Causes behind error rates for predictive biomarker testing:the utility of sending post-EQA surveys

External quality assessment (EQA) schemes assess the performance of predictive biomarker testing in lung and colorectal cancer and have previously demonstrated variable error rates. No information is currently available on the underlying causes of incorrect EQA results in the laboratories. Participants in EQA schemes by the European Society of Pathology between 2014 and 2018 for lung and colorectal cancer were contacted to complete a survey if they had at least one analysis error or test failure in the provided cases. Of the 791 surveys that were sent, 325 were completed including data from 185 unique laboratories on 514 incorrectly analyzed or failed cases. For the digital cases and immunohistochemistry, the majority of errors were interpretation-related. For fluorescence in situ hybridization, problems with the EQA materials were reported frequently. For variant analysis, the causes were mainly methodological for lung cancer but variable for colorectal cancer. Post-analytical (clerical and interpretation) errors were more likely detected after release of the EQA results compared to pre-analytical and analytical issues. Accredited laboratories encountered fewer reagent problems and more often responded to the survey. A recent change in test methodology resulted in method-related problems. Testing more samples annually introduced personnel errors and lead to a lower performance in future schemes. Participation to quality improvement projects is important to reduce deviating test results in laboratories, as the different error causes differently affect the test performance. EQA providers could benefit from requesting root cause analyses behind errors to offer even more tailored feedback, subschemes, and cases

Transgenic mice with mammary gland targeted expression of human cortactin do not develop (pre-malignant) breast tumors: studies in MMTV-cortactin and MMTV-cortactin/-cyclin D1 bitransgenic mice

BACKGROUND: In human breast cancers, amplification of chromosome 11q13 correlates with lymph node metastasis and increased mortality. To date, two genes located within this amplicon, CCND1 and EMS1, were considered to act as oncogenes, because overexpression of both proteins, respectively cyclin D1 and cortactin, correlated well with 11q13 amplification. Cyclin D1 is involved in cell cycle regulation and the F-actin-binding protein cortactin in cytoskeletal dynamics and cell migration. To study the role of cortactin in mammary gland tumorigenesis, we examined mouse mammary tumor virus (MMTV)-cortactin transgenic mice and MMTV-cortactin/-MMTV-cyclin D1 bitransgenic mice. METHODS: MMTV-cortactin transgenic mice were generated and intercrossed with previously described MMTV-cyclin D1 transgenic mice. Immunohistochemical, Northern and Western blot analyses were performed to study the expression of human transgene cortactin during mammary gland development and in mammary tumors. For tumor incidence studies, forced-bred, multiparous mice were used to enhance transgene expression in the mammary gland. Microscopical examination was performed using haematoxylin and eosin staining. RESULTS: Mammary gland tumors arose stochastically (incidence 21%) with a mean age of onset at 100 weeks. This incidence, however, did not exceed that of aged-matched control FVB/N mice (38%), which unexpectedly, also developed spontaneous mammary gland tumors. We mimicked 11q13 amplification by generating MMTV-cortactin/-MMTV-cyclin D1 bitransgenic mice but did not observe any synergistic effect of cortactin on cyclin D1-induced mammary hyperplasias or carcinomas, nor development of distant metastasis. CONCLUSION: From this study, we conclude that development of (pre-malignant) breast tumors in either wild type or MMTV-cyclin D1 mice was not augmented due to mammary gland targeted overexpression of human cortactin

"The leading role of pathology in assessing the somatic molecular alterations of cancer:Position Paper of the European Society of Pathology": letter to the Editor

Contains fulltext : 232005.pdf (Publisher’s version ) (Open Access

Variation in nomenclature of somatic variants for selection of oncological therapies:Can we reach a consensus soon?

A standardized nomenclature for reporting oncology biomarker variants is key to avoid misinterpretation of results and unambiguous registration in clinical databases. External quality assessment (EQA) schemes have revealed a need for more consistent nomenclature use in clinical genetics. We evaluated the propensity of EQA for improvement of compliance with Human Genome Variation Society (HGVS) recommendations for reporting of predictive somatic variants in lung and colorectal cancer. Variant entries between 2012 and 2018 were collected from written reports and electronic results sheets. In total, 4,053 variants were assessed, of which 12.1% complied with HGVS recommendations. Compliance improved over time from 2.1% (2012) to 22.3% (2018), especially when laboratories participated in multiple EQA schemes. Compliance was better for next-generation sequencing (20.9%) compared with targeted techniques (9.8%). In the 1792 reports, HGVS recommendations for reference sequences were met for 31.9% of reports, for 36.0% of noncommercial, and 26.5% of commercial test methods. Compliance improved from 16.7% (2012) to 33.1% (2018), and after repeated EQA participation. EQA participation improves compliance with HGVS recommendations. The residual percentage of errors in the most recent schemes suggests that laboratories, companies, and EQA providers need to collaborate for additional improvement of harmonization in clinical test reporting

Immune microenvironment composition in non-small cell lung cancer and its association with survival

Objectives In non-small cell lung cancer (NSCLC), the immune system and possibly its composition affect survival. In thisin silicostudy, the immune infiltrate composition in NSCLC patients was evaluated. Methods Gene expression data of tumors from early NSCLC patients were obtained from Gene Expression Omnibus (GEO). With CIBERSORT, 22 immune cell fractions were estimated. Results The immune infiltrate of 1430 pretreatment NSCLC patients contained mostly plasma cells, macrophages and CD8 T cells. Higher fractions of resting mast and CD4 T-helper cells were associated with longer overall survival (OS) (HR = 0.95,P <0.01; HR = 0.98, = 0.04, respectively) and higher fractions of M2 macrophages and active dendritic cells with shorter survival (HR = 1.02,P = 0.03; HR = 1.03,P = 0.05, respectively). Adenocarcinoma patients with survival data (n = 587) showed higher fractions of resting mast and resting CD4 T cells, and lower M0 macrophages than squamous cell carcinoma (n = 254), which were associated with OS (HR = 0.95,P = 0.04; HR = 0.97,P = 0.01; HR = 1.03,P = 0.01, respectively). Fractions of memory B cells, naive CD4 T cells and neutrophils had different associations with survival depending on the subtype. Smokers had had higher fractions of regulatory T cell, follicular helper T cell, neutrophil and M2 macrophage, which were associated with shorter survival (HR = 1.3,P <0.01; HR = 1.13,P = 0.02; HR = 1.09,P = 0.03; HR = 1.04,P = 0.02, respectively). Conclusion Pretreatment differences in immune cell composition in NSCLC are associated with survival and depend on smoking status and histological subtype. Smokers' immune composition is associated with lower survival

External Quality Assessment Schemes for Biomarker Testing in Oncology:Comparison of Performance between Formalin-Fixed, Paraffin-Embedded-Tissue and Cell-Free Tumor DNA in Plasma

Liquid biopsies have emerged as a useful addition to tissue biopsies in molecular pathology. Literature has shown lower laboratory performances when a new method of variant analysis is introduced. This study evaluated the differences in variant analysis between tissue and plasma samples after the introduction of liquid biopsy in molecular analysis. Data from a pilot external quality assessment scheme for the detection of molecular variants in plasma samples and from external quality assessment schemes for the detection of molecular variants in tissue samples were collected. Laboratory performance and error rates by sample were compared between matrices for variants present in both scheme types. Results showed lower overall performance [65.6% (n = 276) versus 89.2% (n = 1607)] and higher error rates [21.0% to 43.5% (n = 138) versus 8.7% to 16.7% (n = 234 to 689)] for the detection of variants in plasma compared to tissue, respectively. In the plasma samples, performance was decreased for variants with an allele frequency of 1% compared to 5% [56.5% (n = 138) versus 74.6% (n = 138)]. The implementation of liquid biopsy in the detection of circulating tumor DNA in plasma was associated with poor laboratory performance. It is important both to apply optimal detection methods and to extensively validate new methods for testing circulating tumor DNA before treatment decisions are made

Detection of NTRK Fusions and TRK Expression and Performance of pan-TRK Immunohistochemistry in Routine Diagnostics:Results from a Nationwide Community-Based Cohort

Gene fusions involving NTRK1, NTRK2, and NTRK3 are rare drivers of cancer that can be targeted with histology-agnostic inhibitors. This study aimed to determine the nationwide landscape of NTRK/TRK testing in the Netherlands and the usage of pan-TRK immunohistochemistry (IHC) as a preselection tool to detect NTRK fusions. All pathology reports in 2017–2020 containing the search term ‘TRK’ were retrieved from the Dutch Pathology Registry (PALGA). Patient characteristics, tumor histology, NTRK/TRK testing methods, and reported results were extracted. NTRK/TRK testing was reported for 7457 tumors. Absolute testing rates increased from 815 (2017) to 3380 (2020). Tumors were tested with DNA/RNA-based molecular assay(s) (48%), IHC (47%), or in combination (5%). A total of 69 fusions involving NTRK1 (n = 22), NTRK2 (n = 6) and NTRK3 (n = 41) were identified in tumors from adult (n = 51) and pediatric (n = 18) patients. In patients tested with both IHC and a molecular assay (n = 327, of which 29 NTRK fusion-positive), pan-TRK IHC had a sensitivity of 77% (95% confidence interval (CI), 56–91) and a specificity of 84% (95% CI, 78–88%). These results showed that pan-TRK IHC has a low sensitivity in current routine practice and warrants the introduction of quality guidelines regarding the implementation and interpretation of pan-TRK IHC

Preferences and Experiences Regarding the Use of the Self-Sampling Device in hrHPV Screening for Cervical Cancer

BACKGROUND: To improve participation in the Dutch cervical cancer screening, a self-sampling device (SSD) was introduced in 2017 into the Dutch population-based screening programme (PBS) for the early detection of cervical cancer. The aim of this study was to gather potential preferences and experiences that might influence a woman’s decision to use the SSD in the Dutch PBS. METHODS: A scoping review was performed in the PubMed database. Studies that assessed preferences and experiences of women regarding the SSD were included, and preferences and experiences were extracted. In addition, in a qualitative study, the list of potential preferences and experiences specific for the Dutch PBS was extended based on semi-structured interviews with SSD users as well as non-SSD users who recently participated in the PBS, analysed in a structured manner by translating full sentences to key words. RESULTS: Ninety-eight studies were included in the scoping review and 16 interviews were performed. Frequently mentioned reasons for using the SSD, in both the interviews and the literature, were practicality and comfort. Frequently mentioned reasons for not using the SSD were fear of not performing the SSD procedure correctly and doubts on whether the results of the high-risk human papillomavirus (hrHPV) test will be reliable. A new positive experience elicited in the interviews was accessibility. Negative preferences and experiences were not being aware the SSD was an option, and the inconvenience that after an hrHPV-positive test result of the SSD, an additional smear test at the GP is necessary. CONCLUSION: Several preferences and experiences play a role in the choice whether or not to use the SSD. Based on the currently found preferences and experiences, an app will be developed in order to assess which of these are the most important for women participating in the Dutch population-based cervical screening programme. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40271-021-00550-y

Cortactin overexpression results in sustained epidermal growth factor receptor signaling by preventing ligand-induced receptor degradation in human carcinoma cells

The chromosome 11q13 region is frequently amplified in human carcinomas and results in an increased expression of various genes including cortactin, and is also associated with an increased invasive potential. Cortactin acts as an important regulator of the actin cytoskeleton. It is therefore very tempting to speculate that cortactin is the crucial gene within the 11q13 amplicon that mediates the invasive potential of these carcinomas. Cortactin also participates in receptor-mediated endocytosis, and recent findings have shown that, during receptor internalization, cortactin overexpression inhibits the ubiquitylation-mediated degradation of the epidermal growth factor receptor, resulting in a sustained ligand-induced epidermal growth factor receptor activity

Microsieves for the detection of circulating tumor cells in leukapheresis product in non-small cell lung cancer patients

Background: Circulating tumor cells (CTC) in non-small cell lung cancer (NSCLC) patients are a prognostic and possible therapeutic marker, but have a low frequency of appearance. Diagnostic leukapheresis (DLA) concentrates CTC and mononuclear cells from the blood. We evaluated a protocol using two VyCAP microsieves to filter DLA product of NSCLC patients and enumerate CTC, compared with CellSearch as a gold standard. Methods: DLA was performed in NSCLC patients before starting treatment. DLA product equaling 2×108 leukocytes was diluted to 9 mL with CellSearch dilution buffer in a Transfix CTC tube. Within 72 hours the sample was filtered with a 7 μm pore microsieve and subsequently over a 5μm pore microsieve. CTC were defined as nucleated cells which stained for cytokeratin, but lacked CD45 and CD16. CellSearch detected CTC in the same volume of DLA. Results: Of 29 patients a median of 1.4 mL DLA product (range, 0.5-4.1) was filtered (2% of total product) successfully in 93% and 45% of patients using 7 and 5 μm pores, respectively. Two DLA products were unevaluable for CTC detection. Clogging of the 5 μm but not 7 μm microsieves was positively correlated with fixation time (ρ=0.51, P<0.01). VyCAP detected CTC in 44% (12/27) of DLA products. Median CTC count per mL DLA was 0 [interquartile range (IQR): 0-1]. CellSearch detected CTC in 63% of DLA products (median =0.9 CTC per mL DLA, IQR: 0-2.1). CTC counts detected by CellSearch were significantly higher compared with VyCAP (P=0.05). Conclusions: VyCAP microsieves can identify CTC in DLA product, but workflows need to be optimized