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Introduction Voided urine cytology is a non-invasive method for diagnosing and following up on bladder carcinoma. But this exam suffers from low sensitivity and a lack of reproducibility due to pathologist examiner dependency, especially in low-grade tumours. Cystoscopy is chosen as the gold-standard exam, although it is considered aggressive by patients, especially when it is repeated. The aim of this study is to explore artificial intelligence (AI) to improve voided cytology. Material and method A national prospective multicentre trial (14 centres) was conducted on 1,360 patients, divided in two groups. The first group included bladder carcinoma diagnosis with different histological grades and stages, and the second group included control patients based on negative cystoscopy and cytology results. The first step of this VISIOCYT1 trial focused on algorithm development and the second step on validating this algorithm. A total of 598 patients were included in this first step, 449 patients with bladder tumours (219 high-grade and 230 low-grade) and 149 as negative control. The VisioCyt test was compared to voided urine cytology performed by experienced uro-pathologists from each centre. Results Overall sensitivity was highly improved by the VisioCyt test compared to cytology, respectively 84.9% vs. 43%. For high-grade tumours, VisioCyt test sensitivity was 92.6% compared to 61.1% for uro-pathologists. Regarding low-grade tumours, VisioCyt test sensitivity was 77% versus 26.3% for uro-pathologists. Conclusion Compared to routine cytology, the first phase of VISIOCYT1 results shows very clear progress in term of sensitivity. It is particularly visible and interesting for low-grade tumours. If the validation cohort confirms these results, it could lead to the VisioCyt test being considered as a very useful aid for pathologists. Moreover, as this test is in fact software based on artificial intelligence, it should become more and more efficient as more data is collected.

BACKGROUND Overexpression of p16INK4a has been identified in urothelial malignancies both cytologically and histologically. In addition, p16/Ki-67 dual labeling has been shown to identify high-grade urothelial cancer cells and some progression cases within a 12-month delay. The Paris System for Reporting Urinary Cytology (TPS) was published in late 2015. Its aim is to clarify the criteria for diagnosing or, conversely, excluding high-grade urothelial carcinoma (HGUC). METHODS Dual labeling was performed on archived ThinPrep-based Papanicolaou slides. A total of 208 samples (negative for high-grade urothelial carcinoma [NHGUC], 59; consistent with low-grade urothelial neoplasia [LGUN], 24; atypical urothelial cells [AUC], 15; and suspicious for or showing HGUC, 110) were analyzed for p16/Ki-67 after reclassification according to TPS. We assessed the oncologic status of the patients with cystoscopy, urinary cytology, histology, and prolonged 36-month follow-up data. RESULTS The sensitivity of p16/Ki-67 for life-threatening lesions was not different from that of urinary cytology (82.8% vs 83.6%; P = 1). However, among patients with samples classified as NHGUC and AUC, disease-free survival was significantly shorter for dual-labeled cases versus cases with negative dual labeling (P < .0001). The same tendency was observed in patients with histologically proven LGUN (P < .0001). As for specificity in patients with negative cystoscopy and cytology combined, prolonged follow-up showed 90% overall survival at 24 months. CONCLUSIONS A long-term evaluation of p16/Ki-67 dual labeling may identify HGUC and progression in cases with negative/low-grade urinary cytology results, and there are potential implications for the clinical management of patients after the conservative treatment of non–muscle-invasive urothelial carcinoma. Cancer Cytopathol 2017;125:552–62. © 2017 American Cancer Society.

Objective Since the guidelines of the International Committee for Standardisation in Haematology (ICSH) in 1984 and those of the European Committee for External Quality Assessment Programmes in Laboratory Medicine (EQALM) in 2004, no leading organisation has published technical recommendations for the preparation of air-dried cytological specimens using May-Grunwald–Giemsa (MGG) staining. Data sources Literature data were retrieved using reference books, baseline-published studies, articles extracted from PubMed/Medline and Google Scholar, and online-available industry datasheets. Rationale The present review addresses all pre-analytical issues concerning the use of Romanowsky's stains (including MGG) in haematology and non-gynaecological cytopathology. It aims at serving as actualised, best practice recommendations for the proper handling of air-dried cytological specimens. It, therefore, appears complementary to the staining criteria of the non-gynaecological diagnostic cytology handbook edited by the United Kingdom National External Quality Assessment Service (UK-NEQAS) in February 2015.

Objectifs La cytologie urinaire mictionnelle (CUM) est non invasive, mais souffre d’un manque de sensibilite et reste pathologiste dependante, notamment dans les lesions urotheliales de bas grade. Afin d’ameliorer le diagnostic sur les urines, VitaDX a developpe dans le cadre de l’essai clinique VISIOCYT un dispositif medical qui utilise des algorithmes de traitement d’image et de machine learning (IA) pour predire le caractere tumoral des CUM. Methodes VISIOCYT est une etude clinique prospective multicentrique, portant sur 1360 patients repartis en deux groupes : les temoins negatifs (TNeg) confirmes par une CUM et une endoscopie negatives et les patients porteur d’une tumeur urotheliale (PTU) par le resultat d’une histologie positive de carcinome urothelial. Les resultats de cette communication portent sur 586 patients ; 127 TNeg et 459 PTU (242 haut grade et 217 bas grade). Les performances ont ete evaluees par validation croisee, une approche statistique standard en IA. Les 586 patients ont donc ete subdivises aleatoirement 42 fois dans les proportions suivantes : 5/6 des donnees pour l’apprentissage et 1/6 pour l’evaluation. Resultats A l’issue de la validation croisee, les mesures de performance obtenues sur chacune des 42 partitions sont moyennees pour evaluer les performances de l’algorithme. La sensibilite moyenne de l’algorithme est de 82,6 % pour une specificite de 71,4 %. En detail, on observe une sensibilite moyenne de 89,3 % pour les diagnostics de haut grades et de 76,4 % pour les diagnostics de bas grade. Sur les memes partitions, a titre de comparaison, la sensibilite moyenne des praticiens de l’essai clinique est de 43 % sur l’ensemble des malades, avec en particulier une sensibilite de 61,1 % pour les haut grades et de 26,3 % pour les bas-grades. La specificite des praticiens est par defaut de 100 % car la cytologie negative est un critere d’inclusion pour le groupe temoin ( Tableau 1 ). Conclusion L’approche algorithmique (IA) semble permettre un meilleur diagnostic des lesions urotheliales de bas grade par rapport a la CUM traditionnelle. Ces resultats devront etre confirmes a l’issu de l’essai clinique en sachant que les performances devraient augmenter avec l’accumulation de donnees supplementaires.

May-Grunwald-Giemsa (MGG) stain is a Romanowsky-type, polychromatic stain as those of Giemsa, Leishman and Wright. Apart being the reference method of haematology, it has become a routine stain of diagnostic cytopathology for the study of air-dried preparations (lymph node imprints, centrifuged body fluids and fine needle aspirations). In the context of their actions of promoting the principles of quality assurance in cytopathology, the French Association for Quality Assurance in Anatomic and Cytologic Pathology (AFAQAP) and the French Society of Clinical Cytology (SFCC) conducted a proficiency test on MGG stain in 2013. Results from the test, together with the review of literature data allow pre-analytical and analytical steps of MGG stain to be updated. Recommendations include rapid air-drying of cell preparations/imprints, fixation using either methanol or May-Grunwald alone for 3-10minutes, two-step staining: 50% May-Grunwald in buffer pH 6.8 v/v for 3-5minutes, followed by 10% buffered Giemsa solution for 10-30minutes, and running water for 1-3minutes. Quality evaluation must be performed on red blood cells (RBCs) and leukocytes, not on tumour cells. Under correct pH conditions, RBCs must appear pink-orange (acidophilic) or buff-coloured, neither green nor blue. Leukocyte cytoplasm must be almost transparent, with clearly delineated granules. However, staining may vary somewhat and testing is recommended for automated methods (slide stainers) which remain the standard for reproducibility. Though MGG stain remains the reference stain, Diff-Quik(®) stain can be used for the rapid evaluation of cell samples.

BACKGROUND p16INK4a Is overexpressed in almost all precancerous and carcinomatous lesions of the uterine cervix, secondary to interference between high-risk human papillomaviruses (hr-HPVs) and the retinoblastoma gene product. Overexpression of p16INK4a has also been identified in patients with high-grade urothelial lesions, both cytologically and histologically. However, the etiological role of HPV has not been documented except in inverted papillomas, low-grade bladder tumors, and younger patients. We therefore attempted to verify if HPV DNA was detectable in p16INK4a-positive urothelial tumors. METHODS A total of 90 urinary cytology samples (33 negative/low-grade cases and 57 high-grade proliferations) were analyzed for p16INK4a and HPV DNA. HPV genotyping was performed by polymerase chain reaction using a low-density DNA microarray enabling the detection of 35 HPVs. A reasoned approach combining tissue genotyping and in situ hybridization (ISH) for hr-HPVs was used in patients with urinary HPV. RESULTS Low-risk HPV (HPV-84) and hr-HPVs (HPV-16, -31, and -70) were detected. The prevalence of hr-HPVs in the urine was low: 5 of 82 patients (6.1%) and only 4 of 50 patients (8.0%) with high-grade urothelial malignancy. p16INK4a overexpression was noted in 49 high-grade samples (85.9%). In patients with p16INK4a-positive tumor cells and hr-HPV in the urine, HPV genotyping and ISH for hr-HPVs were negative in matched tissue sections. CONCLUSIONS Our study shows a low prevalence of hr-HPVs in the urine of patients with high-grade urothelial malignancy. In those, p16INK4a overexpression occurs in the absence of demonstrable HPV DNA in the tissue sections, contrary to what is noted in gynecopathology. Cancer (Cancer Cytopathol) 2014;122:760–769. © 2014 American Cancer Society.

Objective Co-expression of p16INK4a protein and Ki-67 (p16/Ki-67) is noted in almost all high-grade urothelial lesions. However, the aetiological role or, conversely, the absence of causative effect of high-risk human papillomaviruses (hr-HPVs) has not been documented. The purpose of this study is to evaluate HPV DNA in p16/Ki-67-positive, high-grade urothelial tumour cells. Methods Fifty-seven urine samples collected from 50 patients, including 55 histologically proven high-grade proliferations and two cases with clinical evidence of malignancy, were analysed for p16/Ki-67. Immunolabelling was performed in destained Papanicolaou-stained slides after ThinPrep® processing. HPV genotyping was performed by polymerase chain reaction (PCR) using a DNA microarray for 35 HPV types. Confirmation of the presence (or absence) of HPV in tissue samples was verified using a reasoned approach combining PCR and in situ hybridization (ISH) for hr-HPVs. Results Co-expression of p16/Ki-67 was noted in 43 of 57 (75.4%) cases. In these, hr-HPVs 16, 31 and 70, and low risk HPV 84, were detected in the urine in four patients (8%). Upregulation of p16INK4a protein was confirmed on bladder biopsy or transurethral resection specimens, but PCR and ISH for hr-HPVs were both negative on the tissue sections. Conclusion Our results show a low prevalence of HPV infection in the urinary tract of patients with p16/Ki-67-positive urothelial malignancy. The study confirms that the deregulated cell cycle, as demonstrated by p16/Ki-67 dual labelling, is independent of the oncogenic action of hr-HPVs in high-grade urothelial proliferations.

BACKGROUND Taking into consideration the known overexpression of p16INK4a in histologically demonstrated high-grade urothelial malignancies, the objective of the current study was to examine the value of p16INK4a overexpression and of p16/Ki-67 dual labeling versus urinary cytology in the detection of urothelial lesions. METHODS Immunolabeling was performed on demounted and destained Papanicolaou slides after liquid-based ThinPrep processing. Actual diagnoses were ascertained by cystoscopy controls and histopathology. Negative cases, papillary urothelial neoplasia of low malignant potential/low-grade tumor, and high-grade lesions were considered separately. RESULTS A total of 216 urine samples were collected from new patients with symptoms who were referred for cystoscopy (92 cases) or patients who were being followed after conservative treatment for lesions involving the bladder (117 cases) or the upper urinary tract (7 cases). p16INK4a positivity was assessed in 171 of the 216 cases (79.2%) and in 93 of 99 high-grade cases with positive cytology (93.9%). Coexpression of p16/Ki-67 in the same cells was observed in 119 of 216 cases (55.1%) and was noted in 18 of 51 cases of negative or papillary urothelial neoplasia of low malignant potential/low-grade tumor (35.3%) and in 80 of 101 high-grade tumors (79.2%) (P

In digestive cancers, the prognostic significance of intraperitoneal free cancer cells remains unclear (IPCC). The main objective of this study was to assess the prognostic significance of IPCC in colorectal and gastric adenocarcinoma. The secondary objectives were to evaluate the predictive significance of IPCC for the development of peritoneal carcinomatosis (PC) and to evaluate the prevalence of synchronous PC and IPCC.This was a prospective multicentre study. All patients undergoing surgery for a digestive tract cancer had peritoneal cytology taken. Patients with gastric and colorectal cancer with no residual tumour after surgery and no evidence of PC were followed-up for 2 years. The primary end point was overall survival.Between 2002 and 2007, 1364 patients were enrolled and 956 were followed-up over 2 years. Prevalence of IPCC was 5.7% in colon cancer, 0.6% in rectal cancer and 19.5% in gastric cancer. The overall 2-year survival rate for patients with IPCC was 34.7% versus 86.8% for patients with negative cytology (p0.0001). By multivariate analysis, IPCC was not an independent prognostic factor. No relationship between cytology and recurrence was found.The presence of IPCC was not an independent prognostic and didn't add any additional prognostic information to the usual prognostic factors related to the tumour (pTNM and differentiation). Moreover the presence of IPCC detected with this method didn't appear to predict development of PC. Peritoneal cytology using conventional staining doesn't seem to be a useful tool for the staging of colorectal and gastric cancers.

Resume Le demembrement des cytologies urinaires atypiques est un enjeu moderne de l’hybridation in situ en fluorescence (FISH). Un resultat negatif obtenu avec le kit multisonde UroVysion ® en presence de cellules urotheliales atypiques est en faveur d’alterations reactionnelles benignes et doit inciter a limiter les explorations invasives. Toutefois, une FISH negative en cas de cytologie negative ou atypique n’exclut pas une neoplasie urotheliale de bas grade. Les atypies urotheliales sont particulierement marquees apres chirurgie conservatrice de la vessie, et notamment apres BCG-therapie pour carcinome in situ ou les alterations benignes peuvent prendre des aspects inquietants. Dans cette circonstance, une FISH positive malgre l’absence de haut grade cytologique indiquera une persistance ou une recurrence du carcinome. Il est a noter toutefois que les aberrations chromosomiques ne sont pas caracteristiques du cancer mais peuvent survenir dans des cellules benignes : la tetraploidie avec duplication equilibree du genome ou la polyploidie peut etre observee dans les cellules Decoy, apres radiotherapie et en cas de lithiase. Ainsi, une FISH positive apres radiotherapie pelvienne pour adenocarcinome prostatique n’indiquera un cancer que s’il existe une deletion du 9p21, plus specifique. Les etudes recentes montrent qu’une attitude agressive n’est pas justifiee en cas de test UroVysion ® positif avec cytologie suspecte, si la cystoscopie est negative. Toutefois, la FISH reste un outil excellent pour ameliorer les performances diagnostiques de la cytologie urinaire, a condition de prendre en compte les donnees cliniques et de se souvenir que la FISH n’est d’aucune utilite en cas de cytologie positive de haut grade.

E. Piaton, L. Villeneuve, C. Maurice, C. Paulin, M. Cottier, B. Fontaniere, M. Salle, D. Seigneurin, S. Vancina, E. Decullier, F. N. Gilly and E. Cotte Intraperitoneal free cancer cells in non-gynaecological adenocarcinomas: a reproducibility study Objective: In recent years, therapeutic approaches including cytoreductive surgery followed by intraperitoneal chemotherapy have proven effective in peritoneal carcinomatosis of colorectal origin. If cytology is to be used to include patients in aggressive treatment regimens, it is necessary to evaluate its performance, particularly in terms of specificity. The aim of this study was to assess interobserver agreement for the detection of intraperitoneal free cancer cells (IFCCs) in patients with non-gynaecological adenocarcinomas. Methods: Over a 5-year period, 1223 patients were recruited in 19 French surgery departments. Peritoneal samples were examined in 14 dispersed pathology laboratories. Giemsa-stained slides were sent to a control reader blind to the previous diagnosis. Discordant cases, concordant positive results and a random selection of negative concordant cases were reviewed by a panel of seven cytopathologists. The ‘final diagnosis’ was that of the consensus meetings but took into account locally-processed slides. Results: Gathering dubious cases with negative results, a 95.6% concordance was achieved between local readers and the control reader. IFCCs were ascertained by the panel in 85 cases (7.0%). Eight of 873 colorectal cancers cases viewed locally were falsely positive (0.9%). Radiotherapy and neoadjuvant therapy had no impact on the false-positive rate as assessed by final validation by the panel (P > 0.05). Samples initially considered as dubious were reclassified as negative by the panel in 24 of 25 cases (96.0%). Conclusions: The panel consensus allowed reclassification of most dubious/equivocal peritoneal cytology cases, whereas clearcut distinction between benign and malignant cases was correctly achieved in almost all cases.

V. Hofman, E. Long, M. Ilie, C. Bonnetaud, J. M. Vignaud, J. F. Flejou, S. Lantuejoul, E. Piaton, N. Mourad, C. Butori, E. Selva, C. H. Marquette, M. Poudenx, S. Sibon, S. Kelhef, N. Venissac, J. P. Jais, J. Mouroux, T. J. Molina, P. Vielh and P. Hofman Morphological analysis of circulating tumour cells in patients undergoing surgery for non-small cell lung carcinoma using the isolation by size of epithelial tumour cell (ISET) method Background and objective: Recurrence rates after surgery for non-small cell lung cancer (NSCLC) range from 25 to 50% and 5-year survival is only 60–70%. Because no biomarkers are predictive of recurrence or the onset of metastasis, pathological TNM (pTNM) staging is currently the best prognostic factor. Consequently, the preoperative detection of circulating tumour cells (CTCs) might be useful in tailoring therapy. The aim of this study was to characterize morphologically any circulating non-haematological cells (CNHCs) in patients undergoing surgery for NSCLC using the isolation by size of epithelial tumour cell (ISET) method. Methods: Of 299 blood samples tested, 250 were from patients with resectable NSCLC and 59 from healthy controls. The presence of CNHCs was assessed blindly and independently by 10 cytopathologists on May-Grunwald–Giemsa stained filters and the cells classified into three groups: (i) malignant cells, (ii) uncertain malignant cells, and (iii) benign cells. We assessed interobserver agreement using Kappa (κ) analysis as the measure of agreement. Results: A total of 123 out of 250 (49%) patients showed CNHCs corresponding to malignant, uncertain malignant and benign cells, in 102/250 (41%), 15/250 (6%) and 6/250 (2%) cases, respectively. No CNHCs were detected in the blood of healthy subjects. Interobserver diagnostic variability was absent for CNHCs, low for malignant cells and limited for uncertain malignant and benign cells. Conclusion: Identification of CTCs in resectable NSCLC patients, using ISET technology and according to cytopathological criteria of malignancy, appears to be a new and promising field of cytopathology with potential relevance to lung oncology.

Detection of circulating tumor cells (CTCs) morphologically may be a promising new approach in clinical oncology. We tested the reliability of a cytomorphologic approach to identify CTCs: 808 blood samples from patients with benign and malignant diseases and healthy volunteers were examined using the isolation by size of epithelial tumor cell (ISET) method. Cells having nonhematologic features (so-called circulating nonhematologic cells [CNHCs]) were classified into 3 categories: CNHCs with malignant features, CNHCs with uncertain malignant features, and CNHCs with benign features. CNHCs were found in 11.1% and 48.9% of patients with nonmalignant and malignant pathologies, respectively (P < .001). CNHCs with malignant features were observed in 5.3% and in 43.1% of patients with nonmalignant and malignant pathologies, respectively. Cytopathologic identification of CTCs using the ISET method represents a promising field for cytopathologists. The possibility of false-positive diagnosis stresses the need for using ancillary methods to improve this approach.

The diagnosis of “suspicious for high-grade urothelial carcinoma (SHGUC)” is meant to reflect the presence of urothelial cells with severe atypia that falls short for a diagnosis of high-grade urothelial carcinoma (HGUC) but beyond atypia that is associated with the “atypical urothelial cells” (AUC). This chapter lists the quantitative and qualitative cytomorphologic criteria needed for a diagnosis of SHGUC.

Atypia is admittedly a difficult topic to cover no matter what the organ system it is in. The most important reason for this is its subjective nature – it is in the eye of the beholder. In keeping with the mantra of The Paris System for Reporting Urinary Cytology, the goal for the “atypical urothelial cells” (AUC) category is to capture the cases worrisome for high-grade urothelial carcinoma that fall short of the “suspicious for high-grade urothelial carcinoma” (SHGUC) category. It is important to ensure that known causes of atypia such as polyomavirus change, treatment effect, etc., are classified as “negative for high-grade urothelial carcinoma” (NHGUC), and not under the AUC category. In order to make this a tangible and more objective category we have set forth criteria for AUC using four main cytomorphologic features: nuclear to cytoplasmic (N/C) ratio, nuclear chromasia, chromatin pattern, and features of the chromatinic rim. From a clinical standpoint, whenever the diagnosis of AUC is rendered on a case, this inconclusive category may create confusion and anxiety in the patient, and may lead to unnecessary procedures. Therefore, we recommend that the “AUC” interpretation should be kept at a minimum, and not be used as a waste basket.

Cells from serous effusions can easily be concentrated by centrifugation. Thereafter, various procedures allow cells to be deposited on glass slides. Standard stains give excellent morphologic details for analysis. However, unsatisfactory specimens are frequently observed in daily practice. Standardization brought by liquid based cytology may be of some help for immunocytochemistry, but conventional methods remain essential. Several decades of experience and analysis of literature allow the authors to select the best criteria for interpretation, with emphasis on the differential diagnosis of malignant vs. benign effusions.

Immunocytochemistry may be of valuable help in typing most malignant serous effusions. Various preparatory methods are described including centrifugation and smearing, cytocentrifugation, cell blocks and liquid based cytology. The value and usefulness of immunocytochemistry depends on the experience of each pathologist. The use of a panel of antibodies associating markers of carcinomas with markers of mesothelial cells is recommended.

p53 could help identify bladder tumour cases with a risk of progression from superficial to invasive disease. Semiautomatic, liquid-based cytology (LBC) techniques offer an opportunity to standardise molecular techniques. The aim of our study was to investigate whether LBC could improve p53 immunolabelling, and to assess whether urinary p53 could have a prognostic value. Immunoreactivity for p53 was studied in 198 urine samples after treatment with the Cytyc Thinprep processor. After antigen retrieval, cells were labelled with a monoclonal antibody that recognises both wild-type and mutant form of the p53 protein (Clone DO-7, Dako), 1/1000. Positivity for p53 was assessed in 17.2% of the cases. High-grade (G3) tumours were positive in 74.1% of the cases. Comparatively, low-grade (G1-2) urothelial carcinomas were positive in 23.5% of the cases. During a median follow-up period of 26 months, recurrence was observed in 52.9% of the cases with p53 overexpression, and in only 10.9% of negative cases (P0.001). The progression rate was 35.3% of p53-positive cases vs 5.5% of p53-negative cases (P0.001). Progression-free survival was significantly shorter in patients with p53 accumulation (P = 0.007). In a multivariate analysis stratified on grade and stage, p53 was an independent predictor of overall survival (P = 0.042). The results show that using Thinprep LBC, p53 immunolabelling of voided urothelial cells allows most high-grade tumours to be detected and may help identify cases with a higher risk of recurrence and progression.

Objective We studied whether atypical, non-superficial urothelial cells (AUC) could be separated into new subcategories including AUC ‘of undetermined significance’ (AUC-US) and ‘cannot exclude high grade’’ (AUC-H) in order to help to standardize urine cytopathology reports, as it is widely accepted in the Bethesda system for gynaecological cytopathology. Methods We investigated whether AUC-US and AUC-H, defined by distinctive cytological criteria, might be separated with statistical significance according to actual diagnosis and follow-up data. A series of 534 cyto-histological comparisons taken in 139 patients, including 221 AUC at various steps of their clinical history was studied. There were 513 (96.1%) postcystoscopy and 469 (87.8%) ThinPrep® liquid-based specimens (95.9% and 89.1% of AUC cases, respectively). Patients viewed between 1999 and 2011 had histological control in a 0- to 6-months delay and were followed-up during an additional 5.9 ± 9.2 (0- to 56-) months period. Results The 221 AUC represented 0.8–2% of the specimens viewed during the study period. Among AUC-H cases, 70 out of 185 (37.8%) matched with high-grade lesions, compared with 3 of 38 (8.3%) of AUC-US cases (P = 0.0003). Conservatively treated patients with AUC-H more frequently developed high-grade lesions than those with AUC-US (54.1% versus 16.7%, P = 0.0007) with a 17.6-months mean delay. Nuclear hyperchromasia, a nuclear to cytoplasm (N/C) ratio > 0.7 and the combination of both were the more informative diagnostic criteria, all with P

Overexpression of p16(INK4a) independent of the presence of E6-E7 oncoproteins of high-risk papillomaviruses has been identified in bladder carcinoma in situ lesions with or without concurrent papillary or invasive high-grade (HG) urothelial carcinoma. As p16(INK4a) and Ki-67 co-expression clearly indicates deregulation of the cell cycle, the aim of this study was to investigate the frequency of p16(INK4a) /Ki-67 dual labelling in urinary cytology samples.Immunolabelling was performed in demounted, destained Papanicolaou slides after ThinPrep(®) processing. A total of 84 urinary cytology samples (18 negative, 10 low grade, 19 atypical urothelial cells and 37 high grade) were analysed for p16(INK4a) /Ki-67 co-expression. We assessed underlying urothelial malignancy with cystoscopy, histopathology and follow-up data in every case.Compared with raw histopathological results, p16 (INK4a) /Ki-67 dual labelling was observed in 48 out of 55 (87.3%) HG lesions and in 11 out of 29 (37.9%) negative, papillary urothelial neoplasia of low malignant potential or low-grade carcinomas (P = 0.05). All cases with high-grade/malignant cytology were dual labelled. Sixteen out of 17 (94.1%) carcinoma in situ cases and eight out of 14 (57.1%) cases with atypical urothelial cells matching with HG lesions were dual labelled. Extended follow-up allowed three cases of progression to be diagnosed in dual-labelled cases with negative/low-grade cytology results after a 9- to 11-months delay.The data show that p16(INK4a) /Ki-67 co-expression allows most HG cancer cells to be detected initially and in the follow-up period. Additional studies are needed in order to determine whether dual labelling can be used as a triage tool for atypical urothelial cells in the urine.

E. Piaton, A. S. Advenier, C. Carre, M. Decaussin-Petrucci, F. Mege-Lechevallier and A. Ruffion p16INK4a/Ki-67 dual labelling as a marker for the presence of high-grade cancer cells or disease progression in urinary cytopathology Objective: Overexpression of p16INK4a independent of the presence of E6–E7 oncoproteins of high-risk papillomaviruses has been identified in bladder carcinoma in situ lesions with or without concurrent papillary or invasive high-grade (HG) urothelial carcinoma. As p16INK4a and Ki-67 co-expression clearly indicates deregulation of the cell cycle, the aim of this study was to investigate the frequency of p16INK4a/Ki-67 dual labelling in urinary cytology samples. Methods: Immunolabelling was performed in demounted, destained Papanicolaou slides after ThinPrep® processing. A total of 84 urinary cytology samples (18 negative, 10 low grade, 19 atypical urothelial cells and 37 high grade) were analysed for p16INK4a/Ki-67 co-expression. We assessed underlying urothelial malignancy with cystoscopy, histopathology and follow-up data in every case. Results: Compared with raw histopathological results, p16 INK4a/Ki-67 dual labelling was observed in 48 out of 55 (87.3%) HG lesions and in 11 out of 29 (37.9%) negative, papillary urothelial neoplasia of low malignant potential or low-grade carcinomas (P = 0.05). All cases with high-grade/malignant cytology were dual labelled. Sixteen out of 17 (94.1%) carcinoma in situ cases and eight out of 14 (57.1%) cases with atypical urothelial cells matching with HG lesions were dual labelled. Extended follow-up allowed three cases of progression to be diagnosed in dual-labelled cases with negative/low-grade cytology results after a 9- to 11-months delay. Conclusions: The data show that p16INK4a/Ki-67 co-expression allows most HG cancer cells to be detected initially and in the follow-up period. Additional studies are needed in order to determine whether dual labelling can be used as a triage tool for atypical urothelial cells in the urine.

The search for biological and clinical significance of equivocal urinary cytology has emerged as the most promising application of fluorescence in situ hybridization (FISH). Using the multiprobe UroVysion(®) assay, a negative FISH result in the presence of atypical urothelial cells favors the presence of reactive, benign alterations and helps to avoid unnecessary invasive procedures. However, a negative FISH result in case of a negative or equivocal cytology does not exclude low-grade urothelial neoplasia. Equivocal findings are a notorious problem after conservative treatment, particularly after BCG immunotherapy of carcinoma in situ, where even benign reactive changes can appear worrisome. In this situation, a positive FISH result in spite of non-high grade cytology independently indicates persistent or recurrent urothelial carcinoma. However, chromosomal abnormalities are not restricted to malignancy but may also occur in benign cells. Tetraploidy with a balanced duplication of the whole genome, or polyploidy can occur in non-neoplastic conditions of the bladder such as Decoy cells, radiotherapy-induced changes and urolithiasis. Thus, a positive FISH result in a patient with a history of pelvic irradiation does not prove cancer unless there is unequivocal 9p21 deletion. Recent studies show that an aggressive workup of patients with a suspicious cytology+positive UroVysion(®) result and negative cystoscopy is not currently justified. However, multi-target UroVysion(®) FISH remains an excellent tool to improve diagnosis in urinary cytopathology, provided that FISH results are interpreted in the light of the clinical situation, and that one reminds that FISH adds no diagnostic value in case of clearly positive, high-grade cytology.

In recent years, therapeutic approaches including cytoreductive surgery followed by intraperitoneal chemotherapy have proven effective in peritoneal carcinomatosis of colorectal origin. If cytology is to be used to include patients in aggressive treatment regimens, it is necessary to evaluate its performance, particularly in terms of specificity. The aim of this study was to assess interobserver agreement for the detection of intraperitoneal free cancer cells (IFCCs) in patients with non-gynaecological adenocarcinomas.Over a 5-year period, 1223 patients were recruited in 19 French surgery departments. Peritoneal samples were examined in 14 dispersed pathology laboratories. Giemsa-stained slides were sent to a control reader blind to the previous diagnosis. Discordant cases, concordant positive results and a random selection of negative concordant cases were reviewed by a panel of seven cytopathologists. The 'final diagnosis' was that of the consensus meetings but took into account locally-processed slides.Gathering dubious cases with negative results, a 95.6% concordance was achieved between local readers and the control reader. IFCCs were ascertained by the panel in 85 cases (7.0%). Eight of 873 colorectal cancers cases viewed locally were falsely positive (0.9%). Radiotherapy and neoadjuvant therapy had no impact on the false-positive rate as assessed by final validation by the panel (P0.05). Samples initially considered as dubious were reclassified as negative by the panel in 24 of 25 cases (96.0%).The panel consensus allowed reclassification of most dubious/equivocal peritoneal cytology cases, whereas clearcut distinction between benign and malignant cases was correctly achieved in almost all cases.

Recurrence rates after surgery for non-small cell lung cancer (NSCLC) range from 25 to 50% and 5-year survival is only 60-70%. Because no biomarkers are predictive of recurrence or the onset of metastasis, pathological TNM (pTNM) staging is currently the best prognostic factor. Consequently, the preoperative detection of circulating tumour cells (CTCs) might be useful in tailoring therapy. The aim of this study was to characterize morphologically any circulating non-haematological cells (CNHCs) in patients undergoing surgery for NSCLC using the isolation by size of epithelial tumour cell (ISET) method.Of 299 blood samples tested, 250 were from patients with resectable NSCLC and 59 from healthy controls. The presence of CNHCs was assessed blindly and independently by 10 cytopathologists on May-Grünwald-Giemsa stained filters and the cells classified into three groups: (i) malignant cells, (ii) uncertain malignant cells, and (iii) benign cells. We assessed interobserver agreement using Kappa (κ) analysis as the measure of agreement.A total of 123 out of 250 (49%) patients showed CNHCs corresponding to malignant, uncertain malignant and benign cells, in 102/250 (41%), 15/250 (6%) and 6/250 (2%) cases, respectively. No CNHCs were detected in the blood of healthy subjects. Interobserver diagnostic variability was absent for CNHCs, low for malignant cells and limited for uncertain malignant and benign cells.Identification of CTCs in resectable NSCLC patients, using ISET technology and according to cytopathological criteria of malignancy, appears to be a new and promising field of cytopathology with potential relevance to lung oncology.

Purpose: Pathologic TNM staging is currently the best prognostic factor for non-small cell lung carcinoma (NSCLC). However, even in early-stage NSCLC, the recurrence rates after surgery range from 25% to 50%. The preoperative detection of circulating tumor cells (CTC) could be useful to tailor new therapeutic strategies in NSCLC. We assessed the presence of CTC in NSCLC patients undergoing surgery, using cytologic analyses, after their isolation by size of epithelial tumor cells (ISET method). The presence and the number of CTCs were considered and correlated with clinicopathologic parameters including patient follow-up. Experimental Design: Of the 247 blood samples tested, 208 samples were from patients with resectable NSCLC and 39 from healthy subjects. The mean follow-up was 24 months. An image of detected cells with presumably nonhematologic features [initially defined as “circulating nonhematologic cells” (CNHC)] was recorded. The presence of CNHC was assessed blindly and independently by 10 cytopathologists, using cytologic criteria of malignancy on stained filters. The count of detected CNHCs was made for each filter. Results: One hundred two of 208 (49%) patients showed CNHCs corresponding to CNHC with malignant cytopathologic features in 76 of 208 (36%) cases. CNHCs were not detected in the control group. A level of 50 or more CNHCs corresponding to the third quartile was associated with shorter overall and disease-free-survival, independently of disease staging, and with a high risk of recurrence and death in early-stage I + II-resectable NSCLC. Conclusion: A high percentage of NSCLC patients show preoperative detection of CNHC by the ISET method. The presence and level of 50 or more CNHCs are associated with worse survival of patients with resectable NSCLC. Clin Cancer Res; 17(3); 505–13. ©2010 AACR.

Background The aim of our study was to objectively compare Cytyc Thinprep® and other methods of obtaining thin layer cytologic preparations (cytocentrifugation, direct smearing and Millipore® filtration) in urine cytopathology. Methods Thinprep slides were compared to direct smears in 79 cases. Cytocentrifugation carried out with the Thermo Shandon Cytospin® 4 was compared to Thinprep in 106 cases, and comparison with Millipore filtration followed by blotting was obtained in 22 cases. Quality was assessed by scoring cellularity, fixation, red blood cells, leukocytes and nuclear abnormalities. Results The data show that 1) smearing allows good overall results to be obtained, 2) Cytocentrifugation with reusable TPX® chambers should be avoided, 3) Cytocentrifugation using disposable chambers (Cytofunnels® or Megafunnel® chambers) gives excellent results equalling or surpassing Thinprep and 4) Millipore filtration should be avoided, owing to its poor global quality. Despite differences in quality, the techniques studied have no impact on the diagnostic accuracy as evaluated by the rate of abnormalities. Conclusion We conclude that conventional methods such as cytocentrifugation remain the most appropriate ones for current treatment of urinary samples. Cytyc Thinprep processing, owing to its cost, could be used essentially for cytology-based molecular studies.

Objectifs Determiner la valeur pronostique et la valeur predictive pour l’apparition d’une carcinose peritoneale de la cytologie peritoneale dans les adenocarcinomes colorectaux et gastriques. Materiels et Methodes De 2001 a 2007, 1300 patients presentant un cancer d’origine digestive ont ete inclus dans le protocole EVOCAP 2, etude prospective multicentrique (19 centres). Une cytologie peritoneale etait realisee au debut de chaque intervention. La survie globale, la survie sans recidive a deux ans et la survenu d’une carcinose peritoneale ont ete analysees en fonction des resultats de la cytologie. Resultats Les resultats intermediaires sur 367 patients etaient les suivants : la presence de cellules malignes detectee par la cytologie peritoneale est un facteur pronostique pejoratif en termes de survie globale pour les cancers coliques (Hazard ratio = 13,3, p = 0,0018) et en termes de survie sans recidive pour les cancers gastriques (hazard ratio = 8,49, p = 0,0376). L’incidence des cytologies positives est correlee significativement avec le stade pTNM, la presence d’ascite ou de carcinose peritoneale et la realisation d’un traitement neo-adjuvant. Le controle des dernieres cytologies est actuellement en cours, et l’analyse statistique sera realisee en mai ce qui permettra de presenter en octobre les resultats definitifs sur la serie globale de 1300 patients. Conclusion La cytologie peritoneale est un facteur pronostique pejoratif pour les adenocarcinomes coliques et gastriques. La survie globale des patients presentant des cellules malignes sur une cytologie peritoneale est equivalente a ceux presentant un cancer de stade IV. Les resultats definitifs sur la serie de 1300 patients permettront d’evaluer le caractere predictif de la cytologie peritoneale sur la survenue d’une carcinose peritoneale.

Buts du travail 1) Analyser l’expression immunocytochimique (ICC) de l’ACE, du CA 15-3, du CA 19-9 et du CA 125 au niveau des cellules tumorales d’epanchements neoplasiques de primitif connu ou inconnu et 2) etudier la correlation entre l’ICC et les dosages seriques des marqueurs dans la recherche de l’organe d’origine. Methodes Une serie de 79 patients presentant une ascite symptomatique dans 36 cas (45,6 %) et/ou une pleuresie dans 43 cas (54,4 %) avec presence de cellules tumorales dans le liquide d’epanchement dans tous les cas etudies. Parmi ceux-ci, 50/79 (63,3 %) avaient un cancer primitif connu, de type adenocarcinome (ADC) dans 43 cas. Dans 35/79 cas (44,3 %), il existait au moment du bilan initial un envahissement mesothelial certain (greffes ou nodules tumoraux de la sereuse dans le cadre d’une evolution polymetastatique, carcinomatose peritoneale et/ou pleurale). Tous ces cas « vrais positifs » ont ete documentes par l’imagerie et/ou l’anatomie pathologique, que le cancer primitif soit connu ou non. Tous les patients ont eu un bilan initial cytochimique, cytopathologique et ICC (etalements immediatement fixes au methanol/acetone a +4 C puis congeles a -20 C). L’ICC etait completee par l’etude de l’expression de Vimentine, de CK20 et d’HBME-1 au niveau des cellules mesotheliales reactionnelles et des cellules tumorales. Resultats Vingt-quatre cas d’ADC sur 43 (55,8 %) avaient une expression ICC significative de l’ACE et une co-expression du CA 15-3 dans 8 cas, du CA 19-9 dans 9 cas et du CA 125 dans 8 cas. Le profil ICC de l’ensemble des marqueurs permettait une orientation correcte vers le primitif connu dans 88,9 % des ADC mammaires (8 septembre cas plus ou moins compatibles dont 6/8 avec CA 15-3 predominant), 72,7 % des ADC gastro-intestinaux (8/11 cas plus ou moins compatibles dont 4/8 avec ACE et/ou CA 19-9 predominant) et 27,3 % des ADC ovariens (6/11 cas plus ou moins compatibles dont 3/6 avec CA 125 predominant). Les autres types de cancers primitifs avaient des profils d’expression variable. La correlation du dosage serique des marqueurs avec l’ICC montrait que l’ACE etait superieur a la valeur seuil de 5 mg/L dans seulement 18/31 cas (58,1 %), et que le CA 15-3 etait superieur a la valeur seuil de 40 KUI/L dans 23/39 cas (58,9 %). Conclusion La recherche ICC des marqueurs tumoraux semble avoir une valeur diagnostique superieure a celle du dosage serique, d’autant qu’elle ne porte que sur l’expression specifique des cellules tumorales. Les dosages seriques sont probablement affectes d’une sensibilite et d’une specificite faibles, non etudiees dans cette serie preliminaire. L’association de l’ACE et des CA permet une orientation ICC satisfaisante dans la plupart des epanchements neoplasiques, avec un meilleur pouvoir discriminant de l’association ACE/CA 15-3 et ACE/CA 19-9 dans les ADC d’origine mammaire et gastro-intestinale, respectivement. Le marqueur le moins utile semble etre le CA 125, aussi bien au niveau ICC que serique.

To compare the DNA content measured by flow cytometry (FCM) and image analysis (IA) from prostatic fine needle aspiration (FNA) samples.A total of 48 samples were studied. FCM was performed on propidium iodide-stained nuclei according to the Vindelov method, and image analysis was performed on Feulgen-stained slides. Positive FNA results were grade (1-3) and compared with Gleason grades.Aneuploidy was closely related to positive FNA results (P.01). DNA histograms were found to be aneuploid in all grade 3 carcinomas (n = 13) by IA and in 11 cases (84.6%) by FCM. Grade 2 carcinomas (n = 9) were found to be aneuploid with both methods. In grade 1 carcinomas (n = 10), 2 cases exhibited IA aneuploid profiles, whereas all FCM cases were diploid. Aneuploid profiles were more often associated with high Gleason scores than were diploid ones (P.01). Among the 16 patients with negative FNA results, two cases had tetraploid DNA profiles related to contaminating seminal vesicle cells. The difference in DNA measurements reached 10.4% but was not statistically significant.These findings show that the two methods, as applied to prostatic FNA samples, give comparable results and that seminal vesicle cells may be responsible for false tetraploid profiles.

We have induced tumors by feeding guinea pigs with a diet containing 25 or 30% dried bracken fern for 100 or 150 days. A high incidence of bladder tumors was obtained. All but one animal had preneoplastic or neoplastic lesions after 4 months; after one year, 24 or 25 exposed animals had carcinoma. Bladder tumors obtained were essentially pure transitional cell carcinomas, although 4 cases (7% of the exposed animals and 10% of the 39 transitional cell carcinoma observed) showed areas of focal squamous metaplasia. Immunohistological detection of cytokeratins 10, 13, and 18 confirmed the transitional nature of these tumors. Tumor development can be followed by ultrasonography and cytology. Bladder tumors arose through several steps. Dysplasia and preneoplastic hyperplasia were seen after 4 months and papillary carcinomas appeared after 6 months, whereas muscle-invasive carcinomas required 1 year. Thus this model reproduces the full spectrum of preneoplastic and neoplastic bladder lesions observed in humans. Interestingly, when tumors were induced in older guinea pigs, none of them progressed to a muscle-invasive stage. This phenomenon should provide the opportunity to study the molecular mechanisms associated with these two different growth patterns, a major issue in understanding human bladder tumor progression.

We analysed 150 cases of primary lung cancers investigated by bronchial aspirate and biopsy methods with clinical, radiologic and bronchoscopic findings. Among the 150 cases studied, three were characterized by mixed tumor cell components, thus allowing 153 cyto-histological comparisons. The cytologic and histopathologic typing agreed strictly in 102 cases of 153 (66.7%) and was considered as correct in 40 cases (26.1%). Only cases with divergent evaluation between small-cell carcinoma and non small-cell carcinoma were considered as discordant: such misclassification occurred in 11 specimens (7.2%), including two cases with mixed patterns. The cytologic typing was in agreement with the final diagnosis in all epidermoid carcinomas, adenocarcinomas, large-cell carcinomas and poorly differentiated carcinomas. In small-cell carcinomas, cytology was in agreement with histopathology in 20 of 26 cases (77%), and could only indicate undifferentiated features in four cases (15.4%). The analysis of bronchial aspirate specimens gave reliable typing in 92% of cases, and indicated a better tumor cell differentiation than histopathology in 6.5% of cases. The results obtained show that aggressive treatments can be reliably proposed on the basis of cytologic findings, even without tissue corroboration. This proposal is particularly helpful in cases where biopsy cannot be performed (peripheral lesions) or creates a danger to the patient (iatrogenic hemorrhage).

In order to investigate the relationship between genetic instability and DNA aneuploidy of malignant cells in human solid tumors, we studied the variations of DNA index within a single tumor. Multiple sampling (mean of 6.4 samples per tumor) was performed in 24 renal cell carcinomas (RCC). Based on the variations in DNA indices within a single tumor, RCC were divided into three groups: 1) tumors with stable DNA indices (variation within the range of the measurement error), including all DNA diploid tumors (n = 8), all hyperdiploid tumors (n = 3), a hypodiploid tumor (n = 1), and only 2 of 11 tetraploid and hypotetraploid tumors; 2) tumors with moderate variations in DNA indices, which were all close to tetraploidy (n = 5); and 3) tumors with large variations of DNA indices. In this last group, subclones present within a tumor varied widely in their DNA indices from the tetraploid to the triploid region, reflecting the DNA content instability within the tumor cell population. These results suggest that DNA aneuploidy can arise by two different mechanisms: 1) loss or gain of chromosomes leading to hypodiploid or hyperdiploid tumors with no apparent increase in DNA content instability, and 2) doubling of the chromosome set followed by random loss of chromosomes as suggested by the DNA indices ranging from tetraploid to triploid region found in three tumors. Differences in DNA indices within one tumor characterize subclones which may arise by chromosome loss or gain.

The proposal of an early diagnosis of prostate cancer through mass screening with digital rectal examination (DRE), transrectal ultrasound (TRUS) and serum tumor markers remains controversial: there is no high risk population. No study has proven that mass screening reduces the mortality from prostate cancer. However, when clinical and biological data give arguments for the presence of a cancer, every patient requires a prostate biopsy. We have studied the Positive Predictive Value (PPV) of each test in a selected population: 200 men over 50 years of age in which rectal examination or PSA assay was suspicious were investigated. Without any reference to the prostate volume, we considered that the PSA level was "suspicious" when it reached 3 times the upper reference value, or 12 ng/ml. DRE was suspicious in 73%, comprising 50% with prostate carcinoma. PSA assay was suspicious in 65%, comprising 61% with prostate carcinoma. 88% of cancers had a suspicious DRE or PSA assay. TRUS was suspicious in 89%, comprising 45% with prostate carcinoma. Ultrasound guided core biopsies were performed in each case, and allowed a positive diagnosis in 42% of cases, whereas bilateral fine-needle cyto-aspirates were positive in 87% of histologically proven carcinomas. Cytology alone was positive in 3 patients with negative biopsies. Both results show that the PPV of a suspicious DRE associated with an elevated PSA level is 84%. An increased PSA level is correlated with a cancer in 61%.(ABSTRACT TRUNCATED AT 250 WORDS)

The diagnostic sensitivity (Se) and specificity (Sp) of fine-needle aspiration cytology (FNAC) of the prostate can be evaluated by comparing its results to a histological reference: rates of reported Se range from 65-98%, Sp being equal or superior to 95%. Published series are heterogeneous in terms of cancer prevalence, with a 25-85% proportion of histologically proven adenocarcinomas, irrespective of the anatomical stage of the disease. The overall accuracy of screening by core biopsies or FNAC is lower than 5%, and does not justify wide-scale application of these tests. In 75%, cytological assessment of the tumor grade correlates with Gleason's histological score and grade. Severe intraductal dysplasias (DIC 3) are probably involved in some of the cytological grade I cases. Ultrasonographic guidance of FNAC is not recommended in comparison with histologically obtained data. The indications for performing FNAC of the prostate should be different from those of standard biopsies: the former should be carried out on suspicious lesions revealed by digital rectal examination or ultrasonography, or in a staging attempt. FNAC should be reserved for early diagnosis of prostate cancer in patients presenting with non-specific urologic symptoms. Samples should be obtained by digitally-guided transrectal bilateral FNAC.

Cytology is an old method of diagnosis: it is the microscopical observation of cells in the aim of their identification, of the recognition of morphological abnormalities to obtain diagnosis or prognosis informations. During the last 20 years, new technics have been developed: 1) Immunocytochemistry permits the identification of the tissue or the organ of which a tumoral cell population is born, the determination of its stage of differentiation, its kinetic and the diagnosis of bone marrow or lymph node metastasis. 2) With in situ molecular biology, it becomes possible to detect, at the cell level, virus, oncogenes or chromosomic abnormalities. 3) Flow and image cytometry are easy means for DNA content determination and for an objective and reproducible cell definition. None of these methods are able, at this time, to replace the cytological analysis, but they are useful, sometimes necessary, for the diagnosis and the prognosis evaluation of malignancy.

In this prospective study, flow cytometry DNA profile of 169 stage D2 prostatic carcinomas were compared with conventional cytologic data. Two transrectal fine-needle aspiration biopsies were performed in each patient. Aspirates were immediately fixed in 2% polyethylene glycol 1500* alcoholic solution (Merck). Suspensions were mixed and studied by a conventional cytologic technique (Papanicolaou) and by flow cytometry (propidium iodide stain on isolated nuclei). A highly significant correlation was found between the DNA profile and the cytologic grade (p less than 0.001). The DNA profile was bimodal in 14% (3/21) of aspirates containing benign or atypical cells, 18% (3/17) of grade I aspirates, 30% (13/43) of grade II aspirates and 71% (24/34) of grade III aspirates. Routine cytologic evaluation of cell suspensions evaluated by flow cytometry is important in clinical practice since both falsely unimodal and falsely bimodal profiles occur. Leukocytes or benign epithelial cells can interfere with the tumor cell population. In fine-needle aspirates, the broad range of non-malignant contaminating cells limits the value of immunocytochemistry and emphasizes the usefulness of routine conventional cytologic evaluation.

Thirty seven prostatic carcinomas at a D2 metastatic stage were studied after transrectal fine-needle aspiration in a prospective multicentric study. The number and the distribution of nucleoli were compared, in each cytological grade, to the flow cytometric nuclear DNA content. In 9 grade I, 13 grade II and 15 grade III the proportion of bimodal DNA profiles was 11%, 30.7% and 66.7%. The total number of nucleoli per 100 nuclei was respectively 114.11 (SD = 8.12), 158.31 (SD = 47.97) and 194.40 (SD = 58.42). A statistically significant difference (t = 2.78; P = 0.009) was found between the total number of nucleoli of unimodal (22 cases) and bimodal DNA profiles (15 cases). However, no significant difference in nucleolar parameters was found between unimodal (14 cases) and bimodal DNA profiles (14 cases) in cytological grades II and III. These results suggest that the nucleolar parameters do not necessarily parallel the increase in the nuclear DNA content, and that the two factors are independent of one another.

Systematic fine-needle aspiration biopsies and core biopsies were simultaneously obtained on 200 patients with suspected prostatic cancer over a 12-month period. The technical aspect of cellular aspiration and fixation was carefully adjusted. 6 to 8 transrectal bilateral aspirations per patient were performed, and their results were compared to those of core biopsies guided on suspicious areas revealed by rectal examination or transrectal ultrasound. The sensitivity of aspiration for the diagnosis of prostatic cancer is 87%. The specificity is 95%, with a positivity in 3 cases of prostatic cancer in which core biopsy is negative. No cytologic specimen is inadequate for diagnosis. The correlation between cytologic and histologic patterns shows that the proportion of Gleason grade 2 decreases with the cytologic grade, whereas the proportion of Gleason grade 5 increases gradually. Both results show the importance of multiple sampling in the cytological procedure, and confirm the diagnostic value of prostatic fine-needle aspiration method.

During a two-year period, 69 positive prostatic fine-needle cyto-aspirates were selected for a comparison between the cytological grade and the clinical data, and also with the Gleason score and grade obtained with needle-biopsy. The cytological grade is independent of stage, except for grade III that indicates an advanced tumor (T3-T4). On the other hand, the cytological grade provides valuable indications on the histopathological differenciation according to Gleason, with no grade II inferior to Gleason score 5, and no grade III inferior to Gleason 7. 90% of cytological grades III are Gleason grade 4. According to recent studies, we believe that the cytological grade I is related to invasive or in-situ carcinoma, but also to high-grade intra-ductal dysplasia. Thus, it is not surprising to find no correlation between grade I and the clinical status, nor between grade I and the Gleason score and grade, though grade I (or DIC 3) has a good diagnostic value.

BackgroundThe combination of cytology and biopsies improves the recognition and typing of small cell (SCLC) versus non small cell (NSCLC) lung cancers in the fiberoptic bronchoscopy assessment of centrally located tumours.MethodsWe studied whether bronchial aspirates performed before biopsies (BA) and washings performed after biopsies (BW) could increase the diagnostic yield of fiberoptic bronchoscopy. A series of 334 consecutive samples taken in patients with suspicious fiberoptic bronchoscopy findings was studied. Two hundred primary tumours were included in the study. The actual diagnosis was based on surgical tissue specimen analysis and/or imaging techniques. The typing used was that of the 1999 WHO/IASLC classification.ResultsThe diagnosis of malignancy and tumour typing were analyzed according to the sequential (combined) or single use of tests. Malignancy was assessed by cytology in 144/164 (87.8%) positive biopsy cases and in 174/200 tumour cases (87.0%). BA before biopsies allowed 84.0% of cancers to be diagnosed, whereas BW after biopsies allowed 79.0% of cancers to be found (p = ns). However, combining biopsies with BW allowed 94.0% of cancers to be diagnosed, whereas 82.0% were diagnosed by biopsies alone (p < 0.001). The highest diagnostic yield was obtained with the combination of BA, biopsies and BW, with 97.0% sensitivity. Exact concordance in typing was obtained in 83.8% of cases. The six surgically resected cases (3.0%) with negative cytology and biopsy results included four squamous cell carcinomas with necrotizing or fibrous surface and two adenocarcinomas, pT1 stage.ConclusionFiberoptic bronchoscopy may reach a yield of close to 100% in the diagnosis and typing of centrally located, primary lung cancers by combining bronchial aspirates, biopsies and washings.

The diagnostic value of fine-needle aspiration biopsy of the prostate (FNAP) has been evaluated by many authors [1, 2] over the past few years to investigate whether this simple technique could challenge prostatic core biopsy (PCB) and even replace it in the diagnosis of prostatic cancer. These studies have demonstrated that FNAP has a good specificity but a lower sensitivity than PCB, even in series realized under apparently favorable conditions, i. e., with a high rate of prostatic cancer and with clinically palpable tumors. This explains why PCB has, up to now, remained the best clinical diagnostic reference, to which any new technique has to be matched.