High-risk human papillomavirus testing in cytology aspiration samples from the head and neck part 1: a review of the literature on available testing options.

Human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma is increasing in incidence and is often first diagnosed on a cytology fine needle aspiration (FNA) specimen of metastatic nodal disease of the neck. In the setting of oropharyngeal squamous cell carcinoma, HPV status defines the disease with HPV-associated tumors having better overall prognosis than those that are HPV negative. Furthermore, metastatic squamous cell carcinoma of the neck of unknown origin requires testing for HPV as a positive result suggests an oropharyngeal primary. As a result, HPV testing in aspirate samples is increasingly important for the proper diagnosis and treatment of patients with head and neck squamous cell carcinoma. Although HPV testing in cervicovaginal cytology specimens is common and well-established, testing in head and neck FNA samples remains challenging. p16 immunohistochemistry is an excellent surrogate marker for HPV in tumors of known or suspected oropharyngeal origin, but the criteria used in histologic specimens may not be appropriate in cytology samples. FNA samples are more frequently hypocellular, and cytology cell blocks have variable fixation and processing steps, limiting the utility of p16 immunohistochemistry. Other potential testing options have been reported in the literature including staining of aspirate smears and molecular testing of liquid-based samples. The American Society of Cytopathology Clinical Practice Committee recently surveyed the American Society of Cytopathology membership to determine the current state of HPV testing in aspirate samples, and this review article is designed to provide a summary of the current literature on various testing options in FNA samples.

Results from the 2019 American Society of Cytopathology survey on rapid on-site evaluation-Part 1: objective practice patterns.

INTRODUCTION: Rapid on-site evaluation (ROSE) is a service provided by cytologists that helps ensure specimen adequacy and appropriate triage for ancillary testing. However, data on the current usage patterns across different practice settings have been lacking. MATERIALS AND METHODS: To obtain an accurate and timely assessment of the current state of practice of ROSE, a 14-question online survey was constructed by the Clinical Practice Committee of the American Society for Cytopathology. The survey was available to the membership of the American Society for Cytopathology for a 3-week period in early 2019. RESULTS: A total of 541 responses were received, including from 255 cytopathologists/pathologists, 261 cytotechnologists, 19 cytology resident/fellow trainees, and 6 others. ROSE was offered as a clinical service by 95.4% of the respondents, with telecytology for ROSE used in 21.9% of the practices. Endobronchial ultrasound-guided transbronchial needle aspiration was the procedure most frequently reported to use ROSE (mean, 59.1%; median, 70%). Cytotechnologists were involved in ROSE in most practices. The number of daily ROSE procedures correlated with the annual nongynecologic cytology volumes. Approximately 70% of ROSE procedures were reported to require >30 minutes, on average, for the cytologist. CONCLUSIONS: The results from our survey of cytologists have shown that the reported practice patterns for the usage of ROSE vary considerably. The presented data can help inform future guideline recommendations and the implementation of ROSE in different clinical settings.

Cytological diagnosis of papillary thyroid carcinoma with tall cells on ThinPrep liquid-based cytology.

BACKGROUND: The tall cell variant of papillary thyroid carcinoma (PTC-TC) has been associated with aggressive features including extrathyroidal extension, higher rate of lymph node and distant metastases, and higher recurrence rate. We aimed to evaluate the cytomorphologic features of PTC-TC on ThinPrep (TP) along with its diagnostic efficacy to detect PTC-TC. METHODS: Preoperative cytology samples from 30 cases of histologically-proven PTC-TC and 30 classical PTC controls were selected for this study. TP preparations were evaluated for varying architectural and cytomorphologic features. RESULTS: Tall cells were present in the majority of PTC-TC cases and were located at the periphery of cell clusters and as single cells. Cytoplasmic cuff along the periphery of cell clusters and soap-bubble pseudoinclusions were very specific features of PTC-TC, when present. PTC-TC cases were more likely to show abundant oncocytic cytoplasm and distinct cell borders. Cytoplasmic tails were more likely to be present and more numerous in PTC-TC. The presence of nuclear grooves, papillary architecture, and giant cells were not reliable distinguishing features of PTC-TC vs controls. CONCLUSION: Our results indicate that tall cell cytomorphologic and architectural features in PTC are identifiable on TP.

Decreased UCHL1 expression as a cytologic biomarker for aggressive behavior in pancreatic neuroendocrine tumors.

BACKGROUND: There are currently no reliable markers associated with aggressive behavior in well-differentiated and moderately differentiated pancreatic neuroendocrine tumors. We aimed to determine whether expression of ubiquitin carboxyl-terminal hydrolase L1 in conjunction with Ki67 can identify metastatic potential of well-differentiated and moderately differentiated pancreatic neuroendocrine tumors from fine-needle aspiration samples obtained by endoscopic ultrasound. METHODS: Retrospective review of 48 patients with well-differentiated and moderately differentiated pancreatic neuroendocrine tumors diagnosed by endoscopic ultrasound fine-needle aspiration at a single center identified 35 biopsy samples with adequate material for analysis. Ubiquitin carboxyl-terminal esterase L1 immunocytochemistry of primary pancreatic neuroendocrine tumors was performed along with Ki67 staining and scored semiquantitatively. The combination of ubiquitin carboxyl-terminal esterase L1 score ≤4 (weak) and Ki67 ≥3% (high) was considered a positive test for predicting tumors associated with metastases. RESULTS: Weak ubiquitin carboxyl-terminal hydrolase L1 staining had 80% sensitivity, 65% specificity, 63% positive predictive value, and 81% negative predictive value to identify primary tumors associated with metastatic disease. The combination of weak ubiquitin carboxyl-terminal hydrolase L1 staining and high Ki67 staining increased the test specificity to 95%. On multivariable analysis, combined positive test of weak ubiquitin carboxyl-terminal esterase L1 staining and high Ki67 staining was an independent predictor of metastatic disease (P = .047). CONCLUSION: Ubiquitin carboxyl-terminal hydrolase L1 is a novel biomarker for identifying malignant potential of primary well-differentiated and moderately differentiated pancreatic neuroendocrine tumors and in combination with Ki67 is an independent predictor of development of metastatic disease.

Accuracy of next-generation sequencing for the identification of clinically relevant variants in cytology smears in lung adenocarcinoma.

BACKGROUND: Minimally invasive diagnostic procedures such as needle-core biopsy and fine-needle aspiration provide adequate material for molecular analyses. Advances in precision oncology are trending toward the interrogation of limited amounts of genomic material to guide clinical and therapeutic decisions. The aim of this study was to investigate the minimum cellularity needed on cytologic smears for the identification of clinically relevant variants with next-generation sequencing (NGS). METHODS: Thirty cases of cytologically diagnosed, resection-proven primary lung adenocarcinoma were identified. Nineteen of the 30 cases were known to harbor actionable variants. One Diff-Quik (DQ)-stained slide and 1 Papanicolaou (Pap)-stained slide were selected from each case. Cases were categorized as containing fewer than 100 tumor cells, 100 to 500 tumor cells, or more than 500 tumor cells. NGS was performed on the Ion Torrent platform. RESULTS: NGS was successfully performed on all cell blocks and on 90% of the smears. Paired DQ and Pap smears showed similar cellularity, and cases that differed in cellularity were within 1 category of each other. The cases with more than 100 tumor cells had a 93% success rate; this was significantly different from the situation for cases with fewer than 100 tumor cells, which were successfully sequenced only 67% of the time. Overall, NGS was able to provide clinically relevant information for 83% of DQ smears and for 90% of Pap smears tested. CONCLUSIONS: The data show a significantly higher likelihood of successful NGS with cytologic smears with more than 100 tumor cells. There was a trend for a higher NGS success rate with Pap smears versus DQ smears. Cancer Cytopathol 2017;125:398-406. © 2017 American Cancer Society.

Diagnostic yield of cytopathology in evaluating pericardial effusions: Clinicopathologic analysis of 419 specimens.

BACKGROUND: Pericardial effusions can cause considerable morbidity and potentially may lead to mortality. Malignant pericardial effusions are uncommon, and data on malignancies encountered in pericardial effusion cytology specimens are limited. METHODS: Relevant records of all pericardial effusions from January 2008 to September 2014 were examined and compared with pericardial biopsy results when performed. Discrepant cases were reviewed to determine the cause of the disagreement. RESULTS: In total, 419 pericardial effusion specimens obtained from 364 patients were examined. Cytologic diagnostic categories included: negative for malignancy (332 specimens; 79%), equivocal (25 specimens; 6%), and positive (62 specimens from 51 patients; 15%). Forty-seven patients who had positive effusions were known to have malignancy. The most common primary malignancies were breast (39.3%) and lung (39.3%) cancers in women and lung cancer (47.4%) in men. A concurrent pericardial biopsy was performed in 46% of patients. Excluding equivocal cytologic diagnoses, cytology and biopsy were concordant in 153 of 173 paired samples (88.4%). The sensitivity of cytology in diagnosing malignancy was 92.1% compared with 55.3% for pericardial biopsy. CONCLUSIONS: Cytologic examination has significant diagnostic utility in the evaluation of pericardial effusions and exhibits a lower false-negative rate compared with pericardial biopsy. Submission of pericardial biopsy alongside effusion cytology is associated with increased sensitivity for detecting malignancy and may be especially useful in the setting of low-volume pericardial effusion. Cancer Cytopathol 2017;125:128-137. © 2016 American Cancer Society.

Cytology Preparations of Formalin Fixative Aid Detection of Giardia in Duodenal Biopsy Samples.

Giardiasis is the most common intestinal parasitic infection in the United States. The organism elicits no, or minimal, inflammatory changes in duodenal biopsy samples, so it can be easily overlooked. We performed this study to determine whether Giardia could be isolated from the formalin fixative of biopsy samples, and to evaluate the value of fluid analysis in the assessment for potential infection. We prospectively evaluated duodenal biopsy samples from 92 patients with a clinical suspicion of giardiasis or symptoms compatible with that diagnosis (ie, diarrhea, bloating, or abdominal pain) Biopsy samples were routinely processed and stained with hematoxylin and eosin. Histologic diagnoses included giardiasis (5 cases, 4%), normal findings (64 cases, 70%), peptic injury/active duodenitis (12 cases, 13%), and intraepithelial lymphocytosis with villous blunting (10 cases, 12%). Fifteen cases (13%) showed detached degenerated epithelial cells or mucus droplets in the intervillous space that resembled Giardia. Cytology slides were prepared from formalin in the biopsy container using the standard Cytospin protocol and reviewed by a cytopathologist blinded to the biopsy findings. Cytologic evaluation revealed Giardia spp. in all 5 biopsy-proven cases, and identified an additional case that was not detected by biopsy analysis. Organisms were significantly more numerous (mean: 400 trophozoites; range, 120 to 810) and showed better morphologic features in cytology preparations compared with tissue sections (mean: 129 trophozoites; range, 37 to 253 organisms; P=0.05). Our findings suggest that cytology preparations from formalin fixative can resolve diagnostically challenging cases and even enhance Giardia detection in some cases.

Metastatic Breast Carcinoma in Cerebrospinal Fluid: A Cytopathological Review of 15 Cases.

The incidence of metastatic carcinoma to the meninges ("meningeal carcinomatosis" [MC]) is increasing due to longer survival of patients and improved imaging techniques. Currently, MC is best diagnosed by cytopathological evaluation of cerebrospinal fluid (CSF). Breast primaries are the commonest cause of MC; although cytopathological features thereof have not been, as yet, fully characterized. In this study of meningeal mammary carcinomatosis, relevant clinicopathological data and archived cytopathological preparations of all "suspicious" and "positive" CSF specimens (1992-2015), from patients with a history of breast carcinoma, were retrieved and reviewed. Twenty-three "positive" CSF specimens, derived from 15 patients formed the basis of this study. All specimens were processed as Cytospin preparations, and stained by Papanicolaou and Diff-Quik techniques. All patients were female, with a mean age of 57 (range: 32-85) years. Mean interval between initial diagnosis of breast carcinoma and "positive" CSF was 32 (range 6-84) months. All 23 specimens (100%) were "cellular" (>10 carcinoma cells). Eighteen (78%) specimens showed only isolated nonclustered cells, and 5 (22%) specimens showed both cell clusters and isolated cells. In most "positive" cases, metastatic breast carcinoma cells showed variation in cell size, with hyperchromatic nuclei, irregular nuclear membranes, prominent nucleoli and cytoplasmic vacuolization. The background in some CSF samples showed red blood cells and fibrin admixed with rare lymphocytes and histiocytes. One specimen showed necrotic debris. Papanicolaou and Diff-Quik-stained Cytospin preparations were equally diagnostic, as the aforementioned findings were present in both types of preparation.

Committee I: Indications for pulmonary cytology sampling methods.

The Papanicolaou Society of Cytopathology has developed a set of guidelines for pulmonary cytology including indications for bronchial brushings, washings and endobronchial ultrasound-guided fine needle aspiration, technical recommendations for cytologic sampling, recommended terminology and classification scheme, recommendations for ancillary testing and recommendations for postcytologic diagnosis management and follow-up. All recommendation documents are based on the expertise of the authors, extensive literature review and feedback from presentations at national and international conferences. This document selectively presents the results of these discussions. The present document summarizes the recommendations for clinical and imaging evaluation of pulmonary lesions along with the indications for cytologic studies regarding these abnormalities. Preprocedural requirements regarding brushing, washing and needle aspiration procedures are discussed also. Diagn. Cytopathol. 2016;44:1010-1023. © 2016 Wiley Periodicals, Inc.

Diagnostic value of positive urinary cytology in the detection of recurrent urothelial carcinoma: 10-Year Experience at the Papanicolaou Cytology Laboratory.

INTRODUCTION: Urothelial carcinoma (UCC) of the bladder is the most common malignancy of the urinary tract. The performance of urine cytology (UCy) after radical cystectomy (RC) and urinary diversion is quite variable and there is no consensus regarding its role in post-treatment surveillance. The goal of this study is to retrospectively review the diagnostic value of positive (suspicious or positive for malignancy) diverted urine cytology (DUCy) in the detection of urinary tract recurrence of UCC. MATERIALS AND METHODS: A retrospective 10-year (January 2005 to January 2015) computerized search for all DUCy specimens was conducted. All suspicious (Susp) or positive for malignant cells (PMC) ThinPrep cases were identified and retrospectively reviewed. Clinical, surgical, and pathological follow-up for patients with Susp or PMC UCy were tabulated and analyzed. RESULTS: During the 10-year-period, 1,525 DUCy cases from 408 patients were identified. Of these, 25 cases (1.64%) from 10 patients were called either Susp (13; 0.85%) or PMC (12; 0.79%). The 25 DUCy cases occurred within a mean of 20 months post-RC. Out of 10 patients, 9 had a concurrent biopsy or a subsequent resection of the recurrent site. Of these 9 patients, 8 (89%) had subsequent biopsy or resection, which showed recurrent UCC. In 5/8 patients, positive DUCy was the very first manifestation of recurrence, which was subsequently confirmed by imaging or histology. CONCLUSION: It is our experience that patients with positive UCy after RC have a high likelihood of recurrent UCC and should be counseled and managed accordingly. Diagn. Cytopathol. 2016;44:975-979. © 2016 Wiley Periodicals, Inc.

Detection of in situ and invasive endocervical adenocarcinoma on ThinPrep Pap Test: Morphologic analysis of false negative cases.

BACKGROUND: The goal of this study was to calculate the sensitivity and false negative (FN) rate of ThinPrep Pap Test (TPPT) and carefully analyze missed cases for educational purposes. MATERIALS AND METHODS: Patients with histologically proven adenocarcinoma in-situ (AIS) or invasive endocervical adenocarcinoma (EAC) over a 17-year-period (1998-2015) were identified. The TPPT immediately preceding the histological diagnosis of AIS/ECA was designated as index Pap (IP). Paps up to 122 months before histologic diagnosis of AIS/ECA were considered for this study. All available negative and unsatisfactory TPPT were re-reviewed. RESULTS: There were 78 patients with histologically-proven AIS (56) or ECA (22) with 184 TPPTs, and 95 of these TPPTs were abnormal. Of the abnormal cases, 55.7% TPPTs were diagnosed as endocervical cell abnormality (atypical endocervical cells/AIS/ECA). Notably, 44.2% of abnormal TPPTs were diagnosed as squamous cell abnormality (atypical squamous cells of undetermined significance/low-grade squamous intraepithelial lesion/high grade squamous intraepithelial lesion). Including the diagnoses of squamous cell abnormality, the sensitivity of index TPPT for histologically-confirmed AIS/ECA was 88%. Eighty-eight of 184 TPPT, including 10 IP, were negative = 87, or unsatisfactory = 1. Forty-two of these slides were available for re-review. Upon review, 21 TPPT (50%) were confirmed negative and 21 TPPT (50%) were reclassified as abnormal = 20, or unsatisfactory = 1. Of the FN cases, the main difficulty in correct diagnosis was the presence of few diagnostic cell clusters which had less feathering, and consisted of smaller, rounder cells in small and tighter clusters, with nuclear overlap. In particular, nuclear overlap in three-dimensional groups precluded the accurate diagnosis. Rare FN cases showed squamous cell abnormality on re-review, and rare cases showed obscuring blood or inflammation. CONCLUSION: A significant proportion of AIS/EAC is discovered after Pap showing squamous cell abnormality. FN cases were most commonly related to nuclear overlap in tight three-dimensional clusters.

"Intercellular bridges" in a case of well differentiated squamous carcinoma.

Intercellular bridges may aide in definitive identification of malignant cell origin, especially in squamous cell carcinoma. They are difficult to identify in routine cytologic specimens and are especially rare in smear preparations. Herein, we present images of intercellular bridges from a case of well differentiated squamous cell carcinoma of the esophagus in a cytologic specimen obtained from FNA of a paraesophageal lymph node.

High grade endometrial stromal sarcoma on thinprep.

High grade endometrial stromal sarcoma (HGESS) is an uncommon malignancy recently re-defined in the new WHO classification of endometrial stromal tumors. In this article, we discuss the differential diagnoses of metastatic HGESS in a fine needle aspiration (FNA) of a lymph node and compare the cytomorphology of HGESS in ThinPrep [(TP), Hologic Inc., Boxborough, MA] to conventional smears (CS). The patient had a history of stage I HGESS, status-post supracervical hysterectomy without regional lymph node metastases. Her post-operative course was complicated by pelvic fluid collections and enlarging para-aortic lymph nodes. Diff-Quik (DQ)-stained and Papanicolaou (Pap)-stained smears from a para-aortic lymph node FNA demonstrated a cellular specimen with monomorphic population of plump to oval cells with scant, wispy cytoplasm in aggregates and as single cells. The nuclei showed fine chromatin and small inconspicuous nucleoli. Compared to the CS, HGESS cells in the TP showed similar cytological features, with the exception that the nuclei were slightly smaller, hyperchromatic, and the chromatin pattern was attenuated. In the absence of prior clinical history, the cytomorphology of metastatic HGESS in a lymph node can be difficult to differentiate from a lymphoma, a variety of metastatic spindle cell tumors or metastatic carcinoma. Immunohistochemical analysis and comparison with the primary tumor can be useful in proving the nature of the malignant cells. The cytomorphology of HGESS on TP correlated well in both single cells and aggregates when compared to CS. The differences noted were decreased nuclear size, nuclear hyperchromasia, and slightly attenuated nuclear detail on TP.

Negative computer-imaged ThinPrep Pap test and positive hybrid capture2 HPV co-testing results: A quality assurance review.

Women ≥30 years of age with negative (-) Pap tests and positive (+) HPV co-test results have a higher prevalence and cumulative risk of developing high-grade cervical intraepithelial neoplasia (CIN 2+). Thus, the current management in these women is to repeat co-test in 12 months or immediate reflex genotyping for HPV16 or HPV 16/18. If genotyping is not an option, timely quality assurance (QA) rescreen of such Pap tests may be a valuable alternative. All ThinPrep Pap tests (TPPT) interpreted as negative for intra epithelial lesion (NILM) or NILM with reactive cellular changes (NILM/RCC) and a (+) high-risk HPV [Hybrid Capture 2 (HC2), Qiagen, Hilden, Germany] co-test result over a 45-month period (10/2009-06/2013) underwent monthly QA review. The TPPT were screened by the TP Imaging System [TIS, Hologic Inc., Bedford, MA]. Twenty five thousand six hundred and seventy five (18%) NILM and NILM/RCC TPPT of a total of 141,548 TPPT underwent HPV co-test. HPV test was (+) in 2,300 (8.9%) TPPT cases. HPV (+) cases by age group were <30 years, 486 (21%), and ≥30 years, 1,814 (79%). Upon QA review, 10 cases (0.4%) were reclassified, with significant findings in three cases in ≥30 years. Two cases showed high-grade squamous intraepithelial lesion (HSIL) on repeat Pap, and one case showed endocervical adenocarcinoma in situ (AIS) on biopsy. Timely QA review of HPV (+) Pap (-) co-tests is a valuable monitor. Ninety percentage of reclassified cases were in ≥30 age group and 70% were originally signed out by using TIS 22 Field of View (FOV) only. Three reclassified cases had significant findings on follow up (F/U).