Endocervical glandular involvement is associated with an increased detection rate of high-grade squamous intraepithelial lesions on the Papanicolaou test.

INTRODUCTION: Although The Bethesda System for Reporting Cervical Cytopathology does not mandate reporting of endocervical glandular involvement (EGI) in Papanicolaou test specimens with high-grade squamous intraepithelial lesions (HSIL), several studies have suggested that EGI diagnosed on surgical specimens is associated with higher rates of residual or recurrent dysplasia. When suspected, EGI is reported for Papanicolaou test specimens at our institution, but the performance of this diagnosis has not been assessed. MATERIALS AND METHODS: The archives were queried for Papanicolaou test specimens with a diagnosis of HSIL-EGI (2006-2017). All follow-up surgical pathology specimens within a year of the Papanicolaou test diagnosis were evaluated for cytologic-histologic correlation. This same query was repeated for all surgical pathology specimens with a diagnosis of HSIL-EGI. All preceding Papanicolaou test diagnoses within a year were assessed for cytologic-histologic correlation. Twenty Papanicolaou test specimen glass slides were reviewed by 6 observers to assess for interobserver variability. RESULTS: Patients with HSIL-EGI on surgical specimens were more likely to have a preceding Papanicolaou diagnosis of HSIL and atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (ASC-H) (32.3% versus 25.5%, P = 0.03, and 16.7% versus 11.8%, P = 0.04, respectively). Patients with an HSIL-EGI diagnosis on a Papanicolaou test were significantly more likely to have HSIL-EGI detected on a follow-up histology (41.6% versus 24.0%, P < 0.001). Interobserver concordance was poor for the assignment of EGI in Papanicolaou test specimens. CONCLUSIONS: Overall, the diagnosis of HSIL-EGI on Papanicolaou test specimens is complicated by poor sensitivity and interobserver concordance.

Variability among observers utilizing the CellSolutions BestCyte Cell Sorter imaging system for the assessment of urinary tract cytology specimens.

INTRODUCTION: Image analysis systems are not currently commonly used for evaluating urinary cytology specimens. We evaluated whether the BestCyte Cell Sorter (CellSolutions, Greensboro, NC) imaging system can reliably identify atypical cells in urinary cytology specimens. METHODS: Fifty-three consecutive urine cytology specimens underwent 2 preparations: one slide using SurePath (SP; BD Diagnostics, Sparks, MD)™ for routine clinical evaluation, and a second slide using the CellSolutions F50 system for analysis by the BestCyte Cell Sorter (BCCS) scanning system. Eight observers reviewed atypical cells flagged by BCCS and assigned a BCCS diagnosis to each of the 53 specimens. The observers also blindly reviewed the SP preparation (when available) and assigned an SP diagnosis. The SP diagnoses given by one "expert" observer was considered as a reference diagnosis. RESULTS: There was fair-to-moderate agreement among observers for identifying any atypia and high-grade atypia (Fleiss kappa: 0.417 and 0.338, respectively) using BCCS. Review of SP preparations had slightly better agreement (Fleiss kappa: 0.558 and 0.564, respectively). Intraobserver agreement between the two methods varied greatly between individuals (Cohen's kappa range: 0.260 to 0.647). When a consensus diagnosis could be reached among the observers for cases with surgical follow-up, the consensus diagnosis was concordant in 11 of 12 instances, with one instance being a one-step discrepancy. CONCLUSIONS: Specimen review by BCCS resulted in slightly greater interobserver variability than review of routine SP preparations. This may have been due to variations in observer experience and comfort with the use of a digital imaging system, which is further suggested by the wide range of intraobserver agreement among individuals.

cancer of unknown primary: Ancillary testing of cytologic and small biopsy specimens in the era of targeted therapy.

Initial evaluation of patients presenting with a suspected malignancy without a clear clinical or radiologic impression of a primary site (so-called cancer of unknown primary, or CUP) frequently includes fine needle aspiration (FNA) to both establish a diagnosis and procure tissue for additional studies. Careful management of limited specimen material is especially important in the modern era, when molecular studies may take precedence over a more definitive tissue diagnosis. This review summarizes our practice habits when encountering a CUP. Broadly, key morphologic patterns in context of patient history and clinical-radiologic impression should guide the selection of a limited panel of IPOX stains to establish first a broad lineage. Subsequently 1-2 site-specific markers may be used to confirm the diagnosis. In difficult cases, it is helpful to convey the situation to the treating physicians, who may prefer to prioritize molecular testing over pursuit of a more definitive diagnosis. Prudent choice of immunoperoxidase (IPOX) studies, based on both key cytomorphologic criteria and continued communications with the clinical team regarding their needs for treatment planning, is critical in CUP cases.

Evaluation of Sienna Cancer Diagnostics hTERT Antibody on 500 Consecutive Urinary Tract Specimens.

OBJECTIVES: Telomerase activity can be detected in up to 90% of urothelial carcinomas (UC). Telomerase activity can also be detected in urinary tract cytology (UTC) specimens and indicate an increased risk of UC. We evaluated the performance of a commercially available antibody that putatively binds the telomerase reverse transcriptase (hTERT) subunit on 500 UTC specimens. STUDY DESIGN: Unstained CytospinTM preparations were created from residual urine specimens and were stained using the anti-hTERT antibody (SCD-A7). Two algorithms were developed for concatenating the hTERT result and cytologic diagnosis: a "no indeterminates algorithm," in which a negative cytology and positive hTERT result are considered positive, and a "high-specificity algorithm," in which a negative cytology and positive hTERT result are considered indeterminate (and thus negative for comparison to the gold standard). RESULTS: The "no indeterminates algorithm" and "high-specificity algorithm" yielded a sensitivity of 60.6 and 52.1%, a specificity of 70.4 and 90.7%, a positive predictive value of 39.1 and 63.8%, and a negative predictive value of 85.0 and 85.8%, respectively. CONCLUSIONS: A positive hTERT result may identify a subset of patients with an increased risk of high-grade UC (HGUC) who may otherwise not be closely followed, while a negative hTERT immunocytochemistry result is associated with a reduction in risk for HGUC.

The Paris System for Reporting Urinary Cytology: early review of the literature reveals successes and rare shortcomings.

The Paris System for Reporting Urinary Cytology (TPS) provides recommendations for the diagnosis of urinary tract cytology (UTC) specimens and has found acceptance on an international level. Since the official release of TPS in 2016, numerous research studies have been published analyzing its impact. This review summarizes the studies published since the release of TPS, highlighting areas in which TPS has performed well and other areas in which TPS may need improvement.

MAML2 Rearrangements in Variant Forms of Mucoepidermoid Carcinoma: Ancillary Diagnostic Testing for the Ciliated and Warthin-like Variants.

Mucoepidermoid carcinoma (MEC) is the most common salivary gland malignancy. Recent studies have shown that most MECs harbor gene fusions involving MAML2-an alteration that appears to be specific for MEC, a finding that could be diagnostically useful. While most cases of MEC are histologically straightforward, uncommon variants can cause considerable diagnostic difficulty. We present 2 variants of MEC for which MAML2 studies were crucial in establishing a diagnosis: a previously undescribed ciliated variant, and the recently described Warthin-like variant. All cases of ciliated and Warthin-like MEC were retrieved from the archives of The Johns Hopkins Hospital. Break-apart fluorescence in situ hybridization for MAML2 was performed on all cases. One ciliated MEC and 6 Warthin-like MECs were identified. The ciliated MEC presented as a 4.6 cm cystic lymph node metastasis originating from the tongue base in a 47-year-old woman. The Warthin-like MECs presented as parotid masses ranging in size from 1.2 to 3.3 (mean, 2.7 cm) in 4 women and 2 men. The ciliated MEC consisted of macrocystic spaces punctuated by tubulopapillary proliferations of squamoid cells and ciliated columnar cells. The Warthin-like MECs were comprised of cystic spaces lined by multilayered oncocytic to squamoid cells surrounded by a circumscribed cuff of lymphoid tissue with germinal centers. In these cases, the Warthin-like areas dominated the histologic picture. Conventional MEC, when present, represented a minor tumor component. MAML2 rearrangements were identified in all cases. Warthin-like MEC, and now a ciliated form of MEC, are newly described variants of a common salivary gland carcinoma. Unfamiliarity with these novel forms, unanticipated cellular features (eg, cilia), and morphologic overlap with mundane benign processes (eg, developmental ciliated cysts, Warthin tumor) or other carcinomas (eg, ciliated human papillomavirus-related carcinoma) may render these variants susceptible to misdiagnosis. These unusual variants appear to consistently harbor MAML2 fusions-a finding that establishes a clear link to conventional MEC and provides a valuable adjunct in establishing the diagnosis.

Improved risk stratification for patients with high-grade urothelial carcinoma following application of the Paris System for Reporting Urinary Cytology.

BACKGROUND: The Paris System for Reporting Urinary Cytology (TPS) requires 4 cytomorphologic criteria for a definitive diagnosis of high-grade urothelial carcinoma (HGUC) in urinary tract cytology (UTC) specimens: an elevated nuclear-to-cytoplasmic (N/C) ratio (at or above 0.7), markedly atypical nuclear borders, moderate to severe hyperchromasia, and coarse chromatin. However, malignant UTC specimens often contain degenerative changes, and this limits the number of malignant cells meeting all 4 TPS cytomorphologic criteria. METHODS: One hundred twelve UTC specimens from patients with a subsequent diagnosis of HGUC were reviewed and reclassified according to TPS criteria. The presence of TPS cytomorphologic criteria for HGUC in each specimen was recorded, as was the proportion of atypical cells meeting all 4 criteria. RESULTS: The number of specimens definitively diagnosed as HGUC did not significantly change upon reclassification. However, approximately 40% of indeterminate specimens (21 of 51) were reclassified into a higher risk category. The most restrictive cytomorphologic criterion was an N/C ratio of 0.7 or higher (seen in 78% of specimens), and approximately half of specimens containing all 4 cytomorphologic criteria did not meet TPS's numerical criterion for HGUC (at least 5 malignant cells). In the majority of specimens qualifying for HGUC by TPS standards, only a small fraction of atypical cells (10%-20%) met all the criteria. CONCLUSIONS: When applied to malignant UTC specimens, TPS criteria improved specimen risk stratification by upgrading approximately 40% of indeterminate specimens into higher risk categories while not significantly changing the frequency of HGUC diagnoses. Cancer Cytopathol 2017;125:427-34. © 2017 American Cancer Society.

Detection of TERT promoter mutations in primary adenocarcinoma of the urinary bladder.

TERT promoter mutations (TERT-mut) have been detected in 60% to 80% of urothelial carcinomas. A molecular urine-based screening assay for the detection of TERT-mut is currently being pursued by our group and others. A small but significant number of bladder carcinomas are adenocarcinoma. The current study assesses the incidence of TERT-mut in primary adenocarcinomas of urinary bladder. A retrospective search of our institutional pathology records identified 23 cystectomy specimens with a diagnosis of adenocarcinoma (2000-2014). All slides were reviewed by a senior urologic pathologist to confirm tumor type and select a representative formalin-fixed, paraffin-embedded block for mutational analysis. Adequate material for DNA testing was available in 14 cases (7 enteric type and 7 not otherwise specified). TERT-mut sequencing analysis was performed using previously described SafeSeq technique. Overall, 28.5% of primary adenocarcinoma harbored TERT-mut. Interestingly, 57% of nonenteric adenocarcinomas were mutation positive, whereas none of the enteric-type tumors harbored mutations. Similar to urothelial carcinoma, we found a relatively higher rate of TERT-mut among nonenteric-type adenocarcinomas further supporting the potential utility of TERT-mut urine-based screening assay for bladder cancer.

Mesothelin Expression in Advanced Gastroesophageal Cancer Represents a Novel Target for Immunotherapy.

The identification of new therapeutic targets is of profound importance if we are to improve outcomes in gastroesophageal cancer. This study assessed the rate of mesothelin expression in tumors of western patients with upper gastrointestinal tract carcinomas. In addition, the AGS gastric cancer cell line was tested for sensitivity to SS1(dsFv)PE38, a mesothelin-targeting immunotoxin. Previously constructed tissue microarrays containing samples from 127 patients with gastroesophageal adenocarcinomas were examined by immunohistochemistry (IHC) for mesothelin expression. Labeling for HER2-neu, E-cadherin, and c-met were also assessed. Tumors were considered positive for mesothelin if at least moderate cytoplasmic/membranous or luminal staining was present in minimum 10% of the neoplastic cells. The AGS gastric cancer cell line was assessed for surface mesothelin expression by flow cytometry and the viability of cells treated with SS1P was measured. Gastroesophageal cancers were mesothelin positive in 64 of 127 tumors [50.4%; 95% confidence interval (CI), 41.4%-59.4%], whereas only 9 carcinomas (7.1%; 95% CI, 3.3%-13.0%) were HER2-neu IHC 3+ positive and 8 (6.6%; 95% CI, 2.9%-12.5%) were c-met positive. Mesothelin expression increased from stage I to stage IV tumors (37.5% to 56.3%, respectively, P=0.10). The AGS gastric cancer cell line was sensitive to the immunotoxin with an EC50 value in the low picomolar range (0.4 ng/mL). A gastric cancer cell line derived from a western patient was exquisitely sensitive to the mesothelin-targeted immunotoxin SS1P. Clinical trials involving novel mesothelin targeted immunotherapeutics in gastroesophageal cancer are currently in development.

Benign and low-grade fibroepithelial neoplasms of the breast have low recurrence rate after positive surgical margins.

Breast phyllodes tumors are uncommon fibroepithelial neoplasms with a range of histologic features. Surgical excision is the primary management, but the need for excision to negative margins in benign and borderline phyllodes tumors is unclear. Here, we review the surgical management patterns and outcomes of 90 patients with benign and low-grade fibroepithelial lesions of the breast treated at our institution, including 19 borderline phyllodes tumors, 52 benign phyllodes tumors, and 19 representative neoplasms with overlapping features of fibroadenoma and benign phyllodes tumors, which were classified as 'fibroadenomas with phyllodal features'. In total, 52 (58%) had positive surgical margins on first excision, and of these 17 (33%) underwent re-excision to achieve negative margins. Residual tumor was identified in three (18%) re-excisions. Patients with fibroadenoma with phyllodal features were more likely to have a positive surgical margin than with benign phyllodes tumors or borderline phyllodes tumors (89 vs 49%, P=0.0015), and were less likely to undergo re-excision for positive margins (12 vs 43%, P=0.031). In total, there were three recurrences (3%), with one per fibroadenoma with phyllodal features, benign phyllodes tumor, and borderline phyllodes tumor. There was no statistically significant difference in recurrence rates between patients with positive or negative margins, or between patients with positive margin with or without re-excision. The extent of the positive margin did not predict recurrence. In conclusion, the recurrence rate of benign and low-grade fibroepithelial lesions is low and not associated with the original margin status. Patients with fibroadenomas with phyllodal features, benign phyllodes tumors, or selected borderline phyllodes tumors and positive margins on initial excision may be managed conservatively, with close follow-up and timely re-excision of any potential recurrence.

Low-Grade Fibromyxoid Sarcoma of the Head and Neck: A Clinicopathologic Series and Review of the Literature.

Low-grade fibromyxoid sarcoma (LGFMS) is a deceptively bland malignancy with potential for late recurrence and metastasis, which usually occurs in the deep soft tissues of the extremities and trunk. Most LGFMSs harbor a characteristic gene fusion of FUS-CREB3L2, and recently MUC4 immunostaining has been found to be highly sensitive and specific for the diagnosis. We present a dedicated series of head and neck LGFMS, including the first reported laryngeal case, as well as a review of reported head and neck cases. The surgical pathology archives of our three institutions were searched for cases of LGFMS arising within the head and neck, and four cases were identified. The H&E slides were reviewed, and immunohistochemistry were performed for pancytokeratin, p63, p40, EMA, S100 protein, ß-catenin, actin, CD34, and MUC4. The patients were 6, 43, 45, and 73 years old (mean 41.8 years) and included three males and one female. The tumors were located in the posterior cervical spine, facial skin, mandible, and larynx. The tumors were treated with surgical excision, and all four had histologic features typical for LGFMS including alternating myxoid and fibrous areas with prominent curvilinear vasculature. All tumors were MUC4 positive (100%), 2/4 (50%) were p63 positive, 1/4 (25%) showed focal EMA positivity; all 4 were negative for pancytokeratin, p40, S100 protein, ß-catenin, actin, and CD34. LGFMS is a low grade sarcoma that rarely develops in the head and neck. Due to its rarity, a pathologist may not consider LGFMS in the differential diagnosis of spindle cell neoplasms within the head and neck. Immunohistochemical staining is helpful, but stains should be selected carefully to avoid misdiagnosis.

CDX-2 Expression in Primary Lung Adenocarcinoma.

Adenocarcinoma with enteric differentiation is a rare subtype of lung adenocarcinoma that is recognized as a variant type of primary adenocarcinoma in the 2015 World Health Organization classification of lung tumors. Published immunohistochemistry studies show variable staining pattern for CDX-2 ranging from positivity in 71% of the cases to no staining. As little is known about CDX-2 expression in lung adenocarcinomas lacking histologic features of enteric differentiation, our aim was to determine the rate of CDX-2 positivity in non-enteric-type lung adenocarcinomas. We performed immunohistochemistry for CDX-2, CK7, CK20, TTF-1, napsin A, and p40 using 4-µm sections of a previously constructed tissue microarray containing 93 primary lung adenocarcinomas that lack morphologic evidence of enteric differentiation. The cohort included 22 well, 54 moderately, and 17 poorly differentiated tumors (55 female, 38 male; age range: 42 to 86, median: 64.5). All 93 tumors were strongly CK7 positive, whereas variable CK20 staining was seen in 4 tumors (1 strong, 1 moderate, and 2 focal). Both TTF-1 and napsin A were positive in 81 of 93 (87%) tumors with only 6 of 93 (6.5%) tumors negative for both the markers. Eleven tumors were CDX-2 positive (5 strong, 3 moderate, and 3 weak), all of which were also TTF-1 and napsin A positive and p40 negative. One CDX-2-positive tumor showed focal CK20 staining. Mutation studies for EGFR/K-ras/ALK were performed in four CDX-2-positive tumors and detected a K-ras mutation in one of them. CDX-2 positivity can be seen in a subset (12%) of lung adenocarcinoma. These tumors are CK7, TTF-1, and napsin A positive and p40 negative. Focal CK20 staining is only seen in rare cases. CDX-2 positivity should not be used as the only criteria to exclude lung origin.

Characterization of status epilepticus induced by two organophosphates in rats.

Organophosphates (OPs) inhibit the enzyme cholinesterase and cause accumulation of acetylcholine, and are known to cause seizures and status epilepticus (SE) in humans. The animal models of SE caused by organophosphate analogs of insecticides are not well characterized. SE caused by OPs paraoxon and diisopropyl fluorophosphate (DFP) in rats was characterized by electroencephalogram (EEG), behavioral observations and response to treatment with the benzodiazepine diazepam administered at various stages of SE. A method for SE induction using intrahippocampal infusion of paraoxon was also tested. Infusion of 200nmol paraoxon into the hippocampus caused electrographic seizures in 43/52 (82.7%) animals tested; and of these animals, 14/43 (30%) had self-sustaining seizures that lasted 4-18h after the end of paraoxon infusion. SE was also induced by peripheral subcutaneous injection of diisopropyl fluorophosphate (DFP, 1.25mg/kg) or paraoxon (1.00mg/kg) to rats pretreated with atropine (2mg/kg) and 2-pralidoxime (2-PAM, 50mg/kg) 30min prior to OP injection. SE occurred in 78% paraoxon-treated animals and in 79% of DFP-treated animals. Diazepam (10mg/kg) was administered 10min and 30min after the onset of continuous EEG seizures induced by paraoxon and it terminated SE in a majority of animals at both time points. DFP-induced SE was terminated in 60% animals when diazepam was administered 10min after the onset of continuous EEG seizure activity but diazepam did not terminate SE in any animal when it was administered 30min after the onset of continuous seizures. These studies demonstrate that both paraoxon and DFP can induce SE in rats but refractoriness to diazepam is a feature of DFP induced SE.