Implementation of PD-L1 22C3 IHC pharmDx^TM in Cell Block Preparations of Lung Cancer: Concordance with Surgical Resections and Technical Validation of CytoLyt® Prefixation

Abstract

Background: Programmed death ligand-1 (PD-L1) assessed by immunohistochemistry (IHC) is used as biomarker for pembrolizumab therapy in advanced stage lung cancer patients. However, data permitting direct performance comparison between cytology and surgical specimen types are limited since both specimens from a single tumor site are infrequently available. In addition, alcohol fixation used with cytology specimens requires technical validation of the PD-L1 IHC assay before clinical use. We here report our experience with implementation of the PD-L1 22C3 IHC pharmDxTM assay for cytologic samples at a large tertiary cancer center. Study Design: Archival formalin-fixed (FF), paraffin-embedded cell blocks (CBs) and subsequent lung tumor resections (LTRs) from the same anatomical site were used for a direct comparison of PD-L1 tumor proportion scores (TPSs). TPS values were independently determined by one surgical lung pathologist and two cytopathologists blinded to the specimen pairs. An interim analysis was performed to facilitate the pooling of expertise among observers. After PD-L1 22C3 IHC pharmDxTM implementation for FF cytology specimens, dual-processed samples were used for a prospective technical validation of CytoLyt® prefixation (CF). Digital image analysis was performed for a subset of dual-processed specimens. Results: Eighty-one CBs and LTRs were included for comparison of the specimen types. PD-L1 assessment in CBs had an accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of 88.9/72.8, 66.7/73.5, 95.2/72.3, 80.0/65.8, and 90.9/79.1% for the ≥50/≥1% cutoff, respectively. The intraclass correlation coefficient was 0.84 (95% confidence interval [CI]: 0.76, 0.90), and it improved after interim analysis (before: 0.79 and after: 0.92). The overall concordance between CF and FF for the categories defined by the ≥50/≥1% cutoff values was 90.4% (95% CI: 79.0, 96.8). Similar assay performance was confirmed by digital analysis. Conclusions: PD-L1 22C3 IHC pharmDxTM shows good reliability if used with CB preparations. CF does not impact assay results significantly. Clinical validation with outcome data is needed, and digital methods of assessment should be further investigated.

PD−L1 immunostaining: what pathologists need to know

“…However, several subsequent studies using paired histology and cytology specimens were published. These studies showed that adequately cellular cytological cell blocks (more than 100 tumor cells) may be suitable for PD-L1 testing and can provide equally reliable results [ 21 , 22 ]. This avenue may be advantageous when histologic material is not feasible due to the patient’s condition or other circumstances.…”

PD-L1 Testing in Cytological Non-Small Cell Lung Cancer Specimens: A Comparison with Biopsies and Review of the Literature

“…In studies with cytology fixed in formalin, that is, excluding studies with nonformalin or mixed formalin/nonformalin fixation, the concordance with paired histological specimens was 81–82% (333–336/410 cases) at cutoff 1% and 88–89% (316–317/358 cases) at cutoff 50% based on 7 studies [38, 39, 48, 50, 58, 60, 62]. If instead, only including studies with cytology fixed in nonformalin fixatives, excluding mixed formalin/nonformalin fixation, the concordance with paired histological specimens was 79–81% (500–512/633 cases) at cutoff 1% and 88–89% (554–565/633 cases) at cutoff 50% based on 9 studies including the Lund cohort from the present study [41, 45, 46, 49, 52, 55, 56, 62].…”

“…However, the compiled literature data did not show any obvious difference between formalin and nonformalin fixation. Also, Lou et al [60] presented a very good concordance for formalin-fixed cell blocks with or without prefixation with CytoLyt® [60], while a perfect concordance was seen in the study by Gosney et al [65] with EBUS cytology fixed in formalin versus alcohol. Fixation in, for example, CytoLyt®, is very rapid, and it has been suggested that <1-h fixation has no effect on IHC staining (personal communication).…”

Lung cancer biopsies: Comparison between simple 22G, 22G upgraded and 21G needle for EBUS-TBNA

“…In advanced stage disease molecular biomarkers such epidermal growth factor (EGFR), anaplastic lymphoma kinase (ALK), proto-oncogene B-Raf (BRAF), proto-oncogene tyrosine-protein kinase (ROS1) and programmed death-ligand 1 (PD-L1), provide us with useful information regarding the treatment of the patient [8][9][10][11]. PD-L1 can be efficiently assessed with cellblocks [11][12][13]. However; an important issue still remains for the sample size obtained from the EBUS-TBNA system.…”