Diagnostic interobserver agreement for thyroid fine-needle aspirates: Effects of reviewer experience and molecular diagnostics.

OBJECTIVES: Few cytologically indeterminate thyroid fine-needle aspirations (FNAs) harbor BRAF V600E. Here, we assess interobserver agreement for The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) category III (atypia of undetermined significance [AUS]) FNAs harboring BRAF V600E and contrast their features with those harboring non-BRAF V600E alterations, with attention to cytopathology experience. METHODS: Seven reviewers evaluated 5 AUS thyroid FNAs harboring BRAF V600E. To blind reviewers, cases were intermixed with 19 FNAs falling within other TBSRTC categories and in which genetic alterations other than BRAF V600E had been identified (24 FNAs total). Interobserver agreement against both "index" and most popular ("mode") diagnoses was calculated. Four additional BRAF V600E cases were independently reviewed. RESULTS: Reviewers included 3 trainees and 3 American Board of Pathology (board)-certified cytopathologists. Board-certified cytopathologists, whose experience ranged from 2 to more than 15 subspecialty practice years, had known AUS rates. BRAF V600E was identified in 5 of 260 (2%) AUS FNAs. Interobserver agreement was higher among cytopathologists with more experience. Mode diagnosis differed from index diagnosis in 6 of 11 cases harboring RAS-like alterations; mode diagnosis was AUS in 4 of 5 BRAF V600E FNAs. CONCLUSIONS: Atypia of undetermined significance of thyroid FNAs harboring BRAF V600E is uncommon yet relatively reproducible, particularly among pathologists with experience. It is advisable to sequence BRAF across V600 in such cases.

Combining molecular testing and the Bethesda category III:VI ratio for thyroid fine-needle aspirates: A quality-assurance metric for evaluating diagnostic performance in a cytopathology laboratory.

BACKGROUND: Molecular testing (MT) of thyroid fine-needle aspiration (FNA)-derived genetic material is commonly used to assess malignancy risk for indeterminate cases. The Bethesda System for Reporting Thyroid Cytopathology (TBS) provides limited guidance for the appropriate use of category III (atypia of undetermined significance [AUS]). The authors combined MT with cytomorphology to monitor AUS diagnoses in a cytopathology laboratory. METHODS: Neoplasia-associated genetic alterations (NGAs) were determined by MT of preoperative FNA biopsies or resected malignancies and were categorized as BRAF V600E mutations, RAS-like mutations (HRAS, NRAS, or KRAS mutations or non-V600E BRAF mutations), or other mutations. RESULTS: Among 7382 thyroid FNA biopsies, the AUS rate was 9.3% overall and ranged from 4.3% to 24.2% among 6 cytopathologists (CPs) who evaluated >150 cases. The ratio of specimens falling into TBS category III to specimens falling into category VI (malignant) (the III:VI ratio) was 2.4 overall (range, 1.1-8.1), and the ratio of specimens falling into TBS categories III and IV (follicular neoplasm or suspicious for follicular neoplasm) combined (III+IV) to specimens falling into category VI (the [III+IV]:VI ratio) was 2.9 overall (range, 1.4-9.5). MT was performed on 588 cases from 560 patients (79% women) with a median age of 56 years (range, 8-89 years). BRAF V600E mutation was the most common (76% of cases) in TBS category VI and was rare (3%) in category III. RAS-like mutations were most common in TBS categories III (13%), IV (25%), and V (suspicious for malignancy) (17.5%). The NGA rate in AUS cases fell between 5% and 20% for 5 of 6 CPs and did not correlate with the III:VI ratio or the (III+IV):VI ratio. CONCLUSIONS: Lack of correlation between the NGA rate and easily calculable diagnostic ratios enables the calibration of diagnostic thresholds, even for CPs who have normal metrics. Specifically, calculation of the NGA rate and the III:VI ratio may allow individual CPs to determine whether they are overcalling or undercalling cases that other CPs might otherwise recategorize.

Nuclear hypochromasia: Shedding light on the "lightness" of high-grade urothelial carcinoma.

BACKGROUND: The Paris system (TPS) has provided a standardized classification for reporting urinary cytology, specifically high-grade urothelial carcinoma (HGUC). While hyperchromasia is well described in HGUC, there exists little data on nuclear hypochromasia in HGUC. Our focus was to investigate the incidence of hypochromasia in HGUC and if it should become a criterion for HGUC. DESIGN: All cases of HGUC at our institution for a 3-year interval (2017-2019) using TPS were reviewed. Each case had a single ThinPrep slide and concurrent biopsy or resection specimen to confirm the diagnosis within 30 days. The presence of hypochromasia was evaluated, and cases with hypochromasia were stratified based on the tumor cell percentage. Cases with hypochromasia in 5% or greater of the tumor cells were considered "positively identified" for hypochromasia. RESULTS: We reviewed 117 cases of HGUC and identified nuclear hypochromasia in 12 cases (10.2%) within 5% or greater of the tumor cells. These 12 cases were further assessed based on if tumor cells showed hypochromasia in 5%-49% of the sample, or greater than 50% of the sample. Hypochromasia in 5%-49% of tumor cells was present in 8/117 cases (6.8%); whereas in 50% or greater samples 4/117 cases (3.4%) showed hypochromasia. No cases were identified where hypochromasia was noted in less than 5% of the tumor cells. CONCLUSION: TPS and use of hyperchromasia as a feature of HGUC is affirmed. However, hypochromasia, while not diagnostic in isolation, is present in a sub-set of patients with HGUC (10.2%) and should be considered as a variance noted in the nuclei of HGUC.