Collection and Handling of Thoracic Small Biopsy and Cytology Specimens for Ancillary Studies: Guideline From the College of American Pathologists in Collaboration With the American College of Chest Physicians, Association for Molecular Pathology, American Society of Cytopathology, American Thoracic Society, Pulmonary Pathology Society, Papanicolaou Society of Cytopathology, Society of Interventional Radiology, and Society of Thoracic Radiology.

CONTEXT.­: The need for appropriate specimen use for ancillary testing has become more commonplace in the practice of pathology. This, coupled with improvements in technology, often provides less invasive methods of testing, but presents new challenges to appropriate specimen collection and handling of these small specimens, including thoracic small biopsy and cytology samples. OBJECTIVE.­: To develop a clinical practice guideline including recommendations on how to obtain, handle, and process thoracic small biopsy and cytology tissue specimens for diagnostic testing and ancillary studies. METHODS.­: The College of American Pathologists convened an expert panel to perform a systematic review of the literature and develop recommendations. Core needle biopsy, touch preparation, fine-needle aspiration, and effusion specimens with thoracic diseases including malignancy, granulomatous process/sarcoidosis, and infection (eg, tuberculosis) were deemed within scope. Ancillary studies included immunohistochemistry and immunocytochemistry, fluorescence in situ hybridization, mutational analysis, flow cytometry, cytogenetics, and microbiologic studies routinely performed in the clinical pathology laboratory. The use of rapid on-site evaluation was also covered. RESULTS.­: Sixteen guideline statements were developed to assist clinicians and pathologists in collecting and processing thoracic small biopsy and cytology tissue samples. CONCLUSIONS.­: Based on the systematic review and expert panel consensus, thoracic small specimens can be handled and processed to perform downstream testing (eg, molecular markers, immunohistochemical biomarkers), core needle and fine-needle techniques can provide appropriate cytologic and histologic specimens for ancillary studies, and rapid on-site cytologic evaluation remains helpful in appropriate triage, handling, and processing of specimens.

Small cell and non small cell lung cancer form metastasis on cellular 4D lung model.

BACKGROUND: Metastasis is the main cause of death for lung cancer patients. The ex vivo 4D acellular lung model has been shown to mimic this metastatic process. However, the main concern is the model's lack of cellular components of the tumor's microenvironment. In this study, we aim to determine if the intact lung microenvironment will still allow lung cancer metastasis to form. METHODS: We harvested a heart-lung block from a rat and placed it in a bioreactor after cannulating the pulmonary artery, trachea and tying the right main bronchus for 10-15 days without any tumor cells as a control group or with NSCLC (A549, H1299 or H460), SCLC (H69, H446 or SHP77) or breast cancer cell lines (MCF7 or MDAMB231) through the trachea. We performed lobectomy, H&E staining and IHC for human mitochondria to determine the primary tumor's growth and formation of metastatic lesions. In addition, we isolated circulating tumor cells (CTC) from the model seeded with GFP tagged cells. RESULTS: In the control group, no gross tumor nodules were found, H&E staining showed hyperplastic cells and IHC showed no staining for human mitochondria. All of the models seeded with cancer cell lines formed gross primary tumor nodules that had microscopic characteristics of human cancer cells on H&E staining with IHC showing staining for human mitochondria. CTC were isolated for those cells labeled with GFP and they were viable in culture. Finally, all cell lines formed metastatic lesions with cells stained for human mitochondria. CONCLUSION: The cellular ex vivo 4D model shows that human cancer cells can form a primary tumor, CTC and metastatic lesions in an intact cellular environment. This study suggests that the natural matrix scaffold is the only necessary component to drive metastatic progression and that cellular components play a role in modulating tumor progression.

Flat epithelial atypia in directional vacuum-assisted biopsy of breast microcalcifications: surgical excision may not be necessary.

The aim of this study was to analyze the clinicopathological features of patients with flat epithelial atypia, diagnosed in directional vacuum-assisted biopsy targeting microcalcifications, to identify upgrade rate to in situ ductal or invasive breast carcinoma, and determine factors predicting carcinoma in the subsequent excision. We retrospectively evaluated the histological, clinical, and mammographic features of 69 cases from 65 women, with directional vacuum-assisted biopsy-diagnosed flat epithelial atypia with or without atypical ductal hyperplasia or atypical lobular hyperplasia, which underwent subsequent surgical excision. The extent and percentage of microcalcifications sampled by directional vacuum-assisted biopsy were evaluated by mammography. All biopsy and surgical excision slides were reviewed. The age of the women ranged from 40 to 85 years (mean 57 years). All patients presented with mammographically detected microcalcifications only, except in one case that had associated architectural distortion. Extent of calcifications ranged from <1 cm (n = 47), 1-3 cm (n = 15) to > 3 cm (n = 6), and no measurement (n = 1). A mean of 11 cores (range 6-25) was obtained from each lesion. Post-biopsy mammogram revealed >90% removal of calcifications in 81% of cases. Pure flat epithelial atypia represented nearly two-thirds of directional vacuum-assisted biopsy specimens (n = 43, 62%), while flat epithelial atypia coexisted with atypical ductal hyperplasia (18 cases, 26%), or atypical lobular hyperplasia (8 cases, 12%). Upon excision, none of the cases were upgraded to in situ ductal or invasive breast cancer. In one case, however, an incidental, tubular carcinoma (4 mm) was found away from biopsy site. Excluding this case, the upgrade rate was 0%. Our study adds to the growing evidence that diagnosis of flat epithelial atypia on directional vacuum-assisted biopsy for microcalcifications as the only imaging finding is not associated with a significant upgrade to carcinoma on excision, and therefore, excision may not be necessary. Additionally, excision may not be necessary for flat epithelial atypia with atypical ductal hyperplasia limited to ≤2 terminal duct-lobular units, if at least 90% of calcifications have been removed on biopsy.

Correlation Between Programmed Death Receptor-1 Expression in Tumor-Infiltrating Lymphocytes and Programmed Death Ligand-1 Expression in Non-Small Cell Lung Carcinoma.

CONTEXT.­: The interaction between programmed death ligand-1 (PD-L1) and programmed death receptor-1 (PD-1) on activated T cells sends an inhibitory signal that dampens the immune response. Tumors can express PD-L1 and evade the immune system. In advanced non-small cell lung carcinoma, expression of PD-1 in tumor-infiltrating lymphocytes (TILs) correlates with PD-L1 expression in tumor cells (TCs). However, this relationship has not been thoroughly explored in early disease. OBJECTIVE.­: To investigate the correlation of PD-1 and PD-L1 in non-small cell lung carcinoma tumor samples, with emphasis on stage I disease. DESIGN.­: Whole tissue sections from non-small cell lung carcinoma tumors were retrospectively evaluated by immunohistochemistry for PD-1 and PD-L1 expression. The scoring was based on the percentage of cells positive for PD-1 in TILs and PD-L1 in TCs and tumor-infiltrating immune cells (ICs). RESULTS.­: Expression of PD-1 in TILs was observed in 147 of 161 non-small cell lung carcinoma cases (91%). The majority of cases negative for PD-1 also lacked PD-L1 in TCs. The 68 cases with highest PD-1 expression in TILs included 33 (49%) with expression of PD-L1 in TCs and ICs. Strong correlations were observed in patients with elevated PD-1 expression in TILs and PD-L1 in TCs ( P = .01) and ICs ( P = .003). Expression of PD-1 also correlated with increased PD-L1 in TCs and ICs when the 2 were grouped together ( P < .001). Finally, stage I patients with negative PD-1 and PD-L1 expression showed trends toward increased disease-specific survival. CONCLUSIONS.­: Expression of PD-1 in TILs correlates with PD-L1 expression in both TCs and ICs. Furthermore, negative expression of PD-1 and PD-L1 suggest trends toward disease-specific survival, even in early disease stages.

Programmed Death Ligand-1 (PD-L1) Expression in Either Tumor Cells or Tumor-Infiltrating Immune Cells Correlates With Solid and High-Grade Lung Adenocarcinomas.

CONTEXT: - Programmed death ligand-1 (PD-L1) expression in non-small cell lung carcinoma (NSCLC) is heterogeneous and known to be underestimated on small biopsies. Correlation of PD-L1 expression with clinicopathologic features may provide additional useful information. To our knowledge, the clinicopathologic features of NSCLC have not been reported for subsets defined by PD-L1 expression in either tumor cells or tumor-infiltrating immune cells. OBJECTIVE: - To investigate the clinicopathologic characteristics of NSCLC subsets defined by PD-L1 expression in either tumor cells or tumor-infiltrating immune cells. DESIGN: - PD-L1 immunohistochemistry with the SP142 clone was performed on whole-tissue sections and given semiquantitative scores (0/1/2/3) according to percent of PD-L1+ tumor cells (TCs) and percent tumor area with PD-L1+ tumor-infiltrating immune cells (ICs). RESULTS: - Adenocarcinoma cases that were scored either TC 1/2/3 or IC 1/2/3 included most (22 of 34; 65%) high-histologic grade cases and most (25 of 36; 69%) solid subtype cases. Compared with the adenocarcinoma TC 0 and IC 0 subset, the TC 1/2/3 or IC 1/2/3 subset correlated with higher histologic grade (P = .005, χ2 test for trend) and solid subtype (P < .001, Fisher exact test). Compared with the adenocarcinoma TC 0/1 or IC 0/1 subset, the TC 2/3 or IC 2/3 subset correlated with higher histologic grade (P = .002, χ2 test for trend), solid subtype (P < .001, Fisher exact test), and higher smoking pack-years (P = .01, Mann-Whitney test). CONCLUSIONS: - Lung adenocarcinoma subsets defined by PD-L1 expression in either tumor cells or tumor-infiltrating immune cells correlated with high histologic grade, solid subtype, and high smoking pack-years.

Biomarkers for Selection of Therapy for Adenocarcinoma of the Lung.

To suggest that the discovery of targetable driver mutations in many patients with advanced adenocarcinoma of the lung has completely transformed the work-up and therapeutic options for this disease would not be hyperbole. Although not curative, small-molecule tyrosine kinase inhibitors directed at oncogene-addicted tumors have led to significantly improved response rates compared with cytotoxic chemotherapy, with often manageable toxicities and better tolerance. However, the absence of reliable clinical predictors has made molecular testing essential to ensure that patients receive the proper medical management. We outline the many recent advances with regard to diagnosis and treatment of oncogene-addicted advanced nonsquamous non-small-cell lung cancer.

Adverse tissue reactions after total hip arthroplasty.

Hip arthroplasty is commonly performed on patients with debilitating hip disease to relieve symptoms and improve quality of life. Generally, long-term success rates are excellent. However, a subset of patients requires revision due to prosthesis failure. A wide array of microscopic findings can be seen in surrounding tissues and many of the findings are etiologically nonspecific. The aim of this review is to discuss the etiologies and accompanying adverse tissue reactions seen with prosthesis failure, including the findings seen in aseptic lymphocyte-dominated vasculitis-associated lesion. Aseptic lymphocyte-dominated vasculitis-associated lesion is an important diagnostic consideration as its proposed pathogenesis is a type VI hypersensitivity response to metal ions. In addition, we also propose a diagnostic algorithm that incorporates clinical and histopathologic findings to suggest an etiologic cause. This proposed algorithm may be clinically useful as, to date, there is no consensus on nomenclature.

Diagnosis of Acute Cellular Rejection and Antibody-Mediated Rejection on Lung Transplant Biopsies: A Perspective From Members of the Pulmonary Pathology Society.

CONTEXT: - The diagnosis and grading of acute cellular and antibody-mediated rejection (AMR) in lung allograft biopsies is important because rejection can lead to acute graft dysfunction and/or failure and may contribute to chronic graft failure. While acute cellular rejection is well defined histologically, no reproducible specific features of AMR are currently identified. Therefore, a combination of clinical features, serology, histopathology, and immunologic findings is suggested for the diagnosis of AMR. OBJECTIVE: - To describe the perspective of members of the Pulmonary Pathology Society (PPS) on the workup of lung allograft transbronchial biopsy and the diagnosis of acute cellular rejection and AMR in lung transplant. DATA SOURCES: - Reports by the International Society for Heart and Lung Transplantation (ISHLT), experience of members of PPS who routinely review lung allograft biopsies, and search of literature database (PubMed). CONCLUSIONS: - Acute cellular rejection should be assessed and graded according to the 2007 working formulation of the ISHLT. As currently no specific features are known for AMR in lung allografts, the triple test (clinical allograft dysfunction, donor-specific antibodies, pathologic findings) should be used for its diagnosis. C4d staining might be performed when morphologic, clinical, and/or serologic features suggestive of AMR are identified.

Utilization review and reimbursement of cytology services in endobronchial ultrasound-guided procedures: challenge and opportunity.

INTRODUCTION: The roles of pathologists and cytotechnologists (CTs) continually evolve to optimize patient care, particularly with regard to rapid on-site evaluation (ROSE). Having ROSE performed helps ensure sufficient material is obtained for diagnosis and permits appropriate specimen triage for ancillary studies. At our institution, both on-site and telecytology evaluations are increasingly utilized, particularly in endobronchial ultrasound-guided procedures (EBUS). Consequently, time demands placed on the pathologist and CT staff has significantly increased, creating workload management challenges. MATERIAL AND METHODS: A consecutive number of ROSE procedures were documented for a 3-month time period at our institution. Case type and time spent for travel, adequacy assessment, processing, screening, and sign-out was recorded in order to assess time demands placed on staff by different procedures. RESULTS: Average travel/processing time by CTs was variable among ROSE procedures (72.9 minutes), as was adequacy assessment time by pathologists (16.9 minutes). EBUS posed the greatest time challenges with the longest CT travel/processing time as EBUS took almost 40% longer and adequacy assessment took the pathologist 3-4 times longer when compared with other procedures because of the targeting of multiple sites during EBUS with associated procedural delays. Using telecytology, average pathologist adequacy assessment time was reduced from 44.8 minutes to 24.6 minutes for EBUS. The provision of ROSE for EBUS is more challenging from a workload management perspective than for other procedures. CONCLUSIONS: ROSE reimbursement is low, and no greater for EBUS than for other procedures. Use of telecytology can save time for pathologists and make the service more cost-effective if the number of procedures is sufficient to justify investment in the technology.

Glycogenic Hepatopathy: Thinking Outside the Box.

Glycogenic hepatopathy (GH) remains underrecognized in adults as most clinicians mistake it for the more common hepatic abnormality associated with uncontrolled diabetes mellitus in this age group, non-alcoholic fatty liver disease. This is also complicated by the fact that both entities are indistinguishable on liver ultrasound. We herein describe a similar predicament in which a young adult female presented with bilateral upper quadrant abdominal pain, tender hepatomegaly, lactic acidosis and a >10-fold increase in liver enzymes, which worsened after the administration of high-dose steroids. Despite intravenous normal saline resuscitation, serum transaminitis persisted in a fluctuating manner. Ultimately, a liver biopsy confirmed GH. Biochemically, GH is driven by high amounts of both circulating glucose and insulin or by the administration of high-dose steroids. Improving glycemic control is the mainstay of treatment for GH. However, in our case, improvement in glycated hemoglobin of just 0.6% was enough to achieve symptomatic relief, supporting recent claims of the involvement of other identified factors in disease development.

Squamous cell carcinoma of the cervix: a cytology-histology-human papillomavirus correlation in clinical practice.

CONTEXT: Cervical cancer mortality has declined by 74% in the United States since the implementation of the Papanicolaou (Pap) test. Nevertheless, more than 12,000 US women annually develop cervical cancer, and squamous cell carcinoma (SqCa) remains the predominant cervical malignancy. OBJECTIVE: To evaluate screening techniques used in the detection of SqCa of the cervix and provide insights regarding which technique(s) is (are) most efficacious in our study population. DESIGN: We retrospectively reviewed all available cytologic, human papillomavirus (HPV), and histologic malignancy burden data from patients diagnosed with SqCa. The clinical data were collected from 2 geographically and socioeconomically diverse hospital systems. Cases in which identified patients had a Pap test with a negative result/unsatisfactory specimen within 5 years of SqCa tissue diagnosis were considered Pap test screening failures. Cases in which patients were diagnosed with HPV-negative SqCa were considered HPV screening failures. RESULTS: Eighty-eight cases (patients' ages ranging from 19 to 73 years) were identified. Of those, cytologic history was available for 64 cases present in our electronic medical history record. Three cases were cytology screening failures (one being an unsatisfactory specimen) and 3 cases were HPV screening failures (one being the cytologic unsatisfactory case). Although measuring sensitivity in practice has limitations, we calculated the SqCa detection sensitivity at 95.3% by Pap test alone and 97% when HPV DNA testing was incorporated. CONCLUSIONS: Our results highlight the necessity of combining Pap and HPV testing. Although the number of cases identified is relatively small, our data suggest detection failures will decrease as the practice of combining HPV and Pap testing increases.

Glandular Lesions of the Cervix in Clinical Practice: A Cytology, Histology, and Human Papillomavirus Correlation Study From 2 Institutions.

CONTEXT: The Papanicolaou (Pap) test has indisputably decreased cervical cancer mortality, as rates have declined by up to 80% in the United States since its implementation. However, the Pap test is considered less sensitive for detecting glandular lesions than for detecting those of squamous origin. Some studies have even suggested an increasing incidence of cervical adenocarcinoma, which may be a consequence of a relatively reduced ability to detect glandular lesions with cervical cancer screening techniques. OBJECTIVE: To evaluate the detection rate of glandular lesions with screening techniques currently used for cervical cancer screening and to provide insight as to which techniques are most efficacious in our study population. DESIGN: We retrospectively reviewed any available cytology, human papillomavirus (HPV), and histologic malignancy data in patients diagnosed with adenocarcinoma in situ and adenocarcinoma from 2 geographically and socioeconomically disparate hospital systems. Identified patients having had a negative/unsatisfactory Pap test within 5 years of adenocarcinoma in situ or adenocarcinoma tissue diagnosis were considered Pap test screening failures. Patients with negative HPV tests on cytology samples were considered HPV screening failures. RESULTS: One hundred thirty cases were identified (age range, 22-93 years); 39 (30%) had no Pap history in our files. Eight of 91 remaining cases (8.8%) were screening failures. The detected sensitivity for identifying adenocarcinoma in situ/adenocarcinoma in this study was 91.2% by cytology alone and 92.3% when incorporating HPV testing. The most common cytologic diagnosis was atypical glandular cells (25 cases), and those diagnosed with adenocarcinoma were 7.4 years older than those diagnosed with adenocarcinoma in situ (50.3 versus 42.9 years). Nine of 24 HPV-tested cases (37.5%) were called atypical squamous cell of undetermined significance on cytology. CONCLUSIONS: Our results highlight the importance of combined Pap and HPV cotesting. Although the number of cases identified is relatively small, our data suggest screening for squamous lesions facilitates the recognition of glandular lesions in the cervix. Additionally, increased use of combined Pap and HPV cotesting may decrease detection failure rates with regard to glandular lesions.

Pheochromocytoma - review and biochemical workup.

A commonly received question in the clinical laboratory is as follows: what is the best test for pheochromocytoma? A widely variable presentation and potentially catastrophic consequence make this a feared neoplasm despite its infrequent encounter. Because various biochemical testing modalities are available, test selection is often confusing. This selection process can be made easier through a better understanding of catecholamine producing neoplasms. The aim of this article is to provide a review of catecholamine producing neoplasms and give recommendations on appropriate test selection.

Strategic use of preparative chiral chromatography for the synthesis of a preclinical pharmaceutical candidate.

The modern use of preparative chromatography in pharmaceutical development is illustrated by the case of a recent preclinical candidate from these laboratories. The synthesis of the candidate employed a coupling of two enantiopure intermediates, each of which could be resolved using preparative chiral chromatography. SFC screening was employed to identify the enantioselective stationary phases, and semipreparative SFC methods derived from this screening were used to produce gram amounts of enantiopure intermediate for initial studies. However, initial larger scale resolution required the translation of the SFC methods to HPLC conditions. Preparative chiral HPLC on a 30-cm i.d. column was then used to produce enantiopure intermediates which were coupled to give 170 g of the preclinical candidate. Subsequent preparation of the candidate at larger scale for later-stage clinical evaluation employed an improved synthesis in which one component was constructed by asymmetric synthesis. Resolution of the other component, now a more advanced intermediate, was carried out using newly obtained large-scale SFC equipment. Some discussion is presented on the varying strategies whereby preparative chiral chromatography can be used to support either short-term or long-term synthetic goals in preclinical pharmaceutical development.

A convenient oxazole C-2 protecting group: the synthesis of 4- and 5-substituted oxazoles via metalation of 2-triisopropylsilyloxazoles.

[reaction: see text] Metalation of oxazoles at the 4 and 5 position was achieved after regioselective C-2 silyl protection. Removal of the protecting group was then accomplished under mild conditions allowing for a straightforward preparation of C-5 monosubstituted and C-4,5 disubstituted oxazoles. The first practical C-2 protecting group of oxazoles has been demonstrated.

Chiral recognition of cyclic alpha-hydroxyketones by CD-sensitive zinc tetraphenylporphyrin tweezer.

A combined chemical/chiroptical microscale protocol for the determination of absolute configurations of cyclic alpha-hydroxyketones is described. The hydroxyl group in cyclic alpha-hydroxyketones is converted into (3-aminopropylamino)acetate (NH2CH2CH2CH2NHCH2COOR), or more generally, according to a newly developed protocol, into (3-hydroxypropylamino)acetate group (HOCH2CH2CH2NHCH2COOR). The resultant conjugated compound forms a 1:1 host-guest complex with a dimeric zinc porphyrin tweezer, which exhibits exciton-coupled bisignate CD spectrum centered around the 420-nm porphyrin Soret band due to induced helicity between the two porphyrins in the complex. The absolute configurations of the alpha-stereogenic center is then determined by comparison of the sign of the observed CD exciton couplet of the complex with that of the preferred porphyrin twist predicted by the Merck Molecular Force Field (MMFFs) method.

Asymmetric synthesis of cyclic hydroxy ketones derived from enol ethers via sharpless asymmetric dihydroxylation. A study in the correlation of the enol ether chain length and enantioselectivity.

The Sharpless asymmetric dihydroxylation reaction of enol ethers derived from their corresponding cyclic ketones, gave alpha-hydroxyketones with high enantioselectivity. The enantiomeric excess was found to be proportional to the length of the unbranched enol ether chain with a maximum ee for the pentyl enol ether. An efficient synthesis of alpha-hydroxy chromanone in >90% ee was demonstrated using this method.