negative (48%), 1% to 49% (23%), and more than 50% (29%), with differences associated to tissue-type and EGFR status
Hereby we present a comprehensive assessment of the PD-L1 IHC diagnostic test in NSCLC at different levels: (1) a comparative cross-validation of the two more widely used antibodies, namely the Dako 22C3 and Ventana SP263 clones; (2) a description of clinical PD-L1 reflex testing in the first 564 cases in an accredited laboratory; (3) the concordance of PD-L1 overexpression by IHC versus PD-L1 upregulation by in situ RNA (RNA-ISH); and (4) the potential role of digital pathology in the automated scoring using open source available software (QuPath)
Summary
Patient suitability to anti–programmed death ligand 1 (PD-L1) immune checkpoint inhibition is key to the treatment of NSCLC. Immune checkpoint blockade therapy is a new paradigm in cancer treatment with durable tumor regression and prolonged stabilization of disease in patients with advanced cancers, including NSCLC.[1,2] The binding of programmed death-1 receptor (PD-1) to its ligand programmed death ligand 1 (PD-L1) plays a pivotal role in the ability of tumor cells to evade the host’s immune system.[3] PD-L1 may be highly expressed on the surface of tumor cells. Along with PD-L1 expression, other markers such as microsatellite instability and tumor mutation burden, as well as understanding the role of other markers and the function of tumor infiltrating immune and stromal cells in the tumor microenvironment may be important in predicting clinical benefit of immune checkpoint inhibition and determining optimal combination therapies.[5,6,7,8,9,10]
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