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Ki67 has potential clinical importance in breast cancer but has yet to see broad acceptance due to inter-laboratory variability. Here we tested an open source and calibrated automated digital image analysis (DIA) platform to: (i) investigate the comparability of Ki67 measurement across corresponding core biopsy and resection specimen cases, and (ii) assess section to section differences in Ki67 scoring. Two sets of 60 previously stained slides containing 30 core-cut biopsy and 30 corresponding resection specimens from 30 estrogen receptor-positive breast cancer patients were sent to 17 participating labs for automated assessment of average Ki67 expression. The blocks were centrally cut and immunohistochemically (IHC) stained for Ki67 (MIB-1 antibody). The QuPath platform was used to evaluate tumoral Ki67 expression. Calibration of the DIA method was performed as in published studies. A guideline for building an automated Ki67 scoring algorithm was sent to participating labs. Very high correlation and no systematic error (p = 0.08) was found between consecutive Ki67 IHC sections. Ki67 scores were higher for core biopsy slides compared to paired whole sections from resections (p ≤ 0.001; median difference: 5.31%). The systematic discrepancy between core biopsy and corresponding whole sections was likely due to pre-analytical factors (tissue handling, fixation). Therefore, Ki67 IHC should be tested on core biopsy samples to best reflect the biological status of the tumor.

BackgroundObesity and cardiometabolic dysfunction have been associated with cancer risk and severity. Underlying mechanisms remain unclear.ObjectivesThe aim of this study was to examine associations of obesity and related cardiometabolic traits with incident cancer.MethodsFHS (Framingham Heart Study) and PREVEND (Prevention of Renal and Vascular End-Stage Disease) study participants without prevalent cancer were studied, examining associations of obesity, body mass index (BMI), waist circumference, visceral adipose tissue (VAT) and subcutaneous adipose tissue depots, and C-reactive protein (CRP) with future cancer in Cox models.ResultsAmong 20,667 participants (mean age 50 years, 53% women), 2,619 cancer events were observed over a median follow-up duration of 15 years. Obesity was associated with increased risk for future gastrointestinal (HR: 1.30; 95% CI: 1.05-1.60), gynecologic (HR: 1.62; 95% CI: 1.08-2.45), and breast (HR: 1.32; 95% CI: 1.05-1.66) cancer and lower risk for lung cancer (HR: 0.62; 95% CI: 0.44-0.87). Similarly, waist circumference was associated with increased risk for overall, gastrointestinal, and gynecologic but not lung cancer. VAT but not subcutaneous adipose tissue was associated with risk for overall cancer (HR: 1.22; 95% CI: 1.05-1.43), lung cancer (HR: 1.92; 95% CI: 1.01-3.66), and melanoma (HR: 1.56; 95% CI: 1.02-2.38) independent of BMI. Last, higher CRP levels were associated with higher risk for overall, colorectal, and lung cancer (P < 0.05 for all).ConclusionsObesity and abdominal adiposity are associated with future risk for specific cancers (eg, gastrointestinal, gynecologic). Although obesity was associated with lower risk for lung cancer, greater VAT and CRP were associated with higher lung cancer risk after adjusting for BMI.

Aim: Manual detection and scoring of Ki67 hotspots is difficult and prone to variability, limiting its clinical utility. Automated hotspot detection and scoring by digital image analysis (DIA) could improve the assessment of the Ki67 hotspot proliferation index (PI). This study compared the clinical performance of Ki67 hotspot detection and scoring DIA algorithms based on virtual dual staining (VDS) and deep learning (DL) with manual Ki67 hotspot PI assessment. Methods: Tissue sections of 135 consecutive invasive breast carcinomas were immunohistochemically stained for Ki67. Two DIA algorithms, based on VDS and DL, automatically determined the Ki67 hotspot PI. For manual assessment; two independent observers detected hotspots and calculated scores using a validated scoring protocol. Results: Automated hotspot detection and assessment by VDS and DL could be performed in 73% and 100% of the cases, respectively. Automated hotspot detection by VDS and DL led to higher Ki67 hotspot PIs (mean 39.6% and 38.3%, respectively) compared to manual consensus Ki67 PIs (mean 28.8%). Comparing manual consensus Ki67 PIs with VDS Ki67 PIs revealed substantial correlation (r = 0.90), while manual consensus versus DL Ki67 PIs demonstrated high correlation (r = 0.95). Conclusion: Automated Ki67 hotspot detection and analysis correlated strongly with manual Ki67 assessment and provided higher PIs compared to manual assessment. The DL‐based algorithm outperformed the VDS‐based algorithm in clinical applicability, because it did not depend on virtual alignment of slides and correlated stronger with manual scores. Use of a DL‐based algorithm may allow clearer Ki67 PI cutoff values, thereby improving the clinical usability of Ki67. [ABSTRACT FROM AUTHOR]

Aims: Recently, human epidermal growth factor 2 (HER2)‐low (i.e. HER2 score 1+ or 2+ without amplification) breast cancer patients became eligible for trastuzumab–deruxtecan treatment. To improve assay standardisation and detection of HER2‐low in a quantitative manner, we conducted an external quality assessment‐like study in the Netherlands. Dynamic range cell lines and immunohistochemistry (IHC) calibrators were used to quantify HER2 expression and to assess interlaboratory variability. Methods and results: Three blank slides with a dynamic range cell line and an IHC calibrator were stained with routine HER2 assays by 35 laboratories. Four different antibody clones were used: 19 (54.3%) 4B5, six (17.1%) A0485, five (14.3%) DG44 (HercepTest) and five (14.3%) SP3. Laboratories used two different detection kits for 4B5 assays: 14 (73.7%) ultraView and five (26.3%) OptiView. Variability of HER2 expression in cell lines, measured with artificial intelligence software, was median (min–max) = negative core 0.5% (0.0–57.0), 1+ core 4.3% (1.6–71.3), 2+ core 42.8% (30.4–92.6) and 3+ core 96.2% (91.8–98.8). The calibrators DG44 and 4B5 OptiView had the highest analytical sensitivity, closely followed by 4B5 ultraView. SP3 was the least sensitive. Calibrators of A0485 assays were not analysable due to background staining. Conclusions: As assays were validated for detecting HER2‐amplified tumours, not all assays and antibodies proved suitable for HER2‐low detection. Some tests showed distinct expression in the negative cell line. Dynamic range cell line controls and quantitative analysis using calibrators demonstrated more interlaboratory variability than commonly appreciated. Revalidation of HER2 tests by laboratories is needed to ensure clinical applicable HER2‐low assays. [ABSTRACT FROM AUTHOR]

Objectives To compare focus score and other histopathological features between paired labial and parotid salivary gland biopsies in a diagnostic cohort of suspected Sjögren's disease (SjD) patients. Methods Labial and parotid salivary gland biopsies were simultaneously obtained from patients with sicca complaints, suspected of having SjD. Biopsies were formalin fixed and paraffin embedded. Sections were stained with haematoxylin & eosin, and for CD3, CD20, CD45, cytokeratin, CD21, Bcl6, activation-induced deaminase (AID) and IgA/IgG. Focus score and other histopathological features characteristic for SjD were analysed. Results Based on the expert opinion of three experienced rheumatologists, 36 patients were diagnosed as SjD and 63 as non-SjD sicca patients. When taking all patients together, absolute agreement of various histopathological features between labial and parotid biopsies was high and varied between 80% (focus score) and 93% [(pre-)lymphoepithelial lesions (LELs)]. More labial gland biopsies had a focus score ≥1 compared with their parotid counterpart. Accordingly, the area of infiltrate was larger in labial gland biopsies. When considering only SjD patients, labial glands contained significantly fewer B-lymphocytes and germinal centres/mm2, and less severe LELs compared with parotid glands. Conclusion Labial and parotid glands from SjD patients contain similar histopathological key features, and thus both glands can be used for diagnosis and classification of SjD. However, parotid salivary glands reveal more evident B-lymphocyte-related features, while labial glands exhibit more inflammation, which may be partially unrelated to SjD. [ABSTRACT FROM AUTHOR]

Replication stress (RS) is a key trait of cancer cells, and a potential actionable target in cancer treatment. Accurate methods to measure RS in tumour samples are currently lacking. DNA fibre analysis has been used as a common technique to measure RS in cell lines. Here, we investigated DNA fibre analysis on fresh breast cancer specimens and correlated DNA replication kinetics to known RS markers and genomic alterations. Fresh, treatment‐naïve primary breast cancer samples (n = 74) were subjected to ex vivo DNA fibre analysis to measure DNA replication kinetics. Tumour cell proliferation was confirmed by EdU incorporation and cytokeratin AE1/AE3 (CK) staining. The RS markers phospho‐S33‐RPA and γH2AX and the RS‐inducing proto‐oncogenes Cyclin E1 and c‐Myc were analysed by immunohistochemistry. Copy number variations (CNVs) were assessed from genome‐wide single nucleotide polymorphism (SNP) arrays. We found that the majority of proliferating (EdU‐positive) cells in each sample were CK‐positive and therefore considered to be tumour cells. DNA fibre lengths varied largely in most tumour samples. The median DNA fibre length showed a significant inverse correlation with pRPA expression (r = −0.29, p = 0.033) but was not correlated with Cyclin E1 or c‐Myc expression and global CNVs in this study. Nuclear Cyclin E1 expression showed a positive correlation with pRPA levels (r = 0.481, p < 0.0001), while cytoplasmic Cyclin E1 expression exhibited an inverse association with pRPA expression (r = −0.353, p = 0.002) and a positive association with global CNVs (r = 0.318, p = 0.016). In conclusion, DNA fibre analysis performed with fresh primary breast cancer samples is feasible. Fibre lengths were associated with pRPA expression. Cyclin E1 expression was associated with pRPA and the percentage of CNVs. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland. [ABSTRACT FROM AUTHOR]

Objective: To assess whether narrow band imaging (NBI) detects fields of cancerisation around suspicious lesions in the upper aerodigestive tract, which were undetected by white light imaging (WLI). Methods: In 96 patients with laryngeal and pharyngeal lesions suspicious for malignancy, 206 biopsies were taken during laryngoscopy: 96 biopsies of suspicious lesions detected by both WLI and NBI (WLI+/NBI+), 60 biopsies adjacent mucosa only suspicious with NBI (WLI‐/NBI+), and 46 biopsies of NBI and WLI unsuspicious mucosa (WLI‐/NBI‐) as negative controls. Optical diagnosis according to the Ni‐classification was compared with histopathology. Results: Signs of (pre)malignancy were found in 88% of WLI+/NBI+ biopsies, 32% of WLI‐/NBI+ biopsies and 0% in WLI‐/NBI‐ (p <.001). In 58% of the WLI‐/NBI+ mucosa any form of dysplasia or carcinoma was detected. Conclusion: The use of additional NBI led to the detection of (pre)malignancy in 32% of the cases, that would have otherwise remained undetected with WLI alone. This highlights the potential of NBI as a valuable adjunct to WLI in the identification of suspicious lesions in the upper aerodigestive tract. [ABSTRACT FROM AUTHOR]