Considerable interlaboratory variation in PD-L1 positivity for head and neck squamous cell carcinoma in the Netherlands- A nationwide evaluation study.

AIMS: Patients with recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) are eligible for first-line immune checkpoint inhibition if their tumour is positive for programmed death ligand 1 (PD-L1) determined by the combined positive score (CPS). This nationwide study, using real-world data, investigated the developing PD-L1 testing landscape in the first 3 years after introduction of the test in HNSCC and examined interlaboratory variation in PD-L1 positivity rates. METHODS: Pathology reports of HNSCC patients mentioning PD-L1 were extracted from the Dutch Pathology Registry (Palga). Tumour and PD-L1 testing characteristics were analysed per year and interlaboratory variation in PD-L1 positivity rates was assessed using funnel plots with 95% confidence limits around the overall mean. RESULTS: A total of 817 PD-L1 tests were reported in 702 patients among 19 laboratories; 85.2% of the tests on histological material were stated to be positive. The national PD-L1 positivity rate differed significantly per year during the study period (79.7-89.9%). The use of the recommended 22C3 antibody increased from 59.9 to 74.3%. A total of 673 PD-L1 tests on histological material from 12 laboratories were analysed to investigate interlaboratory variation. Four (33%) deviated significantly from the national mean of PD-L1-positive cases using CPS ≥ 1 cut-off, while two (17%) deviated significantly for CPS ≥ 20 cut-off. CONCLUSION: In the first 3 years of PD-L1 assessment in HNSCC, the testing landscape became more uniform. However, interlaboratory variation in PD-L1 positivity rates between Dutch laboratories was substantial. This implies that there is a need for further test standardisation to reduce this variation.

Significant inter- and intra-laboratory variation in grading of invasive breast cancer: A nationwide study of 33,043 patients in the Netherlands.

Accurate, consistent and reproducible grading by pathologists is of key-importance for identification of individual patients with invasive breast cancer (IBC) that will or will not benefit from adjuvant systemic treatment. We studied the laboratory-specific grading variation using nationwide real-life data to create insight and awareness in grading variation. Synoptic pathology reports of all IBC resection-specimens, obtained between 2013 and 2016, were retrieved from the nationwide Dutch Pathology Registry (PALGA). Absolute differences in laboratory-proportions of Grades I-III were compared to the national reference. Multivariable logistic regression provided laboratory-specific odds ratios (ORs) for high- vs. low-grade IBC. 33,792 IBC pathology reports of 33,043 patients from 39 laboratories were included, of which 28.1% were reported as Grade I (range between laboratories 16.3-43.3%), 47.6% as Grade II (38.4-57.8%), and 24.3% as Grade III (15.5-34.3%). Based on national guidelines, the indication for adjuvant chemotherapy was dependent on histologic grade in 29.9% of patients. After case-mix correction, 20 laboratories (51.3%) showed a significantly deviant OR. Significant grading differences were also observed among pathologists within laboratories. In this cohort of 33,043 breast cancer patients, we observed substantial inter- and intra-laboratory variation in histologic grading. It can be anticipated that this has influenced outcome including exposure to unnecessary toxicity, since choice of adjuvant chemotherapy was dependent on grade in nearly a third of patients. Better standardization and training seems warranted.

The effect of an e-learning module on grading variation of (pre)malignant breast lesions.

Histologic grade is a biomarker that is widely used to guide treatment of invasive breast cancer (IBC) and ductal carcinoma in situ of the breast (DCIS). Yet, currently, substantial grading variation between laboratories and pathologists exists in daily pathology practice. This study was conducted to evaluate whether an e-learning may be a feasible tool to decrease grading variation of (pre)malignant breast lesions. An e-learning module, representing the key-concepts of grading (pre)malignant breast lesions through gold standard digital images, was designed. Pathologists and residents could take part in either or both the separate modules on DCIS and IBC. Variation in grading of a digital set of lesions before and after the e-learning was compared in a fully-crossed study-design. Multiple outcome measures were assessed: inter-rater reliability (IRR) by Light's kappa, the number of images graded unanimously, the number of images with both extreme scores (i.e., grade I and grade III), and the average number of discrepancies from expert-consensus. Participants were included as they completed both the pre- and post-e-learning set (DCIS-module: n = 36, IBC-module: n = 21). For DCIS, all outcome measures improved after e-learning, with the IRR improving from fair (kappa: 0.532) to good (kappa: 0.657). For IBC, all outcome measures for the subcategories tubular differentiation and mitosis improved, with >90% of participants agreeing on almost 90% of the images after the e-learning. In contrast, the IRR for the subcategory of nuclear pleomorphism remained fair (kappa: 0.523 vs. kappa: 0.571). This study shows that an e-learning module, in which pathologists and residents are trained in histologic grading of DCIS and IBC, is a feasible and promising tool to decrease grading variation of (pre)malignant breast lesions. This is highly relevant given the important role of histologic grading in clinical decision making of (pre)malignant breast lesions.

Hormone- and HER2-receptor assessment in 33,046 breast cancer patients: a nationwide comparison of positivity rates between pathology laboratories in the Netherlands.

PURPOSE: Patient management of invasive breast cancer (IBC) is to a large extent based on hormone- and HER2-receptor assessment. High-quality, reliable receptor assessment is of key importance as false results may lead to under- or overtreatment of patients. Surveillance of case-mix adjusted positivity rates has been suggested as a tool to identify laboratories with insufficient testing assays, as this covers the whole process of receptor assessment and enables laboratories to benchmark their positivity rates against other laboratories. We studied laboratory-specific variation in hormone- and HER2 positivity rates of 33,046 breast cancer patients using real-life nationwide data. METHODS: All synoptic pathology reports of IBC resection-specimens, obtained between 2013 and 2016, were retrieved from the nationwide Dutch pathology registry (PALGA). Absolute and case-mix adjusted receptor positivity rates were compared to the mean national proportion and presented in funnel plots in separate analyses for estrogen (ER), progesterone (PR) and HER2. Case-mix adjustment was performed by multivariable logistic regression. RESULTS: 33,794 IBC lesions from 33,046 patients of 39 pathology laboratories were included. After case-mix adjustment, mean positivity rates were 87.2% for ER (range 80.4-94.3), 71.3% for PR (62.5-77.5%), and 9.9% for HER2 (5.5-12.7%). Overall, 14 (35.9%), 17 (43.6%) and 11 (28.2%) laboratories showed positivity rates outside the 95% confidence interval for ER, PR and HER2, respectively. CONCLUSION: This nationwide study shows that absolute variation in hormone- and HER2-receptor positivity rates between Dutch pathology laboratories is limited. Yet, the considerable number of outlying laboratories shows that there is still need for improvement. Continuous monitoring and benchmarking of positivity rates may help to realize this.

Significant inter- and intra-laboratory variation in grading of ductal carcinoma in situ of the breast: a nationwide study of 4901 patients in the Netherlands.

PURPOSE: A considerable part of ductal carcinoma in situ (DCIS) lesions may never progress into invasive breast cancer. However, standard treatment consists of surgical excision. Trials aim to identify a subgroup of low-risk DCIS patients that can safely forgo surgical treatment based on histologic grade, which highlights the importance of accurate grading. Using real-life nationwide data, we aimed to create insight and awareness in grading variation of DCIS in daily clinical practice. METHODS: All synoptic pathology reports of pure DCIS resection specimens between 2013 and 2016 were retrieved from PALGA, the nationwide Dutch Pathology Registry. Absolute differences in proportions of grade I-III were visualized using funnel plots. Multivariable analysis was performed by logistic regression to correct for case-mix, providing odds ratios and 95% confidence intervals for high-grade (III) versus low-grade (I-II) DCIS. RESULTS: 4952 DCIS reports from 36 laboratories were included, of which 12.5% were reported as grade I (range 6.1-24.4%), 39.5% as grade II (18.2-57.6%), and 48.0% as grade III (30.2-72.7%). After correction for case-mix, 14 laboratories (38.9%) reported a significantly lower (n = 4) or higher (n = 10) proportion of high-grade DCIS than the reference laboratory. Adjusted ORs (95%CI) ranged from 0.52 (0.31-0.87) to 3.83 (1.42-10.39). Significant grading differences were also observed among pathologists within laboratories. CONCLUSION: In this cohort of 4901 patients, we observed substantial inter- and intra-laboratory variation in DCIS grading, not explained by differences in case-mix. Therefore, there is an urgent need for nationwide standardization of grading practices, especially since the future management of DCIS may alter significantly depending on histologic grade.

Alcohol and Dietary Folate Intake and Promoter CpG Island Methylation in Clear-Cell Renal Cell Cancer.

We investigated whether alcohol and dietary folate intakes were associated with promoter methylation in clear-cell renal cell carcinoma (ccRCC). The Netherlands Cohort Study with a case-cohort design included 120,852 subjects aged 55-69 yr in 1986. Diet was measured with a food-frequency questionnaire. After 20.3 yr of follow-up, paraffin-embedded tumor blocks were collected. Methylation-specific polymerase chain reaction (MSP) was used to analyze promoter methylation of 11 genes. ccRCC cases were classified into low (0-19% of the genes), intermediate (20-39%), and high (40%+) methylation. Multivariable Cox regression analyses were conducted, stratified according to methylation, including 3980 subcohort members and 297 ccRCC cases. Increasing alcohol intake was associated with decreased ccRCC risk, but was not statistically significant; multivariable adjusted hazard ratio (HR) for ≥30 g alcohol/day versus 0 g/day was 0.78 [95% confidence interval (CI): 0.48-1.24], and P-value for trend was 0.46. In strata according to methylation index, no significant heterogeneity was observed. Dietary folate intake was not associated with ccRCC risk. There was no significant heterogeneity between strata according to methylation index. There was no effect modification of alcohol and dietary folate intake on ccRCC risk, nor in strata according to methylation index. Our findings do not support the hypothesis that alcohol and dietary folate intakes are involved in ccRCC.

Nationwide differences in cytology fixation and processing methods and their impact on interlaboratory variation in PD-L1 positivity.

Programmed death ligand-1 (PD-L1) immunostaining, which aids clinicians in decision-making on immunotherapy for non-small cell lung cancer (NSCLC) patients, is sometimes performed on cytological specimens. In this study, differences in cytology fixation and cell block (CB) processing between pathology laboratories were assessed, and the influence of these differences on interlaboratory variation in PD-L1 positivity was investigated. Questionnaires on cytology processing were sent to all Dutch laboratories. Information gathered from the responses was added to data on all Dutch NSCLC patients with a mention of PD-L1 testing in their cytopathology report from July 2017 to December 2018, retrieved from PALGA (the nationwide network and registry of histo- and cytopathology in the Netherlands). Case mix-adjusted PD-L1 positivity rates were determined for laboratories with known fixation and CB method. The influence of differences in cytology processing on interlaboratory variation in PD-L1 positivity was assessed by comparing positivity rates adjusted for differences in the variables fixative and CB method with positivity rates not adjusted for differences in these variables. Twenty-eight laboratories responded to the survey and reported 19 different combinations of fixation and CB method. Interlaboratory variation in PD-L1 positivity was assessed in 19 laboratories. Correcting for differences in the fixative and CB method resulted in a reduction (from eight (42.1%) to five (26.3%)) in the number of laboratories that differed significantly from the mean in PD-L1 positivity. Substantial variation in cytology fixation and CB processing methods was observed between Dutch pathology laboratories, which partially explains the existing considerable interlaboratory variation in PD-L1 positivity.

Considerable interlaboratory variation in PD-L1 positivity in a nationwide cohort of non-small cell lung cancer patients.

OBJECTIVES: Immunohistochemical expression of programmed death-ligand 1 (PD-L1) is used as a predictive biomarker for prescription of immunotherapy to non-small cell lung cancer (NSCLC) patients. Accurate assessment of PD-L1 expression is therefore crucial. In this study, the extent of interlaboratory variation in PD-L1 positivity in the Netherlands was assessed, using real-world clinical pathology data. MATERIALS AND METHODS: Data on all NSCLC patients in the Netherlands with a mention of PD-L1 testing in their pathology report from July 2017 to December 2018 were extracted from PALGA, the nationwide network and registry of histo- and cytopathology in the Netherlands. PD-L1 positivity rates were determined for each laboratory that performed PD-L1 testing, with separate analyses for histological and cytological material. Two cutoffs (1% and 50%) were used to determine PD-L1 positivity. Differences between laboratories were assessed using funnel plots with 95% confidence limits around the overall mean. RESULTS: 6,354 patients from 30 laboratories were included in the analysis of histology data. At the 1% cutoff, maximum interlaboratory variation was 39.1% (32.7%-71.8%) and ten laboratories (33.3%) differed significantly from the mean. Using the 50% cutoff, four laboratories (13.3%) differed significantly from the mean and maximum variation was 23.1% (17.2%-40.3%). In the analysis of cytology data, 1,868 patients from 23 laboratories were included. Eight laboratories (34.8%) differed significantly from the mean in the analyses of both cutoffs. Maximum variation was 41.2% (32.2%-73.4%) and 29.2% (14.7%-43.9%) using the 1% and 50% cutoffs, respectively. CONCLUSION: Considerable interlaboratory variation in PD-L1 positivity was observed. Variation was largest using the 1% cutoff. At the 50% cutoff, analysis of cytology data demonstrated a higher degree of variation than the analysis of histology data.

Significant Inter- and Intralaboratory Variation in Gleason Grading of Prostate Cancer: A Nationwide Study of 35,258 Patients in The Netherlands.

PURPOSE: Our aim was to analyze grading variation between pathology laboratories and between pathologists within individual laboratories using nationwide real-life data. METHODS: We retrieved synoptic (n = 13,397) and narrative (n = 29,377) needle biopsy reports from the Dutch Pathology Registry and prostate-specific antigen values from The Netherlands Cancer Registration for prostate cancer patients diagnosed between January 2017 and December 2019. We determined laboratory-specific proportions per histologic grade and unadjusted odds ratios (ORs) for International Society of Urological Pathologists Grades 1 vs. 2-5 for 40 laboratories due to treatment implications for higher grades. Pathologist-specific proportions were determined for 21 laboratories that consented to this part of analysis. The synoptic reports of 21 laboratories were used for analysis of case-mix correction for PSA, age, year of diagnosis, number of biopsies and positive cores. RESULTS: A total of 38,321 reports of 35,258 patients were included. Grade 1 ranged between 19.7% and 44.3% per laboratory (national mean = 34.1%). Out of 40 laboratories, 22 (55%) reported a significantly deviant OR, ranging from 0.48 (95% confidence interval (CI) 0.39-0.59) to 1.54 (CI 1.22-1.93). Case-mix correction was performed for 10,294 reports, altering the status of 3/21 (14%) laboratories, but increasing the observed variation (20.8% vs. 17.7%). Within 15/21 (71%) of laboratories, significant inter-pathologist variation existed. CONCLUSION: Substantial variation in prostate cancer grading was observed between and within Dutch pathology laboratories. Case-mix correction did not explain the variation. Better standardization of prostate cancer grading is warranted to optimize and harmonize treatment.

Grading variation in 2,934 patients with ductal carcinoma in situ of the breast: the effect of laboratory- and pathologist-specific feedback reports.

BACKGROUND: Histologic grade of ductal carcinoma in situ of the breast (DCIS) may become the single biomarker that decides whether patients will be treated. Yet, evidence shows that grading variation in daily practice is substantial. To facilitate quality improvement, feedback reports, in which laboratory-specific case-mix adjusted proportions per grade were benchmarked against other laboratories, were sent to the individual laboratories by March 1, 2018. One year later, the effect of these feedback reports on inter-laboratory variation was studied. METHODS: Synoptic pathology reports of all pure DCIS resection specimens between March 1, 2017 and March 1, 2019 were retrieved from PALGA (the nationwide Dutch pathology registry). Laboratory-specific proportions per grade were compared to the overall proportion in the year before and after feedback. The absolute deviation for all three grades at once, represented by the overall deviation score (ODS), was calculated as the sum of deviations from the grade-specific overall proportions. Case-mix adjusted, laboratory-specific odds ratios (ORs) for high- (grade III) versus low-grade (grade I-II) DCIS were obtained by multivariable logistic regression. RESULTS: Overall, 2954 DCIS reports from 31 laboratories were included. After feedback, the range between laboratories decreased by 22 and 6.5% for grades II and III, while an increase of 6.2% was observed for grade I. Both the mean ODS (27.2 to 24.1%) and maximum ODS (87.7 to 59.6%) decreased considerably. However, the range of case-mix adjusted ORs remained fairly stable and substantial (0.39 (95% CI: 0.18-0.86) to 3.69 (95% CI: 1.30-10.51)). CONCLUSION: A promising decrease in grading variation was observed after laboratory-specific feedback for DCIS grades II-III, while this was not observed for DCIS grade I. Overall, grading variation remained substantial which needs to be addressed considering its clinical implications. Nationwide consensus on a classification, and training of (expert breast) pathologists, for example by e-learning, may help to further improve grading standardization.

Potential role of gene-environment interactions in ion transport mechanisms in the etiology of renal cell cancer.

We investigated the ion transport mechanism (ITM) in renal cell cancer (RCC) etiology using gene-environment interactions between candidate single nucleotide polymorphisms (SNPs) and associated environmental factors, including dietary intakes of sodium, potassium and fluid, hypertension and diuretic medication. A literature-based selection of 13 SNPs in ten ITM genes were successfully genotyped in toenail DNA of 3,048 subcohort members and 419 RCC cases from the Netherlands Cohort Study. Diet and lifestyle were measured with baseline questionnaires. Cox regression analyses were conducted for main effects and gene-environment interactions. ADD1_rs4961 was significantly associated with RCC risk, showing a Hazard Ratio (HR) of 1.24 (95% confidence intervals (CI): 1.01-1.53) for the GT + TT (versus GG) genotype. Four of 65 tested gene-environment interactions were statistically significant. Three of these interactions clustered in SLC9A3_rs4957061, including the ones with fluid and potassium intake, and diuretic medication. For fluid intake, the RCC risk was significantly lower for high versus low intake in participants with the CC genotype (HR(95% CI): 0.47(0.26-0.86)), but not for the CT + TT genotype (P-interaction = 0.002). None of the main genetic effects and gene-environment interactions remained significant after adjustment for multiple testing. Data do not support the general hypothesis that the ITM is a disease mechanism in RCC etiology.

Promoter Methylation of CDO1 Identifies Clear-Cell Renal Cell Cancer Patients with Poor Survival Outcome.

PURPOSE: In this era of molecular diagnostics, prediction of clear-cell renal cell cancer (ccRCC) survival requires optimization, as current prognostic markers fail to determine individual patient outcome. Epigenetic events are promising molecular markers. Promoter CpG island methylation of cysteine dioxygenase type 1 (CDO1), which was identified as prognostic marker for breast cancer, is studied as a potential marker for ccRCC survival. EXPERIMENTAL DESIGN: We collected primary tissues of 365 ccRCC cases identified within the prospective Netherlands Cohort Study (NLCS). In this population-based series, CDO1 promoter methylation was observed in 124 of 324 (38.3%) patients with successful methylation-specific PCR analysis. Kaplan-Meier curves and Wilcoxon tests were used to evaluate 10-year ccRCC-specific survival. Cox regression analysis was used to obtain crude and multivariate HRs and 95% confidence intervals (CI). The relative prognostic value of multivariate models with and without CDO1 promoter methylation was compared using likelihood-ratio tests. RESULTS: Patients with CDO1 promoter methylation have a significantly poorer survival than those without (Wilcoxon P = 0.006). Differences in survival were independent of other prognostic factors, including age and sex (HR, 1.66; 95% CI, 1.12-2.45) and TNM stage, tumor size, and Fuhrman grade (HR, 1.89; 95% CI, 1.25-2.85). Multivariate models performed better with than without CDO1 promoter methylation status (likelihood-ratio P = 0.003). Survival curves were validated in an independent series of 280 ccRCC cases from The Cancer Genome Atlas (TCGA; Wilcoxon P < 0.001). CONCLUSIONS: CDO1 promoter methylation may not substitute common prognostic makers to predict ccRCC survival, but offers additional, relevant prognostic information, indicating that it might be a novel molecular marker to determine ccRCC prognosis.

Investigating international time trends in the incidence and prevalence of atopic eczema 1990-2010: a systematic review of epidemiological studies.

The prevalence of atopic eczema has been found to have increased greatly in some parts of the world. Building on a systematic review of global disease trends in asthma, our objective was to study trends in incidence and prevalence of atopic eczema. Disease trends are important for health service planning and for generating hypotheses regarding the aetiology of chronic disorders. We conducted a systematic search for high quality reports of cohort, repeated cross-sectional and routine healthcare database-based studies in seven electronic databases. Studies were required to report on at least two measures of the incidence and/or prevalence of atopic eczema between 1990 and 2010 and needed to use comparable methods at all assessment points. We retrieved 2,464 citations, from which we included 69 reports. Assessing global trends was complicated by the use of a range of outcome measures across studies and possible changes in diagnostic criteria over time. Notwithstanding these difficulties, there was evidence suggesting that the prevalence of atopic eczema was increasing in Africa, eastern Asia, western Europe and parts of northern Europe (i.e. the UK). No clear trends were identified in other regions. There was inadequate study coverage worldwide, particularly for repeated measures of atopic eczema incidence. Further epidemiological work is needed to investigate trends in what is now one of the most common long-term disorders globally. A range of relevant measures of incidence and prevalence, careful use of definitions and description of diagnostic criteria, improved study design, more comprehensive reporting and appropriate interpretation of these data are all essential to ensure that this important field of epidemiological enquiry progresses in a scientifically robust manner.