RESUMEN
OBJECTIVES: The objective is to assess the impact of the quality of tissue fixation in surgical pathology on immunohistochemical (IHC) staining and DNA degradation. MATERIALS AND METHODS: Twenty-five non-small cell lung cancer (NSCLC) resection specimens were analyzed. After resection, all tumors were processed according to the protocols in our center. In haematoxylin and eosin (H&E) stained tissue slides, adequately- and inadequately fixed tumor areas were microscopically demarcated, based on basement membrane detachment. In 10 IHC stains ALK (clone 5A4), PD-L (clone 22C3), CAM5.2, CK7, c-Met, KER-MNF116, NapsinA, p40, ROS1, TTF1) the immunoreactivity in H-scores was determined in adequately- and inadequately fixed, and necrotic tumor areas. From the same areas DNA was isolated, and DNA fragmentation in base pairs (bp) was measured. RESULTS: H-scores were significantly higher in H&E adequately fixed tumor areas in IHC stains KER-MNF116 (H-score 256 vs 15, p=0.001) and p40 (H-score 293 vs 248, p=0.028). All other stains showed a trend towards higher immunoreactivity in H&E adequately fixed areas. Independent of H&E adequatelty- or inadequately fixed areas, all IHC stains showed significant different IHC staining intensity within tumors, suggesting heterogeneous immunoreactivity (H-scores: PD-L1 123 vs 6, p = 0.001; CAM5.2 242 vs 101, p=<0.001; CK7 242 vs 128, p=<0.001; c-MET 99 vs 20, p=<0.001; KER-MNF116 281 vs 120, p=<0.001; Napsin A 268 vs 130, p = 0.005; p40 292 vs 166, p = 0.008; TTF1 199 vs 63, p=<0.001). DNA fragments rarely exceeded a length of 300 bp, independent of adequate fixation. However, DNA fragments of 300 and 400 bp had higher concentrations in tumors with short fixation delay (<6 h vs >16 h) and short fixation time (<24 h vs >24 h). CONCLUSIONS: Impaired tissue fixation of resected lung tumors results in decreased IHC staining intensity in some parts of the tumor. This may impact the reliability of IHC analysis.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/cirugía , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/cirugía , Neoplasias Pulmonares/metabolismo , Proteínas Tirosina Quinasas , Fijación del Tejido/métodos , Reproducibilidad de los Resultados , Inmunohistoquímica , Proteínas Proto-Oncogénicas , Antígeno B7-H1/metabolismo , Biomarcadores de Tumor/genéticaRESUMEN
OBJECTIVES: Programmed death-ligand 1 (PD-L1) is the only approved predictive biomarker for immunotherapy in non-small cell lung cancer (NSCLC). However, predictive PD-L1 immunohistochemistry is subject to interobserver variability. We hypothesized that a pathologist's personality influences the interobserver variability and diagnostic accuracy of PD-L1 immunoscoring. MATERIALS AND METHODS: Seventeen pathologists performed PD-L1 immunoscoring on 50 resected NSCLC tumors in three categories (<1%;1-49%;≥50%). Also, the pathologists completed a certified personality test (NEO-PI-r), assessing five personality traits: neuroticism, extraversion, openness, altruism and conscientiousness. RESULTS: The overall agreement among pathologists for a series of 47 tumors was substantial (kappa = 0.63). Of these, 23/47 (49%) tumors were entirely negative or largely positive, resulting in a kappa value of 0.93. The remaining 24/47 (51%) tumors had a PD-L1 score around the cutoff value, generating a kappa value of 0.32. Pathologists with high scores for conscientiousness (careful, diligent) had the least interobserver variability (r = 0.6, p = 0.009). Also, they showed a trend towards higher sensitivity (74% vs. 68%, p = 0.4), specificity (86% vs. 82%, p = 0.3) and percent agreement (83% vs. 79%, p = 0.3), although not significant. In contrast, pathologists with high scores for neuroticism (sensitive, anxious) had significantly lower specificity (80% vs. 87%, p = 0.03) and percent agreement (78% vs. 85%, p = 0.03). Also, a trend towards high interobserver variability (r = -0.3, p = 0.2) and lower sensitivity (68% vs. 74%, p = 0.3) was observed, although not significant. Pathologists with relatively high scores for conscientiousness scored fewer tumors PD-L1 positive at the ≥ 1% cut-off (r = -0.5, p = 0.03). In contrast, pathologists with relatively high scores for neuroticism score more tumors PD-L1 positive at ≥ 1% (r = 0.6, p = 0.017) and ≥ 50% cut-offs (r = 0.6, p = 0.009). CONCLUSIONS: This study is the first to demonstrate the impact of a pathologist's personality on the interobserver variability and diagnostic accuracy of immunostaining, in the context of PD-L1 in NSCLC. Larger studies are needed for validation of these findings.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Antígeno B7-H1 , Biomarcadores de Tumor , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Carcinoma de Pulmón de Células no Pequeñas/patología , Humanos , Inmunohistoquímica , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/patología , Variaciones Dependientes del Observador , Patólogos , PersonalidadRESUMEN
BACKGROUND: Advanced non-small-cell lung cancer (NSCLC) harbours many genetic aberrations that can be targeted with systemic treatments. Whole-genome sequencing (WGS) can simultaneously detect these (and possibly new) molecular targets. However, the exact added clinical value of WGS is unknown. OBJECTIVE: The objective of this study was to determine the early cost effectiveness of using WGS in diagnostic strategies compared with currently used molecular diagnostics for patients with inoperable stage IIIB,C/IV non-squamous NSCLC from a Dutch healthcare perspective. METHODS: A decision tree represented the diagnostic pathway, and a cohort state transition model represented disease progression. Three diagnostic strategies were modelled: standard of care (SoC) alone, WGS as a diagnostic test, and SoC followed by WGS. Treatment effectiveness was based on a systematic review. Probabilistic cost-effectiveness analyses were performed, and threshold analyses (using 80,000 per quality-adjusted life-year [QALY]) was used to explore the early cost effectiveness of WGS. RESULTS: WGS as a diagnostic test resulted in more QALYs (0.002) and costs (1534 [incremental net monetary benefit -1349]), and SoC followed by WGS resulted in fewer QALYs (-0.002) and more costs (1059 [-1194]) compared with SoC alone. WGS as a diagnostic test was only cost effective if it was priced at 2000 per patient and identified 2.7% more actionable patients than SoC alone. Treating these additional identified patients with new treatments costing >4069 per month decreased the probability of cost effectiveness. CONCLUSIONS: Our analysis suggests that providing WGS as a diagnostic test is cost effective compared with SoC followed by WGS and SoC alone if costs for WGS decrease and additional patients with actionable targets are identified. This cost-effectiveness model can be used to incorporate new findings iteratively and to support ongoing decision making regarding the use of WGS in this rapidly evolving field.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Análisis Costo-Beneficio , Pruebas Diagnósticas de Rutina , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Años de Vida Ajustados por Calidad de VidaRESUMEN
AIMS: Investigate the impact of interlaboratory- and interobserver variability of immunohistochemistry on the assessment of programmed death ligand 1 (PD-L1) in non-small cell lung cancer (NSCLC). METHODS: Two tissue microarrays (TMAs) were constructed from 50 (TMA-A) and 51 (TMA-B) resected NSCLC cases, and distributed among eight centres. Immunostaining for PD-L1 was performed using Agilent's 22C3 pharmDx Assay (pharmDx) and/or a 22C3 laboratory developed test (LDT). The interlaboratory variability of staining- and interobserver variability of scoring for PD-L1 were assessed in selected critical samples (samples at the cut-off of positivity) and non-critical samples. Also, PD-L1 epitope deterioration in time in stored unstained slides was analysed. Krippendorff's alpha values (0=maximal, 1=no variability) were calculated as measure for variability. RESULTS: For interlaboratory variability of immunostaining, the percentage of PD-L1 positive cases among centres ranged 40%-51% (1% cut-off) and 23%-30% (50% cut-off). Alpha values at 1% cut-off were 0.88 (pharmDx) and 0.87 (LDT) and at 50% cut-off 0.82 (pharmDx) and 0.95 (LDT). Interobserver variability of scoring resulted in PD-L1 positive cases ranging 29%-55% (1% cut-off) and 14%-30% (50% cut-off) among pathologists. Alpha values were at 1% cut-off 0.83 (TMA-A) and 0.66 (TMA-B), and at 50% cut-off 0.77 (TMA-A) and 0.78 (TMA-B). Interlaboratory variability of staining was higher (p<0.001) in critical samples than in non-critical samples at 50% cut-off. Furthermore, PD-L1 epitope deterioration in unstained slides was observed after 12 weeks. CONCLUSIONS: The results provide insight in factors contributing to variability of immunohistochemical assessment of PD-L1, and contribute to more reliable predictive testing for PD-L1.