Multitarget Fluorescence Research Articles

Comparison of the sensitivity and specificity of urine cytology, urinary nuclear matrix protein-22 and multitarget fluorescence in situ hybridization assay in the detection of bladder cancer

This study aimed to compare the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of urine cytology, BladderChek® nuclear matrix protein-22 (NMP22) and UroVysion™ fluorescence in situ hybridization (FISH) tests in patients with newly diagnosed bladder cancer, those with recurrent bladder cancer, and those with bladder cancer but in remission during surveillance. Voided urine samples obtained from 178 patients with suspected or known bladder cancer about to undergo diagnostic or surveillance cystoscopy and 25 control subjects without the disease were divided into four and used for urine culture and cytology, NMP22 BladderChek and UroVysion FISH tests. The sensitivity, specificity, PPV and NPV for each test were calculated. Comparison was made between the ability of each test to detect bladder cancer in the three category of patients listed. Of the 178 patients with bladder cancer, 43 were newly diagnosed, 58 had recurrent disease and 77 were in remission. The sensitivity of each test in newly diagnosed patients was: urine cytology 28%, NMP22 88% and FISH 80%; and in patients with recurrent disease: urine cytology 33%, NMP22 57% and FISH 85%. The mean specificity for urine cytology, NMP22 and FISH was 95%, 67% and 48%, respectively. Of the tests used in the study for detection of bladder cancer, NMP22 appeared to be most cost-effective and rapid, with relatively high sensitivity and specificity in all categories of patients. The NMP22 test may be considered a new gold standard for the assessment of patients with known or suspected bladder cancer.

Voided Urine Fluorescence In Situ Hybridization Testing for Upper Tract Urothelial Carcinoma Surveillance

Fluorescence in situ hybridization is gaining popularity for transitional cell carcinoma screening. We determined the accuracy of fluorescence in situ hybridization for identifying upper tract transitional cell carcinoma. A retrospective review of our upper tract transitional cell carcinoma database from 2005 to 2008 identified 35 patients with upper tract transitional cell carcinoma who submitted voided urine specimens for fluorescence in situ hybridization at commercial laboratory during a routine office visit. Each patient was evaluated endoscopically in the operating room within 3 months of sampling. Suspicious lesions were biopsied and treated. Transitional cell carcinoma in the lower or upper tract was proved by direct visualization, positive biopsy or upper tract cytology read as positive or highly suspicious for malignancy. Of the patients 35 satisfied study inclusion criteria. A total of 67 fluorescence in situ hybridization specimens were submitted. Upper tract transitional cell carcinoma was identified on 51 operative evaluations, of which 23 showed concurrent bladder tumor. For all encounters the sensitivity of fluorescence in situ hybridization was 56% and specificity was 80%. Sensitivity for low and high grade lesions was 68% and 67%, respectively. Only upper tract tumors were noted in 28 patients, in whom there were 2 false-positive and 13 false-negative voided fluorescence in situ hybridization results. In these cases sensitivity was 54% and specificity was 78% compared to the 18% sensitivity and 100% specificity of bladder cytology. Sensitivity for low and high grade upper tract transitional cell carcinoma was 60% and 50%, respectively. Voided fluorescence in situ hybridization has become an adjunct for bladder transitional cell carcinoma surveillance. However, it has limited value for upper tract tumor surveillance.

Detection of chromosomal anomalies in endometrial atypical hyperplasia and carcinoma by using fluorescence in situ hybridization

Endometrial cancer is the most common pelvic gynecological malignancy. The diagnosis of well-differentiated endometrial adenocarcinoma, atypical hyperplasia, and hyperplasia is often challenging. The authors sought to investigate the utility of chromosomal anomalies for the detection of endometrial hyperplasia and carcinoma using multitarget fluorescence in situ hybridization (FISH). Samples were collected by endometrial Tao brush and processed by liquid-based cytological preparation protocol from consecutive cases to include 50 benign, 50 hyperplasia without atypia, 47 atypical hyperplasia, and 53 endometrial cancers. Each was hybridized using fluorescence-labeled DNA probes to chromosomes 1, 8, and 10. The FISH signals were enumerated in 100 cells per case, and the chromosomal anomalies were correlated with pathologic findings, including histologic diagnoses on matched endometrial tissue samples. Numeric chromosomal anomalies were found in 0% (0 of 50) of benign, 20% (10 of 50) of hyperplasia, 74% (35 of 47) of atypical hyperplasia, and 87% (46 of 53) of carcinoma specimens. The mean percentage of cells with chromosomal changes was 55% in cancer specimens, which was significantly higher than that in hyperplasia without atypia (13%, P < .0001) and atypical hyperplasia (32%, P = .003). The most frequent chromosomal anomaly was gain of chromosome 1. FISH anomalies had an overall sensitivity of 81% and specificity of 90% for the detection of atypical hyperplasia and/or endometrial carcinoma. There was no association with grade of endometrial carcinoma. Multitarget FISH appears to be useful for the differential diagnosis of hyperplasia, atypical hyperplasia, and endometrial adenocarcinoma, with a high level of sensitivity and specificity. It is also a potential tool for the early detection of neoplastic cells in endometrial cytology specimens. Endometrial hyperplasia with FISH-detected chromosomal anomalies may represent a clinically significant subset of cases that warrant close clinical follow-up.

Multi-target fluorescence in situ hybridization for prediction of recurrent superficial bladder cancer

Objective To evaluate validity of fluorescence in situ hybridization (FISH) in predicting tumor recurrence in patients with history of superficial bladder cancer. Methods FISH was performed on 20 normal subjects and 43 patients with negative cystoscopy and urinary cytology at the time of FISH assay. The threshold was established based on the assay of normal subjects, and the correlation between FISH and tumor recurrence was analyzed based on the assay of patients. Results After a median follow-up of 24 months,22 patients developed tumor recurrence,including 19 cases (86% ) of abnormal assay and 3 cases (14% ) of normal assay. Twenty-nine patients didn' t suffer from tumor recurrence, including 16 cases (55% ) of abnormal assay,and 13 cases (45% ) of normal assay. Recurrent tumors were developed in 54% of the patients with abnormal assay compared with 19% of the patients with normal assay (P<0.05). Conclusion FISH may be a available tool for predicting tumor recurrence. Key words: Urinary bladder neoplasm; Fluorescence; In situ hybridization; Prognosis

Detection of chromosomal anomalies in uterine endometrial carcinoma using fluorescence in situ hybridization (UteroFISH)

5533 Background: Endometrial cancer is the most common pelvic gynecological malignancy. The diagnosis of well-differentiated endometrial adenocarcinoma, atypical hyperplasia, and marked hyperplasia is often challenging. We sought to investigate the utility of chromosomal anomalies for the detection of uterine endometrial carcinoma using multitarget fluorescence in situ hybridization (FISH). Methods: Samples were collected by endometrial brush and processed by liquid-based thin-layer cytological preparation protocol. For study, we collected cytology slides from consecutive cases to include 50 benign, 50 hyperplasia without atypia, 50 atypical hyperplasia, and 50 endometrial cancers. Each was hybridized using fluorescence labeled DNA probes to chromosomes 1, 8, and 10 (UteroFISH). The FISH signals were enumerated in 100 cells per case, and the chromosomal anomalies were correlated with pathologic findings, including histologic diagnoses on endometrial tissue samples. Results: Numeric chromosomal anomalies were found in 0% (0/50) of benign, 20% (10/50) of hyperplasia, 76% of atypical hyperplasia (38/50), and 86% (43/50) of carcinoma specimens. The mean percentage of cells with chromosomal changes was 54% in cancer specimens, significantly higher than that in hyperplasia without atypia (13%, p&lt; 0.0001) and atypical hyperplasia (34%, p&lt; 0.0001). The most frequent chromosomal anomaly was gain of chromosome 1. FISH anomalies had an overall sensitivity of 81% and specificity of 90% for the detection of atypical hyperplasia and/or endometrial carcinoma. There was no association with grade of endometrial carcinoma. Conclusions: Multi-target UteroFISH appeared to be useful for the differential diagnosis of reactive hyperplasia, atypical hyperplasia, and endometrial adenocarcinoma, with a high level of sensitivity and specificity. Endometrial hyperplasia with FISH-detected chromosomal anomalies may require close clinical follow-up. No significant financial relationships to disclose.

Quantitative Fluorescence in Situ Hybridization and its Ability to Predict Bladder Cancer Recurrence and Progression to Muscle-Invasive Bladder Cancer

Fluorescence in situ hybridization (FISH) testing is used to detect bladder cancer in urine specimens. The purpose of this study was to determine whether there are associations between the percentage of chromosomally abnormal cells by FISH and time to bladder cancer recurrence and progression to metastasis. Clinical records were searched to identify patients with urine FISH results, history of non-invasive bladder cancer, and at least one follow-up pathological diagnosis. Covariates analyzed included age, gender, smoking status, treatment after FISH, cystoscopy result, and prior stage of bladder cancer. The percentage of abnormal cells (hazard ratio [HR] 1.03, 95% CI 1.02-1.03; P < 0.001), age (HR 1.02, 95% CI 1.00-1.03; P = 0.033), male gender (HR 0.60, 95% CI 0.41-0.87; P < 0.001), treatment (HR 0.37, 95% CI 0.25-0.55; P < 0.001), and history of TIS/T1-stage tumors (HR 1.66, 95% CI 1.23-2.24; P = 0.001) were significantly associated with time to cancer recurrence. Time to invasive cancer was significantly associated with the percentage of abnormal cells (HR 1.02, 95% CI 1.01, 1.03; P < 0.001), history of TIS/T1 tumor (HR 3.73, 95% CI 1.88, 7.38; P = 0.001), and treatment (HR 0.33, 95% CI 0.13, 0.83; P = 0.019), suggesting that the percentage of abnormal cells independently predicts cancer recurrence and progression to invasive disease in patients with a history of non-invasive bladder cancer.

Value of multicolour fluorescence in situ hybridisation (UroVysion) in the differential diagnosis of flat urothelial lesions

Aims:During the past 10 years, multitarget fluorescence in situ hybridisation has been established as a valuable adjunct in the cytological diagnosis of precancerous and malignant lesions of the urinary tract....

Reflex UroVysion Testing of Bladder Cancer Surveillance Patients With Equivocal or Negative Urine Cytology

A proportion of patients under surveillance for recurrent bladder carcinoma with no immediate evidence of bladder tumor recurrence have positive multitarget fluorescence in situ hybridization (FISH; UroVysion, Vysis, Downers Grove, IL) results. The course of these "anticipatory positive" cases and the time to bladder tumor recurrence remains unknown. We followed up 250 patients with urine cytologic results, concurrent multitarget FISH, and cystoscopic examination for recurrent urothelial carcinoma. Of 81 cases (32.4%) with FISH-positive results, tumor recurrence developed in 60 (74.0%). Of 169 (67.6%) FISH-negative cases, recurrent urothelial carcinoma developed in 22 (13.0%). Of 211 patients (84.4%) with negative cystoscopic examination results, 56 (26.5%) had positive FISH results, and in 35 (62.5%) of these patients, recurrent urothelial carcinoma developed. Approximately 27% of patients under bladder carcinoma surveillance without immediate evidence of tumor recurrence will have a positive FISH result, defining the anticipatory positive subset. In about 65% of this anticipatory positive group, recurrent bladder urothelial carcinoma developed within 29 months.

High Frequency of Genetic Alterations in Non-small Cell Lung Cancer Detected by Multi-target Fluorescence In Situ Hybridization

Detection of genetic alterations could provide a tool as an adjuvant for the diagnosis of non-small cell lung cancer (NSCLC) and to define patients at risk for early relapse. In this study, a multi-target fluorescence in situ hybridization (FISH) assay was conducted to investigate the correlation between the alterations of chromosomes, including 5p15.2, 6p11.1-q11, 7p12, and 8q24.12-q24.13 (LaVysion Test), and clinicopathological variables, and to clarify the potential of the multi-target FISH assay in 37 NSCLC. The most notable finding was the higher frequency of a gain in chromosome 5p15.2 in early-stage (I+IIa) lung cancers. The frequency of the gain was 81.3% (16/22) in stage I tumors. The frequencies of gains in 6p11.1-q11 and 8q24.12-q24.13 were 61.5% (8/13) and 84.6% (11/13) in stage IIIa cancers, as compared with lower frequencies in stage I tumors at 25.0% and 31.3%, respectively. There was also a significant difference in the histological type. Our results suggest that a gain in 6p11.1-q11 and 8q24.12-q24.13 plays an important role in tumor progression and is associated with histological differentiation. On the other hand, gene amplification in the 5p region was one of the most consistent alterations in early-stage lung cancer, and thus a series of genes in the critical 5p15.2 region might potentially associated with the development of lung cancer.

UroVysion Compared with Cytology and Quantitative Cytology in the Surveillance of Non–Muscle-Invasive Bladder Cancer

ObjectivesThe multitarget fluorescence in situ hybridization probe set Vysis UroVysion, consisting of probes for chromosomes 3, 7, and 17 and for the 9p21 band, was studied to evaluate its value in the follow-up of patients with bladder cancer. The results were compared with conventional cytology and quantitative cytology (Quanticyt). The aim of this study was to evaluate whether UroVysion is a better adjunct to urethrocystoscopy than cytology and quantitative cytology. MethodsUroVysion, cytology, and quantitative cytology were performed on 113 voided urinary samples of 105 patients under surveillance for non–muscle-invasive bladder cancer. Before urethrocystoscopy or transurethral resection of the bladder, a voided urinary sample was obtained. Results of all tests were compared to evaluate the value of UroVysion. ResultsSixty-four patients had biopsy-proven urothelial cell carcinoma. Sensitivity and specificity were, respectively, 39.1% and 89.7% for UroVysion, 40.6% and 89.7% for cytology, and 42.1% and 67.9% for quantitative cytology. When the UroVysion test and cytology were combined, sensitivity increased to 53.1%, but specificity decreased to 79.5%. Detection of Ta tumours was equal for cytology and UroVysion (26.7%), detection of T1 and T2–T4 samples by UroVysion was 60% and 50%, respectively. Detection of grade 1, 2, and 3 tumours by UroVysion was 21.4%, 36.8%, and 66.7%, respectively. In four cases the UroVysion test was positive, but no abnormalities were seen at cystoscopy. ConclusionsOur data suggest that the use of UroVysion provides no improvement over cytology or quantitative cytology in the diagnosis of recurrent non–muscle-invasive bladder tumours.

MP-15.12: UroVysion® compared to cytology and quantitative cytology in the surveillance of superficial bladder cancer

The multitarget fluorescence in situ hybridization (FISH) probe set UroVysion® (Vysis, Abbott Diagnostics, Hoofddorp, The Netherlands) is a urinary test in the diagnosis of superficial bladder cancer and consists of probes to chromosomes 3, 7, 17 and to the 9p21 band. Aim of this study is to evaluate the diagnostic value of UroVysion® in patients under surveillance for superficial bladder cancer and investigate whether it is a better adjunct to cystoscopy compared to cytology (urine) and quantitative cytology (bladder wash).

An International Telecytologic Quiz on Urinary Cytology Reveals Educational Deficits and Absence of a Commonly Used Classification System

Urinary cytology is limited by high interobserver variability in the evaluation of cells with little atypia. We set up an online quiz on urinary cytology and tested the performance of 246 international participants. The quiz consisted of still images of 42 urinary specimens with equivocal morphologic features and 10 control cases with an unequivocal cytologic diagnosis. The nature of the cells on the 292 quiz images had been verified by multitarget fluorescence in situ hybridization in addition to the information obtained by cystoscopy, clinical follow up, and/or histologic examination. The original quiz cases and the percentage of answers given by the participants can be viewed at: http://kathrin.unibas.ch/urinzyto/. High-grade cancers were diagnosed correctly in 76.0% and low-grade cancers in only 33.9%. Remarkably, 54.5% of all participants misclassified decoy cells as malignant. This study shows that large-scale international online quizzes may be used to find educational deficits in cytopathology.

Multitarget Fluorescence In Situ Hybridization Elucidates Equivocal Lung Cytology

The category of equivocal respiratory cytology is a common diagnostic dilemma to both cytopathologists and clinicians. Chromosomal alterations are a hallmark of cancer but are rare or absent in benign conditions. The goal of this study was to test the ability of multitarget fluorescence in situ hybridization (FISH) for dissecting equivocal respiratory cytology into reactive and malignant categories. A consecutive series of 54 Papanicolaou-stained cytologic specimens of the lung was analyzed. The Papanicolaou-stained atypical cell groups were photographed, and the exact locations on the specimens were saved using automated stage and relocation software. The specimens were hybridized with a multitarget FISH probe that contains a mixture of fluorescent probes to the centromeric region of chromosome 6 and to the 5p15, 8q24 (site of the MYC gene) and 7p12 (site of the EGFR gene) loci. The hybridized atypical cells were selectively scored after relocation. A final diagnosis was available in 45 patients, revealing lung carcinoma in 55.5% (n = 25), no evidence of malignancy in 37.8% (n = 17), and pulmonary metastasis of another primary carcinoma in 6.7% (n = 3). FISH results were negative in all 17 patients with benign pulmonary disease and positive in 20 of the 25 patients (80%) with lung carcinoma (p < 0.0001). The sensitivity, specificity, and positive and negative predictive values for detection of malignancy were 79%, 100%, 100%, and 74%, respectively. These data suggest that multitarget FISH in conjunction with automated relocation is a powerful approach for the elucidation of equivocal lung cytology.

Clinical decisions for treatment of different staged bladder cancer based on multitarget fluorescence in situ hybridization assays?

Non-invasive methods for detecting genetic alterations of bladder cancer are increasingly becoming the focus of attention as diagnostic tools. The fluorescence in situ hybridization we performed to detect genetic alterations of chromosomes 3, 7, 9p21, and 17 (UroVysion Test) showed very high sensitivity, higher even than cytology, in detecting bladder tumors of varying differentiation (pTa-pT4). The use of this test in everyday clinical urology can be a very useful decision aid in treating problem cases. A pT1G3 bladder carcinoma in the presence of multichromosomal alterations should be treated as a muscle-invasive pT2 tumor. Other superficial bladder tumors (pTaGI-III, pT1GI-II) with negative histopathology in follow-up and positive FISH analysis with the UroVysion Test should have bladder mapping performed again. Although FISH analysis is currently the most sensitive marker for bladder tumors, the elaborate handling, the cost of the DNA probes and the laboratory equipment required, limit the use of this method in the urologist's everyday routine.

Multitarget FISH and LOH Analyses at Chromosome 3p in Non–Small Cell Lung Cancer and Adjacent Bronchial Epithelium

Multitarget fluorescence in situ hybridization (FISH; LAVysion, Vysis, Downers Grove, IL) targeting chromosomes 6p11-q11, 7p12, 8q24, and 5p15.2 was compared with results of microsatellite studies at chromosome 3p to identify molecular changes associated with tobacco use and tumor development in non-small cell lung cancer (NSCLC). Analyses were performed on 26 NSCLCs and matched benign bronchial epithelium; samples from 10 patients without NSCLC served as control samples. Significant molecular differences between tumor tissue and corresponding benign bronchi were found using FISH (P = .001) and loss of heterozygosity (LOH) analysis (P = .031). Bronchial epithelium from patients with NSCLC was genetically different from epithelium from patients without NSCLC in FISH analysis (P = .025). Receiver operating characteristic curve analysis revealed an optimal cutoff value of 5% atypical cells for bronchial epithelium. There was no statistical correlation with the patient's smoking history, and LOH analysis of bronchi did not differentiate between patients with and without NSCLC. Multicolor FISH analysis is able to detect a tumor-associated molecular field effect in bronchi adjacent to NSCLC.

Multitarget FISH Analysis in the Diagnosis of Lung Cancer

The aim of the present study was to explore the diagnostic usefulness of the multitarget fluorescence in situ hybridization (FISH) test, LAVysion (Vysis, Downers Grove, IL), for the detection of lung cancer cells in cytologic specimens. Specimens from bronchial washings, bronchial brushings, and transbronchial fine-needle aspirates (TBNAs) from 100 patients with suspected lung cancer and from a control group of 71 patients with nonneoplastic lung disorders were analyzed. FISH positivity was defined as more than 5 cells with gains of at least 2 chromosomes or gene loci. FISH significantly improved the sensitivity of bronchial brushings from 49% to 73%. The specificities of FISH and cytologic examination were 87% and 100%, respectively. In contrast, FISH provided no additional diagnostic information in TBNAs and bronchial washings. There was no increased prevalence of genetic changes in contralateral bronchial washings from patients with lung cancer compared with the control group. The quantity of previous smoking had no effect on the prevalence of chromosomal changes.

New molecular markers for bladder cancer detection.

Bladder cancer continues to be one of the most common genitourinary malignancies. The mainstay of diagnosis remains cystoscopic visualization with transurethral biopsy or resection. As over two-thirds of bladder tumors recur, vigilant surveillance is required. Due to the invasiveness and expense of frequent cystoscopies and the lack of sensitivity of urinary cytology, especially for low-grade superficial lesions, novel molecular markers have been investigated as a means to detect bladder cancer noninvasively. As our understanding of the pathogenesis of urothelial neoplasia improves, coupled with recent advances in molecular biological techniques, an array of new approaches to the diagnosis of bladder cancer has emerged. Several urine-based markers have been tested against the standard of urinary cytology with promising results. However, lack of standardization of technique and heterogeneity of bladder cancer itself may hinder the widespread dissemination of these diagnostic aids. A host of new molecular markers based on the pathogenesis of bladder cancer have been investigated, such as telomerase, survivin, and multitarget fluorescence in situ hybridization, which may eventually improve detection and management of urothelial malignancies. By improving the sensitivity of urinary cytology for low-grade superficial lesions and detecting recurrent disease noninvasively early in its course, these new molecular markers might someday allow changes in the way bladder cancer is diagnosed and followed. At the present time, however, no single molecular marker provides 100% accuracy. Perhaps panels utilizing the most promising of these markers may alter bladder cancer detection and management policy.

Comparison of multitarget fluorescence in situ hybridization in urine with other noninvasive tests for detecting bladder cancer: Friedrich MG, Toma MI, Hellstern A, Pantel K, Weisenberger DJ, Noldus J, Huland H, Department of Urology, University Hospital Hamburg-Eppendorf, University of Hamburg, Germany

Comparison of multitarget fluorescence in situ hybridization in urine with other noninvasive tests for detecting bladder cancer: Friedrich MG, Toma MI, Hellstern A, Pantel K, Weisenberger DJ, Noldus J, Huland H, Department of Urology, University Hospital Hamburg-Eppendorf, University of Hamburg, Germany

267: Prospective Analysis of Urine Cytology Ancillary Multitarget Fluorescence In-Situ Hybridization Assay in the Detection of Recurrent Transitional Cell Carcinoma in Patients with Atypical and Negative Cytology

267: Prospective Analysis of Urine Cytology Ancillary Multitarget Fluorescence In-Situ Hybridization Assay in the Detection of Recurrent Transitional Cell Carcinoma in Patients with Atypical and Negative Cytology

Multiprobe FISH for enhanced prediction of recurrent bladder cancer

The early detection of recurrent urothelial cancer is a great challenge for both urologists and pathologists. Cytology is a sensitive and highly specific tool for the diagnosis of high-grade bladder cancer and carcinoma in situ, but is not reliable in low-grade tumors. Therefore, cystoscopy has remained the gold standard for surveillance after resection of bladder cancer. Recent data suggest that multi-target fluorescence in situ hybridization (FISH) can markedly improve the sensitivity of urinary cytology. The goal of this study was to investigate the utility of the recently developed FISH assay Uro-Vysion for the prediction of recurrence during surveillance. 134 bladder washes obtained during a negative follow-up cystoscopy from 127 patients were analyzed. Positive cytology was strongly associated with subsequent recurrence, emphasizing the importance of conventional cytology for bladder cancer surveillance. A positive UroVysion FISH test significantly predicted recurrence when cases with rare tetraploic cells were considered as negative. Taken together, both cytology and FISH help to better determine the risk of bladder cancer recurrence in order to establish more individualized follow-up schemes.