Evaluation of fine-needle core biopsy specimens for pancreatic ductal adenocarcinoma: Diagnostic utility and helpful histological features.

AIMS: With the advent of new biopsy devices, fine-needle core biopsy specimens can be obtained from pancreas masses. This study aimed to report the histological spectrum of intrapancreatic adenocarcinoma on fine-needle core biopsy and the accuracy of sampling. METHODS AND RESULTS: We identified 423 SharkCore™ fine-needle core biopsies taken from patients with a high clinical concern for pancreatic adenocarcinoma. For each, we recorded patient age and sex, percentage of diagnostic tissue in each sample and tumour site, size and histological findings. The cases came from 392 patients (193 men, 199 women; mean age 69 years). Median diagnostic tissue amount in the samples was 30%. Common histological findings included desmoplasia (36%), single atypical cells (44%), haphazard glandular growth pattern (68%), nuclear pleomorphism > 4:1 (39%), incomplete gland lumens (18%) and detached atypical epithelial strips (37%). Additional levels were ordered on 143 cases. Final clinical diagnoses associated with the 423 cases were adenocarcinoma (n = 343), pancreatitis (n = 22), intraductal neoplasm or other benign/low-grade process (n = 16) and unknown (n = 42, patients lost to follow-up). Of the adenocarcinoma cases, the diagnosis was established by the evaluated fine-needle core biopsy sample alone in 178, by fine-needle aspiration biopsy alone in 30, by both concurrently in 89 and by subsequent biopsy or resection in 37 cases. Among 68 cases called suspicious on fine-needle core biopsy, 78% ultimately represented adenocarcinoma. CONCLUSIONS: Fine-needle core biopsy allows for histological diagnosis of pancreatic adenocarcinoma, using known histological parameters. Common findings include single atypical cells, desmoplasia, haphazard gland growth and nuclear pleomorphism. Cases interpreted as suspicious often represent malignancy.

Comparative histologic features among liver biopsies with biliary-pattern injury and confirmed clinical diagnoses.

Biliary-pattern injury in the liver (eg, duct injury, ductular reaction, cholestasis) can occur in several conditions, including primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC), large duct obstruction (LDO), and drug-induced liver injury (DILI). While the histologic changes in these conditions have been individually well described, distinguishing among them remains often challenging, particularly when biopsy samples are limited in size, robust clinical information is unavailable, and/or the pathologist does not feel confident in evaluating liver disease. This study evaluated histologic features that could aid the diagnosis of biliary-pattern injury on biopsy. We reviewed 121 liver biopsies from clinically confirmed cases of PBC, PSC, chronic LDO, or DILI for multiple clinical and histologic parameters. The rates of these histologic findings were then compared among different entities. Onion-skin fibrosis was seen in 14% of PSC in comparison to 0%, 5%, and 0% of PBC, DILI, and chronic LDO (P = 0.031). Florid duct lesions were identified in 21% of PBC compared to 2% of PSC and 0% of DILI and LDO (P = 0.0065). Similarly, 42% of PBC showed lobular granulomas, compared to 7% of PSC, 11% of DILI, and 33% of chronic LDO (P = 0.0001). Cholestasis was more commonly seen in DILI (42%) and chronic LDO (83%) than in PBC (4%) and PSC (16%) (P < 0.0001). Lobular chronic inflammation was found in a significantly higher percentage of PBC and LDO than of PSC and DILI (P = 0.0009). There were significantly fewer cases of PBC showing neutrophils in ductular reaction than PSC, DILI, and LDO (P = 0.0063). Histologic findings that can help suggest a diagnosis in liver biopsies with biliary-pattern injury include florid duct lesions, lobular granulomas, lack of neutrophils in ductular reaction, and lobular chronic inflammation in PBC; onion-skin fibrosis in PSC; cholestasis and feathery degeneration in DILI; and lobular granulomas, lobular chronic inflammation, cholestasis, and feathery degeneration in chronic LDO. These findings are likely most helpful when complicating factors interfere with biopsy interpretation.

Risk of malignancy associated with diagnostic categories of the proposed World Health Organization International System for Reporting Pancreaticobiliary Cytopathology.

BACKGROUND: The World Health Organization (WHO) has proposed an updated international classification system for reporting pancreaticobiliary cytology. Substantial changes to the prior Papanicolaou Society of Cytopathology (PSC) system have been recommended. Chiefly, the "neoplastic: benign" and "neoplastic: other" categories have been replaced by 2 new categories-"pancreatic neoplasia-low-grade" (PaN-Low) and "pancreatic neoplasia-high-grade" (PaN-High)-stratifying neoplastic mucinous cysts by cytological atypia. Low-grade malignancies are placed in the "malignant" category and benign serous cystadenoma in the "benign/negative" category. Risk of malignancy (ROM) associated with the diagnostic categories of the WHO system has yet to be defined. METHODS: All patients who underwent endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) for a pancreatic lesion at a single institution from January 2016 to December 2016, prospectively classified using the PSC system, were reclassified using the WHO system. Absolute ROM was determined by histologic outcome and/or clinical follow-up of at least 6 months. RESULTS: A total of 334 EUS-FNA samples from 322 patients were reviewed and reclassified. Absolute ROM for the WHO system was 7.7% for "insufficient/inadequate/nondiagnostic" category, 1.0% for "benign/negative for malignancy," 28.0% for "atypical," 4.8% for "PaN-Low," 60.0% for "PaN-High," 100% for "suspicious for malignancy," and 100% for "malignant;" the absolute ROM for the same cohort using the PSC system was 7.7% for "nondiagnostic" category, 1.0% for "negative (for malignancy)," 28.0% for "atypical," 0.0% for "neoplastic: benign," 30.3% for "neoplastic: other," 100% for "suspicious (for malignancy)," and 100% for "positive or malignant." CONCLUSIONS: The WHO international system achieves improved stratification by associated ROM compared to the PSC system.

Immunohistochemistry as predictive and prognostic markers for gastrointestinal malignancies.

Biomarkers play a key role in the comprehensive pathologic evaluation of gastrointestinal malignancies. These biomarkers can be predictive, indicating whether a tumor is likely to respond to a particular therapy, or prognostic, providing information about the likely course and outcome of a disease. This review article will discuss available immunohistochemical stains for assessing these markers, including staining rationale, scoring criteria, associated systemic therapies, and pictorial examples. PD-L1, HER2, and mismatch repair status can be evaluated via immunohistochemistry for esophageal, gastric, and colorectal carcinomas. Biomarkers currently play a more limited role in evaluation of pancreatic and small bowel malignancies. Immunohistochemistry can also be used to evaluate biomarker status in gastrointestinal stromal tumors, gastrointestinal malignancies with NTRK gene fusions, and undifferentiated carcinomas with switch-sucrose non-fermentable complex abnormalities.

Next-generation sequencing in the evaluation of biliary strictures in patients with primary sclerosing cholangitis.

BACKGROUND: Primary sclerosing cholangitis (PSC) is a well-described risk factor for the development of cholangiocarcinoma (CCA). Early detection of CCA in these patients is of great importance because it expands options for therapeutic interventions, including liver transplantation. Current diagnostic tests for the evaluation of biliary strictures are limited to biliary brushing (BB) cytology and fluorescence in situ hybridization (FISH). Next-generation sequencing (NGS) has become an important diagnostic tool in oncology and may be a useful tool for diagnosing CCA on BBs. It is not clear how NGS performs when it is added to BB cytology and FISH in patients with PSC. METHODS: This study reports the authors' experience with NGS performed as a prospective cotest with cytology and FISH on BBs obtained from 60 patients with PSC followed at Massachusetts General Hospital. A duct with malignancy was defined as a high-risk (HR) stricture with either high-grade dysplasia or CCA. RESULTS: NGS was better than FISH and cytology in detecting HR strictures, which showed multiple genetic mutations in all cases. NGS provided specific mutational information, and NGS results were reproducible in longitudinal samples. CONCLUSIONS: Adding NGS to BB cytology and FISH in the evaluation of biliary strictures for patients with PSC may provide additional information that could help to inform clinical management.

ACGME Milestones 2.0: why and what's new for cytopathology?

BACKGROUND: Primary stakeholders in the Accreditation Council for Graduate Medical Education (ACGME) Milestones Project are: ACGME, Residency Programs, Residents, Fellowship Programs, Fellows, and Certification Boards. The intent of the Milestones is to describe the educational and professional developmental trajectory of a trainee from the first stages of their postgraduate education through the completion of their clinical training. The Milestones 2.0 project includes changes made based on experience with Milestones 1.0. METHODS: The ACGME solicited volunteers to participate in the development of subspecialty Milestones 2.0. The workgroup was charged with reviewing/making any additions to the four "Harmonized Milestones", developing subspecialty specific milestones for the Patient Care and Medical Knowledge competencies, and creating a supplemental guide. The Milestones were finalized following review of input from an open comment period. RESULTS: The Cytopathology Milestones 2.0 will go into effect July 2021. They include additional subcompetencies in the 4 harmonized competency areas and cytopathology-specific edits to the patient care and medical knowledge subcompetencies. Although the number of subcompetencies has increased from 18 to 21, within each subcompetency, the number of milestone trajectories has decreased. Additionally, within each subcompetency, the wording has been streamlined. A supplemental guide was created and Milestones 1.0 were compared to 2.0; however, curriculum mapping has been left to programs to develop. CONCLUSIONS: The ultimate goal of the Cytopathology Milestones 2.0 is to provide better real-time documentation of the progress of cytopathology fellows. The expected outcome is to produce highly competent cytopathologists, improving the care they provide, regardless of the program at which they trained.

Critical values in cytology.

The timely reporting of critical values, or values that may be life-threatening if immediate action is not taken, is essential to patient care and safety. Although some guidelines exist for critical diagnoses in cytology, not all laboratories have a specific list of diagnoses that should be considered critical, and the very existence of cytology "critical values" has been called into question. Here we propose a pragmatic system for determining cytology critical values and report our laboratory's critical value list, formulated based on a review of the medical literature regarding clinical urgency and other institutions' cytology critical value lists.

Targeted therapy for upper gastrointestinal tract cancer: current and future prospects.

Gastric and oesophageal carcinoma remain major causes of worldwide mortality and morbidity. Despite incredible progress in understanding tumour biology, few targeted treatment options have proved effective in prolonging survival, and adjuvant therapy is largely interchangeable in these carcinomas. Through large-scale sequencing by the Cancer Genome Atlas and the Asian Cancer Research Group, numerous potential molecular targets have been discovered. Of the approved targeted therapies for gastric and oesophageal cancer, pathologists play a role in patient selection for the majority of them. Trastuzumab has been approved as a first-line therapy in conjunction with standard treatment in adenocarcinomas with either 3+ HER2/neu expression by immunohistochemistry or ERBB2 amplification by FISH. PD-L1 immunohistochemistry showing a combined positive score of 1 or greater qualifies patients for third-line pembrolizumab therapy, and identification of microsatellite instability-high carcinomas may qualify patients for second-line pembrolizumab. Ramucirumab, targeting VEGFR2, has also been approved for second-line therapy in gastric carcinoma. Non-surgical therapy for gastrointestinal stromal tumours relies mainly upon tyrosine kinase inhibitors, while new targeted therapy options for neuroendocrine neoplasms have recently emerged. Potential future options for targeted therapy in all these malignancies are being investigated in clinical trials, as this review will discuss.

Cytomorphologic characteristics of next-generation sequencing-positive bile duct brushing specimens.

INTRODUCTION: Cytology of bile duct brushings (BDBs) is a specific, but insensitive, test for malignancy. Next-generation sequencing (NGS) of BDBs has recently been shown to improve sensitivity. We analyzed the cytologic features of NGS-positive (NGS+) and NGS-negative (NGS-) BDBs and correlated the morphology with the presence of mutations. MATERIALS AND METHODS: A total of 96 BDBs were analyzed for 29 cytologic features by 2 pathologists who were unaware of the original diagnosis and NGS results. Clinicopathologic follow-up was used to determine the patient outcomes (ie, benign, low-grade neoplasm, malignant [carcinoma/high-grade dysplasia]). RESULTS: We analyzed 74 NGS+ BDBs from 66 patients and 22 NGS- BDBs from 22 patients. During follow-up, 58 of 66 NGS+ patients (88%) had malignancy compared with 0% of NGS- patients (P < 0.001). Fewer than 50% of the malignant cases had been interpreted as malignant on cytology; however, 100% had demonstrated mutations using NGS. Within the NGS+ cases, 53% showed late mutations (TP53, SMAD4, and CDKN2A) supportive of a high-risk stricture. Significant morphologic differences were seen in the background appearance, presence of single cells, architectural disarray, nucleomegaly, anisonucleosis, irregular nuclear borders, increased nuclear/cytoplasmic ratio, nuclear hyperchromasia, nucleoli, abnormal groups, clusters, and/or single cells, and overall impression. Naked nuclei, nucleomegaly, anisonucleosis, and coarse chromatin were more common in BDBs with late mutations than in those with KRAS/GNAS (Kirsten rat sarcoma viral oncogene homolog/guanine nucleotide binding protein, α-stimulating complex locus) mutations only. Cytology had a sensitivity of 16% and a specificity of 100% for malignancy. In contrast, NGS had a sensitivity of 100% and a specificity of 73%. Late mutations were 100% specific for malignancy compared with mutations in KRAS/GNAS only, of which 69% were malignant. CONCLUSIONS: We found significant overlap in the cytomorphologic features between neoplastic and non-neoplastic BDBs, and more than one half of cancer cases had been interpreted as "nonmalignant" on cytology. NGS showing late mutations was 100% specific for malignancy. Adding genetic testing to BDB cytology would be a valuable ancillary test for the detection of malignancy, and reflex testing should be considered.

Risk of malignancy in pancreatic cysts with cytology of high-grade epithelial atypia.

BACKGROUND: The risk of malignancy is weighed against the attendant risks of surgery in the clinical management of pancreatic cysts. The latter are a group of histologically diverse and prognostically variable entities, and the risk of malignancy therein is primarily based on imaging characteristics-with or without high-grade atypia. Cytologic criteria for high-grade atypia in intraductal papillary mucinous neoplasms have recently been defined, and its recognition in all pancreatic cysts may help to guide management. METHODS: All patients who underwent endoscopic ultrasound-guided fine-needle aspiration for a pancreatic cyst at Massachusetts General Hospital from June 2015 to October 2016 were prospectively evaluated. Clinical data, radiographic impressions, biochemical analyses, and cytologic diagnoses of 118 pancreatic cyst fine-needle aspiration biopsy specimens from 106 patients were reviewed. Clinical and radiologic data were used as follow-up for 86 patients, and histology was obtained in 20 cases. Cysts were classified by imaging as high-risk, worrisome, or low-risk as defined by the 2012 Fukuoka consensus guidelines. Cytology was categorized as low-grade or high-grade. Malignant histology included mucinous cysts with high-grade dysplasia, invasive adenocarcinomas, and neuroendocrine tumors. The risk of malignancy (ROM) was determined by histological outcome. RESULTS: The presence of high-grade cytology (P < .01) was the only statistically significant predictor of malignancy and was 89% sensitive and 98% specific for malignancy. The positive predictive value (ie, ROM) of high-grade atypia on cytology was 80%. CONCLUSIONS: High-grade atypia is both sensitive and specific for identifying high-risk pancreatic cysts and is associated with a high risk of malignancy, and thus resection is warranted.

Using Machine Learning-Based Multianalyte Delta Checks to Detect Wrong Blood in Tube Errors.

OBJECTIVES: An unfortunate reality of laboratory medicine is that blood specimens collected from one patient occasionally get mislabeled with identifiers from a different patient, resulting in so-called "wrong blood in tube" (WBIT) errors and potential patient harm. Here, we sought to develop a machine learning-based, multianalyte delta check algorithm to detect WBIT errors and mitigate patient harm. METHODS: We simulated WBIT errors within sets of routine inpatient chemistry test results to develop, train, and evaluate five machine learning-based WBIT detection algorithms. RESULTS: The best-performing WBIT detection algorithm we developed was based on a support vector machine and incorporated changes in test results between consecutive collections across 11 analytes. This algorithm achieved an area under the curve of 0.97 and considerably outperformed traditional single-analyte delta checks. CONCLUSIONS: Machine learning-based multianalyte delta checks may offer a practical strategy to identify WBIT errors prior to test reporting and improve patient safety.

Quality Control Practices for Chemistry and Immunochemistry in a Cohort of 21 Large Academic Medical Centers.

OBJECTIVES: In the United States, minimum standards for quality control (QC) are specified in federal law under the Clinical Laboratory Improvement Amendment and its revisions. Beyond meeting this required standard, laboratories have flexibility to determine their overall QC program. METHODS: We surveyed chemistry and immunochemistry QC procedures at 21 clinical laboratories within leading academic medical centers to assess if standardized QC practices exist for chemistry and immunochemistry testing. RESULTS: We observed significant variation and unexpected similarities in practice across laboratories, including QC frequency, cutoffs, number of levels analyzed, and other features. CONCLUSIONS: This variation in practice indicates an opportunity exists to establish an evidence-based approach to QC that can be generalized across institutions.

PD-L1 and IDO1 Are Expressed in Poorly Differentiated Thyroid Carcinoma.

Poorly differentiated thyroid carcinoma (PDTC) is an aggressive form of thyroid cancer that currently has limited effective treatment options. Immune checkpoint inhibitors (ICIs) have shown to be an effective treatment for a variety of carcinomas. In this study, we explore whether immune checkpoint pathways, such as programmed cell death ligand 1 (PD-L1) and indoleamine 2,3-dioxygenase 1 (IDO1), are activated in a cohort of patients with PDTC to determine whether ICIs may be an effective therapy for these patients. PDTC from 28 patients were stained for IDO1, PD-L1, and CD8 using immunohistochemistry. Staining was scored using an H-score, and PD-L1 and IDO1 expression was correlated with clinicopathologic characteristics. Positivity for PD-L1 and IDO1 was set at an H-score cutoff of five. Twenty-five percent (n = 7/28) of the PDTC were positive for PD-L1 expression. Twenty-nine percent (n = 2/7) of the PD-L1 positive PDTCs also co-expressed IDO1. The expression of PD-L1 in PDTC was significantly associated with tumor size and multifocality, with a non-significant trend towards associations with older age, extrathyroidal extension, presence of metastasis, higher stage, increased number of CD8+ T cells, and decreased disease-free and overall survival. PD-L1 expression occurs in a subset of PDTC, and is associated with a subset of clinical features of aggressive thyroid disease. Given the limited effective treatments for this patient population, consideration for ICIs as monotherapy or in combination with an IDO1 inhibitor should be explored as a novel treatment modality for patients with PDTC.

Rivaroxaban Causes Missed Diagnosis of Protein S Deficiency but Not of Activated Protein C Resistance (Factor V Leiden).

CONTEXT: - Rivaroxaban causes a false increase in activated protein C resistance (APCR) ratios and protein S activity. OBJECTIVE: - To investigate whether this increase masks a diagnosis of factor V Leiden (FVL) or protein S deficiency in a "real-world" population of patients undergoing rivaroxaban treatment and hypercoagulation testing. DESIGN: - During a 2.5-year period, we compared 4 groups of patients (n = 60): FVL heterozygous (FVL-HET)/taking rivaroxaban, wild-type/taking rivaroxaban, FVL-HET/no rivaroxaban, and normal APCR/no rivaroxaban. Patients taking rivaroxaban were tested for protein S functional activity and free antigen (n = 32). RESULTS: - The FVL-HET patients taking rivaroxaban had lower APCR ratios than wild-type patients ( P < .001). For FVL-HET patients taking rivaroxaban, mean APCR was 1.75 ± 0.12, versus 1.64 ± 0.3 in FVL-HET patients not taking rivaroxaban ( P = .005). Activated protein C resistance in FVL-HET patients fell more than 3 SDs below the cutoff of 2.2 at which the laboratory reflexes FVL DNA testing. No cases of FVL were missed despite rivaroxaban. In contrast, rivaroxaban falsely elevated functional protein S activity, regardless of the presence or absence of FVL ( P < .001). A total of 4 of 32 patients (12.5%) had low free protein S antigen (range, 58%-67%), whereas their functional protein S activity appeared normal (range 75%-130%). Rivaroxaban would have caused a missed diagnosis of all cases of protein S deficiency during the study if testing relied on the protein S activity assay alone. CONCLUSIONS: - Despite rivaroxaban treatment, APCR testing can distinguish FVL-HET from normal patients, rendering indiscriminate FVL DNA testing of all patients on rivaroxaban unnecessary. Free protein S should be tested in patients taking rivaroxaban to exclude hereditary protein S deficiency.

Ring chromosome in myeloid neoplasms is associated with complex karyotype and disease progression.

Ring chromosome (RC) is a poorly understood genetic anomaly seen in myeloid neoplasms. This study aims to shed light on the clinical significance of this finding. We identified 96 cases of myeloid neoplasms with RC from 3 academic hospitals. Clinicopathologic features and overall (OS) and leukemia-free survival were reviewed and compared to cases of myeloid neoplasms lacking RC. We identified 59 acute myeloid leukemias (AML-RC) and 37 myelodysplastic syndromes (MDS-RC) with RC identified on routine karyotyping. Seventy-five percent of AML-RC and 97% of MDS-RC had complex (>3 independent cytogenetic abnormalities) karyotypes. The median OS of AML-RC with complex karyotype was significantly shorter than AML-RC patients with a non-complex (≤3 independent cytogenetic abnormalities) karyotype (P=.001), but similar to AML patients with complex karyotype lacking RC (P=not significant). Compared to complex-karyotype MDS lacking RC, MDS-RC patients had shorter leukemia-free survival (P=.016) and a trend for shorter OS (P=.10). RCs were sometimes lost after therapy or appeared during disease relapse, suggesting that they may be associated with genetic instability.

Tumor engraftment in patient-derived xenografts of pancreatic ductal adenocarcinoma is associated with adverse clinicopathological features and poor survival.

Patient-derived xenograft (PDX) tumors are powerful tools to study cancer biology. However, the ability of PDX tumors to model the biological and histological diversity of pancreatic ductal adenocarcinoma (PDAC) is not well known. In this study, we subcutaneously implanted 133 primary and metastatic PDAC tumors into immunodeficient mice. Fifty-seven tumors were successfully engrafted and even after extensive passaging, the histology of poorly-, moderately-, and well-differentiated tumors was maintained in the PDX models. Moreover, the fibroblast and collagen contents in the stroma of patient tumors were recapitulated in the corresponding PDX models. Analysis of the clinicopathological features of patients revealed xenograft tumor engraftment was associated with lymphovascular invasion (P = 0.001) and worse recurrence-free (median, 7 vs. 16 months, log-rank P = 0.047) and overall survival (median, 13 vs. 21 months, log-rank P = 0.038). Among successful engraftments, median time of growth required for reimplantation into new mice was 151 days. Reflective of the inherent biological diversity between PDX tumors with rapid (<151 days) and slow growth, differences in their growth were maintained during extensive passaging. Rapid growth was additionally associated with lymph node metastasis (P = 0.022). The association of lymphovascular invasion and lymph node metastasis with PDX formation and rapid growth may reflect an underlying biological mechanism that allows these tumors to adapt and grow in a new environment. While the ability of PDX tumors to mimic the cellular and non-cellular features of the parental tumor stroma provides a valuable model to study the interaction of PDAC cells with the tumor microenvironment, the association of successful engraftment with adverse clinicopathological features suggests PDX models over represent more aggressive forms of this disease.

Cytologic characteristics of circulating epithelioid cells in pancreatic disease.

BACKGROUND: Circulating epithelioid cells (CECs), also known as circulating tumor, circulating cancer, circulating epithelial, or circulating nonhematologic cells, are a prognostic factor in various malignancies that can be isolated via various protocols. In the current study, the authors analyzed the cytomorphologic characteristics of CECs isolated by size in a cohort of patients with benign and malignant pancreatic diseases to determine whether cytomorphological features could predict CEC origin. METHODS: Blood samples were collected from 9 healthy controls and 171 patients with pancreatic disease who were presenting for surgical evaluation before treatment. Blood was processed with the ScreenCell size-based filtration device. Evaluable CECs were analyzed in a blinded fashion for cytomorphologic characteristics, including cellularity; nucleoli; nuclear size, irregularity, variability, and hyperchromasia; and nuclear-to-cytoplasmic ratio. Statistical differences between variables were analyzed via the Fisher exact test. RESULTS: No CECs were identified among the 9 normal healthy controls. Of the 115 patients with CECs (positive or suspicious for), 25 had nonmalignant disease and 90 had malignancy. There were no significant differences in any of the cytologic criteria noted between groups divided by benign versus malignant, neoplastic versus nonneoplastic, or pancreatic ductal adenocarcinoma versus neuroendocrine tumor. CONCLUSIONS: CECs were observed in patients with malignant and nonmalignant pancreatic disease, but not in healthy controls. There were no morphologic differences observed between cells from different pancreatic diseases, suggesting that numerous conditions may be associated with CECs in the circulation and that care must be taken not to overinterpret cells identified by cytomorphology as indicative of circulating tumor cells of pancreatic cancer. Additional studies are required to determine the origin and clinical significance of these cells. Cancer Cytopathol 2017;125:332-340. © 2017 American Cancer Society.

Next-generation sequencing adds value to the preoperative diagnosis of pancreatic cysts.

BACKGROUND: The diagnosis of a pancreatic cyst as mucinous or high-risk dictates the need for follow-up or surgery. Molecular analysis of aspirated pancreatic cyst fluid (PCF) can provide valuable information not obtained by carcinoembryonic antigen (CEA) analysis or cytology. METHODS: All patients who underwent molecular analysis of PCF between March 2013 and June 2015 were reviewed, including pathology, imaging, and follow-up. Molecular testing was performed using a patented, anchored multiplex polymerase chain reaction next-generation sequencing (NGS) platform, which sequenced numerous hotspots in 39 genes linked with malignancy. Performance of NGS and cytology was calculated using final outcome, as determined by clinicopathologic follow-up. RESULTS: The study cohort included 113 PCFs from 105 patients. In total, 119 variants were detected in 67 PCFs (59%). Variants were more common in intraductal papillary mucinous neoplasms (IPMNs)/cancer than in nonmucinous cysts (P < .005). The inclusion of v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS)/guanine nucleotide-binding protein (GNAS) variants improved the classification of IPMNs as mucinous from 50% by microscopy to 100%. Seventy-five percent of cancers had high-grade atypia versus 0% of IPMNs and nonmucinous cysts (P < .002). Variants in tumor protein 53 (TP53), SMAD family member 4 (SMAD4), cyclin-dependent kinase inhibitor 2A (CDKN2A), and notch1 (NOTCH1) were detected only in malignant cysts. Cytology was similarly specific (100%) for detecting malignant cysts but was more sensitive than the identification of late mutations by NGS (75% vs 46%). CONCLUSIONS: The detection of KRAS/GNAS variants improves the identification of mucinous neoplasms. Variants in TP53, SMAD4, CDKN2A, and NOTCH1 support the diagnosis of a high-risk cyst requiring surgery or additional sampling. Although molecular analysis is not a replacement for cytopathology, it does provide valuable information for accurate preoperative diagnosis, helping to classify mucinous neoplasms and high-risk cysts that require surgical resection. Cancer Cytopathol 2017;125:41-47. © 2016 American Cancer Society.

PD-L1 expression in colorectal cancer is associated with microsatellite instability, BRAF mutation, medullary morphology and cytotoxic tumor-infiltrating lymphocytes.

Programmed cell death 1 (PD-1) and its ligand (PD-L1) are key suppressors of the cytotoxic immune response. PD-L1 expression on tumor cells may be induced by the immune microenvironment, resulting in immune escape (adaptive immune resistance), and an adverse prognosis in many malignancies. In colorectal carcinoma the response to PD-1/PD-L1 inhibition is correlated with microsatellite instability. However, little is known about the clinicopathologic, molecular, and prognostic characteristics of colorectal carcinoma with PD-L1 expression. We performed immunohistochemistry for PD-L1 on 181 cases of colorectal carcinoma with known microsatellite instability and mutational status, and correlated PD-L1 expression with clinicopathologic features including tumor-infiltrating lymphocyte burden/immunophenotype, tumor mutational profile, and disease-specific survival. PD-L1 was expressed in tumors from 16 patients (9%) who were more often older (P=0.006) and female (P=0.035), with tumors exhibiting a larger size (P=0.013), but lower stage (P<0.001). PD-L1 expression was associated with increased CD8 and TBET-positive tumor-infiltrating lymphocytes, medullary phenotype, poor differentiation, microsatellite instability, BRAF mutation (P<0.001 for each), and a lower frequency of KRAS mutation (P=0.012). On multivariate analysis, PD-L1 expression was associated with medullary morphology and frequent CD8-positive tumor-infiltrating lymphocytes, suggesting adaptive immune resistance. PD-L1 positivity was not predictive of survival in the entire cohort, but it was associated with a lower disease-specific survival within the microsatellite-instability high cohort. PD-L1 expression in colorectal carcinoma is associated with clinicopathologic and molecular features of the serrated pathway of colorectal carcinogenesis, and is associated with a worse outcome within microsatellite-unstable tumors. These findings support the role of PD-L1 expression in providing normally immunogenic colorectal carcinoma a means of immune evasion and a more aggressive biology, provide a potential mechanistic explanation for the favorable response of microsatellite-unstable colorectal carcinoma to PD-1/PD-L1 pathway blockade, and suggest potential predictive and prognostic roles of PD-L1 immunohistochemistry in colorectal carcinoma.