UroVysion Fluorescence Research Articles

UroVysion Fluorescence in situ hybridization (UroVysion FISH) assay for detecting Turkish bladder cancer patients in voided urine: A preliminary study

Bladder cancer is the fourth most common cancer in men and the fifth most common cancer worldwide. UroVysion FISH has high sensitivity and specificity for urothelial carcinoma detection. We investigated the genetic marker detected by the UroVysion FISH technique in diagnosis of Turkish bladder cancer patients and compared these results with the urine cytology and cystoscopy. Urine specimens were analyzed using UroVysion FISH probes for abnormalities in centromeric chromosomes 3, 7, and 17 and locus-specific 9p21.Morning fresh voided urine samples were collected from each patient for FISH analysis. Cytology and histopathology analysis were performed by the pathology department. Twenty-seven bladder cancer patients (23 male and 4 female) with a history of bladder cancer who provided informed consent were included in this prospective study.The results showed that cancer was detected in 8 patients via FISH; 7 via cytology; 12 via cystoscopy. According to the pathology results, 15 were normal, 10 high-grade carcinoma and 2 low-grade carcinoma. Sensitivity of these methods with FISH, cytology, and cystoscopy was 29.6%, 25.9%, and 44.4%, respectively.In conclusion, all tests have different advantages and disadvantages. Also, larger studies will be needed to confirm these results. But, UroVysion FISH appeared to have good specificity for detecting bladder cancer in urine specimens and also it is important to correlate the FISH results with the cystoscopy and cytological findings.

UroVysion Fluorescence In-Situ Hybridization Demonstrates Aneusomy in High Grade Urothelial Carcinomas with a Hypochromatic Cytologic Appearance

UroVysion Fluorescence In-Situ Hybridization Demonstrates Aneusomy in High Grade Urothelial Carcinomas with a Hypochromatic Cytologic Appearance

Automated signal pattern evaluation of a bladder cancer specific multiprobe‐fish assay applying a user‐trainable workstation

Signal pattern enumeration of Urovysion Fluorescence in Situ Hybridization test is tedious and requires great experience. Our aim was to eliminate human interaction by automating the process, using an adoptable, automated image acquisition, and analysis system. For extensive analytical analysis control, cell populations were used, while preliminary clinical study was performed on 21 patients with clinical suspicion for bladder cancer. All investigations were carried out using an automated user-trainable workstation (Metafer4-Metacyte). The system identified nuclei with a specificity and sensitivity of 92.7 and 96.6%, respectively, while signal detection accuracy was 81.1% on average. Both analytical and diagnostic accuracy of automated analysis was comparable to manual approach (94.8 and 71% vs. 97.9 and 76%, respectively), but classification accuracy increased with degree of polysomy, thus diagnostic sensitivity in low grade, low stage cases was poor. It is possible to automate signal enumeration of Urovysion using a user-trainable system, and achieve efficiency comparable to manual analysis. Previously introduced automated immunophenotypic targeting should further increase diagnostic sensitivity, while resulting in a comprehensively automated method. However, the problem of reduced detection accuracy in cases featured with low polysomy is likely to remain a great challenge of automated signal enumeration.

Lymphoepithelioma-like Carcinoma of the Urinary Bladder

Lymphoepithelioma-like carcinoma (LELC) in the urinary tract is a rare malignancy, named for its resemblance to nasopharyngeal undifferentiated carcinoma or lymphoepithelioma. Investigation of immunohistochemical and molecular characteristics of bladder LELC is limited. The pathogenesis and biological behavior of these tumors are controversial. We examined clinicopathologic features of the urinary tract LELC, including light microscopy; immunohistochemistry for cytokeratin 7 (CK7), CK20, 34βE12, p53, p63, α-methylacyl-CoA racemase, thyroid transcription factor-1, Epstein-Barr virus latent membrane protein-1, and CD30; in situ hybridization for human papillomavirus; and UroVysion fluorescence in situ hybridization (FISH). We identified tumors from 34 patients, the largest series to date, (male:female, 2.8:1), ranging from 54 to 84 years of age (mean, 70 years). Urothelial carcinoma in situ was identified in 50% of patients. 34βE12 (75%), CK7 (57%), and p63 (53%) were frequently positive in tumor cells, whereas thyroid transcription factor-1 and CD30 were consistently negative. Expression of p53 was noted in a subset of tumors (61%), whereas CK20 staining was negative with weak positivity in a single case. UroVysion FISH showed frequent chromosomal abnormalities similar to those of urothelial carcinoma. In tumors with concurrent urothelial, squamous, sarcomatoid, and glandular components, identical FISH abnormalities were noted in both areas. In situ hybridization for human papillomavirus and immunostaining for Epstein-Barr virus were negative in all studied lesions. Five patients with pure or predominant LELC tumors treated with transurethral resection and followed by chemotherapy were alive without evidence of disease at 2 to 5 years. In contrast, 2 patients treated in this manner with <50% LELC morphology had death from disease or distant metastasis. Urinary tract LELC is a rare histologic variant of urothelial carcinoma. The frequent presence of UroVysion FISH abnormalities, urothelial carcinoma in situ, and p53 positivity by immunohistochemistry in cases of urinary tract LELC suggests a similar pathogenesis to high-grade invasive urothelial carcinoma. In contrast to typical urothelial carcinoma, CK20 is frequently negative in LELC. Our findings support the hypothesis that pure or predominant LELC may be treated with transurethral resection and chemotherapy. However, a large-scale study with long-term follow-up is needed to better understand the biological behavior of urinary bladder LELC.

Chromosomal abnormalities in renal cell carcinoma variants detected by Urovysion fluorescence in situ hybridization on paraffin-embedded tissue

Urovysion fluorescence in situ hybridization (UVFISH) identifies malignant cells in urine by detecting specific urothelial carcinoma-related chromosomal abnormalities. Some renal carcinomas (RCCs) share overlapping chromosomal aberrations with urothelial carcinoma. Malignant renal cells that are shed in urine can potentially cause a positive UVFISH result. We evaluated UVFISH in RCCs to determine its potential applicability to the diagnosis and grading of RCCs. Paraffin blocks from 39 RCCs (25 clear cell, 9 papillary, 2 chromophobe, and 3 sarcomatoid) and 15 controls (5 renal oncocytomas and 10 urothelial carcinomas) were tested. Of the RCCs, 15 (40%) were UVFISH-positive (9/25 [40%] clear cell, 3/9 [30%] papillary, 1/2 [50%] chromophobe, and 2/3 [67%] sarcomatoid carcinoma) and 24 (60%) were negative. Of the 15 controls, 8 (∼50%) were UVFISH-positive (2/5 [40%] oncocytomas and 6/10 [60%] urothelial carcinomas) and 7 (∼50%) were UVFISH-negative. Polysomy of chromosome 17 showed a statistically significant correlation with RCC subtype, being absent in most of the clear cell RCCs (P = .0096) compared with other RCCs. Polysomy of chromosome 7 was more frequent in high-grade than low-grade RCC (P = .0197) and more likely in high-grade clear cell than low-grade clear cell RCC (P = .0120). In conclusion, we showed that RCC has overlapping chromosomal abnormalities with urothelial carcinoma and can cause a positive UVFISH result. This has implications for the interpretation of Urovysion in patients whose urine contains malignant cells but who have negative cystoscopy and a concomitant renal mass. The chromosomal abnormalities observed in RCC are not distinct from those in urothelial carcinoma; therefore, UVFISH cannot distinguish these tumor types, nor can it type or grade RCC.

Integrating a FISH imaging system into the cytology laboratory

We have implemented an interactive imaging system for the interpretation of UroVysion fluorescence in situ hybridization (FISH) to improve throughput, productivity, quality control and diagnostic accuracy. We describe the Duet imaging system, our experiences with implementation, and outline the financial investment, space requirements, information technology needs, validation, and training of cytotechnologists needed to integrate such a system into a cytology laboratory. Before purchasing the imaging system, we evaluated and validated the instrument at our facility. Implementation required slide preparation changes, IT modifications, development of training programs, and revision of job descriptions for cytotechnologists. A darkened room was built to house the automated scanning station and microscope, as well as two imaging stations. IT changes included generation of storage for archival images on the LAN, addition of external hard drives for back-up, and changes to cable connections for communication between remote locations. Training programs for cytotechnologists, and pathologists/fellows/residents were developed, and cytotechnologists were integrated into multiple steps of the process. The imaging system has resulted in increased productivity for pathologists, concomitant with an expanded role of cytotechnologists in multiple critical steps, including FISH, scan setup, reclassification, and initial interpretation.

Urothelial carcinoma following augmentation cystoplasty: an aggressive variant with distinct clinicopathological characteristics and molecular genetic alterations

Patients who have undergone intestinal augmentation cystoplasty are at risk for developing latent vesicle malignancy. The aim was to evaluate the histological and immunohistochemical characteristics and molecular genetic alterations in these neoplasms. Four patients developing urothelial neoplasms after augmentation cystoplasty were included in the current study. The mean age of the patients, including two men and two women, was 37 years. The latency from bladder augmentation to developing malignancy ranged from 17 to 21 years (mean 19 years). All patients died of cancer shortly after diagnosis (mean 5 months). In the morphological evaluation, all tumours were high-grade (grade 3) invasive urothelial carcinoma comprising various architectural patterns with brisk mitoses and tumour necrosis. Three harboured glandular differentiation and the remaining one showed squamous differentiation. All cases revealed abnormal decreasing beta-catenin expression. Two tumours showed nuclear expression of CDX2. On UroVysion fluorescence in situ hybridization (FISH) analysis, all tumours displayed characteristic chromosomal abnormalities. Point mutations of both FGFR3 and p53 genes were identified in one case. Urothelial carcinomas developed after augmentation cystoplasty are extremely aggressive and exhibit distinct morphological, immunohistochemical and genetic characteristics. UroVysion FISH analysis may offer a surveillance strategy in patients who undergo augmentation cystoplasty.

Detection of Urothelial Bladder Cancer Cells in Voided Urine Can Be Improved by a Combination of Cytology and Standardized Microsatellite Analysis

To evaluate molecular and immunohistochemical markers to develop a molecular grading of urothelial bladder cancer and to test these markers in voided urine samples. 255 consecutive biopsies from primary bladder cancer patients were evaluated on a tissue microarray. The clinical parameters gender, age, adjacent carcinoma in situ, and multifocality were collected. UroVysion fluorescence in situ hybridization (FISH) was done. Expression of cytokeratin 20, MIB1, and TP53 was analyzed by immunohistochemistry. Fibroblast growth factor receptor 3 (FGFR3) status was studied by SNaPshot mutation detection. Results were correlated with clinical outcome by Cox regression analysis. To assess the predictive power of different predictor subsets to detect high grade and tumor invasion, logistic regression models were learned. Additionally, voided urine samples of 119 patients were investigated. After cytologic examination, urine samples were matched with their biopsies and analyzed for loss of heterozygosity (LOH), FGFR3 mutation, polysomy, and p16 deletion using UroVysion FISH. Receiver operator characteristic curves for various predictor subsets were plotted. In biopsies, high grade and solid growth pattern were independent prognostic factors for overall survival. A model consisting of UroVysion FISH and FGFR3 status (FISH + FGFR3) predicted high grade significantly better compared with a recently proposed molecular grade (MIB1 + FGFR3). In voided urine, the combination of cytology with LOH analysis (CYTO + LOH) reached the highest diagnostic accuracy for the detection of bladder cancer cells and performed better than cytology alone (sensitivity of 88.2% and specificity of 97.1%). The combination of cytology with LOH analysis could reduce unpleasant cystoscopies for bladder cancer patients.

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.

Prognostic Significance of Nondiagnostic Molecular Changes in Urine Detected by UroVysion Fluorescence In Situ Hybridization During Surveillance for Bladder Cancer

To determine the outcomes for patients with nondiagnostic fluorescence in situ hybridization (FISH) (ie, < 4 gains of chromosomes 3, 7, or 17 in < or = 3 cells). FISH detects urothelial carcinoma and is especially beneficial in patients with negative or atypical urine cytology findings. A positive result is defined as a gain of > or = 2 chromosomes (3, 7, or 17) in 4 cells, isolated loss of 9p21 in 12 cells, or isolated gains of only 1 chromosome in > or = 10% of cells. Most FISH-positive patients will develop recurrent urothelial carcinoma within 1 year. We compared the data from 149 patients with a nondiagnostic FISH result and > or = 30 months of follow-up with the data from patients with a negative FISH result from the same period. The time to conversion to a positive FISH result or the development of a bladder tumor was recorded. Patients with nondiagnostic FISH results had significantly greater rates of progression to positive FISH findings or the development of a bladder tumor than did patients with negative FISH findings. Most progression occurred within 1 year. Patients with nondiagnostic FISH results and concurrent negative cytology and cystoscopy had a very low risk of developing recurrent disease, similar to that found with truly negative FISH results. Nondiagnostic FISH results are related to a greater risk of progression to positive FISH results and tumor recurrence than those with negative FISH findings. However, after controlling for negative cytologic and cystoscopic status, a nondiagnostic FISH result does not appear to be an independent predictor of disease recurrence, and aggressive investigation is not warranted.

Combined morphologic and fluorescence in situ hybridization analysis of voided urine samples for the detection and follow-up of bladder cancer in patients with benign urine cytology: Daniely M, Rona R, Kaplan T, Olsfanger S, Elboim L, Freiberger A, Lew S, Leibovitch I, BioView Ltd., Rehovot, Israel.

Bladder cancer is among the five most common malignancies worldwide. Patients with bladder cancer are closely followed with periodic cystoscopies and urine cytology analyses due to the significant risk of tumor recurrence. The UroVysion fluorescence in situ hybridization (FISH) test demonstrated higher sensitivity over urine cytology in detecting bladder cancer by most comparative studies. In the current study, the diagnostic usefulness of a combined cytology and FISH analysis approach was tested using the Duet automatic scanning system in patients with benign urine cytology who were being monitored for recurrent urothelial carcinoma or being assessed for various urologic symptoms. By combining the benefits of conventional cytology with molecular diagnostics, a more sensitive detection of bladder cancer was attained. All patients who had positive cystoscopy concomitantly with urine sampling were detected by combined analysis. Additional patients who developed transitional cell carcinoma during a follow-up period of 24 months had a previous positive result on combined analysis. Only 2 patients with a negative combined analysis result presented with late disease recurrence (20 months and 22 months, respectively, after the negative test). Therefore, negative combined analysis was found to be predictive of a lack of disease recurrence for at least 12 months. In this timeframe, the overall sensitivity, specificity, negative predictive value (NPV), and positive predictive values of the combined analysis test were 100%, 65%, 100%, and 44%, respectively. Given the absolute sensitivity and NPV of the combined analysis test, the management of patients with a negative combined analysis result might be revised and allow for more flexible assessment and management of bladder cancer patients relying more on urine bound tests.

Histogenesis of Clear Cell Adenocarcinoma in the Urinary Tract: Evidence of Urothelial Origin

Clear cell adenocarcinoma in the urinary tract is a rare entity with an appearance resembling its counterpart in the female genital tract. Although several theories have been proposed about its origin, its exact histogenesis has remained uncertain. We integrated molecular genetic evaluation by fluorescence in situ hybridization and X-chromosome inactivation with conventional morphologic and immunohistochemical analyses in 12 patients with clear cell adenocarcinomas in the urinary tract. Concurrent urothelial carcinoma or urothelial carcinoma in situ was present in six cases (50%) and foci of cystitis glandularis were observed in four cases (33%). Neither intestinal metaplasia nor Müllerian component was identified in any case. Cytoplasmic expression of alpha-methylacyl-CoA racemase was demonstrable in 10 of 12 tumors (83%). Moderate to diffuse immunostaining for cytokeratin 7 was identified in all 12 tumors (100%), whereas only 3 of 12 (25%) tumors showed positive immunostaining for cytokeratin 20. Focal uroplakin III staining was seen in 6 of 12 tumors (50%). In five cases (42%), focal to moderate CD10 immunoreactivity was observed. Immunostains for OCT4 and CDX2 were completely negative in all tumors. In UroVysion fluorescence in situ hybridization assays, all tumors displayed chromosomal alterations similar to those commonly found in urothelial carcinoma. Identical patterns of nonrandom X-chromosome inactivation in concurrent clear cell adenocarcinoma and urothelial neoplasia were identified in two informative female cases. Our findings support an urothelial origin for most clear cell adenocarcinomas of the urinary tract, despite their morphologic resemblance to certain Müllerian-derived tumors of the female genital tract.

Detection of tumours of the urinary tract in voided urine

Patients with non-muscle-invasive bladder cancer are treated by transurethral resection. About 60-70% of these patients will develop recurrences and in 11% of these cases progression to a muscle-invasive tumour occurs. Surveillance of patients by cystoscopy is therefore carried out every 3-4 months in the first 2 years and yearly thereafter. Several biomarkers have been developed that potentially can detect recurrent bladder cancer in voided urine samples and may present an alternative for the invasive cystoscopy procedure. Recently, van Rhijn reviewed the performance of several of these biomarkers regarding detection of recurrent disease in patients under surveillance. In general, sensitivities were much lower when only patients under surveillance were taken into account than when the patient cohorts included patients with primary disease or patients with high-grade tumours. In this article recent new data on those markers that displayed a sensitivity and specificity of at least 70% as mentioned in the review by van Rhijn are reviewed. The literature selected was limited to those papers in which the performance of makers was assayed only on urine samples of patients under surveillance. The markers with sensitivity and specificity over 70% that were selected from the previous study are Lewis X, NMP22, microsatellite analysis (MA), CYFRA 21.1, cytokeratin 20 and the UroVysion fluorescence in situ hybridization (FISH) test. Recent new developments such as the use of FGFR3 mutation analysis and methylation detection are also discussed. In conclusion, tests such as the UroVysion FISH test and MA are able to detect most concomitant recurrences and to predict recurrent disease. In general, lesions that are missed are pTa and low grade. With MA several upper tract recurrences were identified that were missed by cystoscopy. The value of the most promising urine tests needs to be established in longitudinal studies and exclusively on patients under surveillance for recurrent disease. A longitudinal setting allows subsequent urine samples to be tested and this increases sensitivity because a negative test outcome sometimes occurs between positive ones. Stratification of patients according to the genetic status of their primary tumours and smoking habits should be investigated. Decision models should be developed that recommend at which points in time cystoscopy or urine testing should be performed.

Urothelial Carcinoma With an Inverted Growth Pattern Can be Distinguished From Inverted Papilloma by Fluorescence In Situ Hybridization, Immunohistochemistry, and Morphologic Analysis

Inverted papilloma of the urinary bladder and urothelial carcinoma with an inverted (endophytic) growth pattern may be difficult to distinguish histologically, especially in small biopsies. The distinction is important as these lesions have very different biologic behaviors and are treated differently. We examined histologic features and undertook immunohistochemical staining and UroVysion fluorescence in situ hybridization (FISH) to determine whether these methods could aid in making this distinction. We examined histologic sections from 15 inverted papillomas and 29 urothelial carcinomas with an inverted growth pattern. Each tumor was stained with antibodies to Ki-67, p53, and cytokeratin 20. In addition, each tumor was examined with UroVysion FISH for gains of chromosomes 3, 7, and 17 and for loss of chromosome 9p21 signals. None of the inverted papillomas stained positively for Ki-67 or for cytokeratin 20. Only 1 of 15 inverted papillomas stained positively for p53. By contrast, 66%, 59%, and 59% of urothelial carcinomas with an inverted growth pattern stained positively for Ki-67, p53, and cytokeratin 20, respectively. Only 3 of the urothelial carcinomas stained negatively for all 3 immunohistochemical markers. UroVysion FISH produced normal results for all cases of inverted papilloma. By contrast, 21 of 29 cases (72%) of urothelial carcinoma with an inverted growth pattern demonstrated chromosomal abnormalities typical of urothelial cancer and were considered positive by UroVysion FISH criteria. Morphologic features, as well as immunohistochemical stains (including stains for Ki-67, p53, and cytokeratin 20) and/or UroVysion FISH can help to distinguish inverted papilloma from urothelial carcinoma with an inverted growth pattern.

Combined morphologic and fluorescence in situ hybridization analysis of voided urine samples for the detection and follow-up of bladder cancer in patients with benign urine cytology

Bladder cancer is among the 5 most common malignancies worldwide. Patients with bladder cancer are closely followed with periodic cystoscopies and urine cytology analyses due to the significant risk of tumor recurrence. The UroVysion fluorescence in situ hybridization (FISH) test demonstrated higher sensitivity over urine cytology in detecting bladder cancer by most comparative studies. In the current study, the diagnostic usefulness of a combined cytology and FISH analysis approach was tested using the Duet automatic scanning system in patients with benign urine cytology who were being monitored for recurrent urothelial carcinoma or being assessed for various urologic symptoms. By combining the benefits of conventional cytology with molecular diagnostics, a more sensitive detection of bladder cancer was attained. All patients who had positive cystoscopy concomitantly with urine sampling were detected by combined analysis. Additional patients that developed transitional cell carcinoma during a follow-up period of 24 months had a previous positive result on combined analysis. Only 2 patients with a negative combined analysis result presented with late disease recurrence (20 months and 22 months, respectively, after the negative test). Therefore, negative combined analysis was found to be predictive of a lack of disease recurrence for at least 12 months. In this timeframe, the overall sensitivity, specificity, negative predictive value (NPV), and positive predictive values of the combined analysis test were 100%, 65%, 100%, and 44%, respectively. Given the absolute sensitivity and NPV of the combined analysis test, the management of patients with a negative combined analysis result might be revised and allow for more flexible assessment and management of bladder cancer patients relying more on urine bound tests.

POD-09.04: A comparison of specificity of urine cytology, urinary nuclear matrix protein 22 [NMP22] and multi-target fluorescent in situ hybridization [FISH] assay in bladder cancer patients with no recurrence at surveillance

The UroVysion fluorescence in situ hybridization assay[FISH][Vysis Inc, Illinois, USA] is a multi-target assay that detects aneuploidy of chromosomes 3, 7 and 17 and loss of the 9p21 band in exfoliated cells in urine from patients with transitional cell carcinoma[TCC]. The nuclear matrix protein 22[NMP22] detects elevated amounts of nuclear mitotic apparatus protein, a component of the nuclear matrix essential for cell division that is released into urine at cell death. Both tests have been reported to have higher sensitivity and specificity than urine cytology in patients with cystoscopic and histologic TCC either at initial diagnosis or recurrence.

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....

Use of a Multitarget Fluorescence In Situ Hybridization Assay to Diagnose Bladder Cancer in Patients With Hematuria

We evaluated the multitarget UroVysion fluorescence in situ hybridization assay for the diagnosis of bladder cancer in patients with hematuria and no history of bladder cancer. A multicenter, blinded trial was performed to compare the sensitivity of the fluorescence in situ hybridization assay to that of voided cytology in patients with gross or microscopic hematuria. Confirmation of hematuria was required. Voided urine was sent to a central laboratory for each study before cystoscopy. Suspicious lesions on cystoscopy were biopsied or resected. A centrally reviewed histopathological interpretation was used to confirm cancer and assign grade and stage. A total of 497 patients were enrolled at 23 centers and in 473 (95.2%) fluorescence in situ hybridization and cytology results were interpretable. Bladder cancer was diagnosed histologically in 50 patients (10.1%) and ureteral cancer was diagnosed in 1. Fluorescence in situ hybridization assay detected 69% of cases and cytology detected 38% (95% CI 25 to 52). When low grade, low stage (TaG1) tumors were excluded, fluorescence in situ hybridization detected 25 of 30 cancers (84%), while cytology detected only 15 (50%). Of 265 current or past smokers with hematuria and positive fluorescence in situ hybridization assay findings bladder cancer was detected in 65% with a history of greater than 40 pack-years compared to 13.6% to 24.2% in those with no, less than a 20 or a 20 to 40-pack-year smoking history. The UroVysion fluorescence in situ hybridization assay is significantly more sensitive than voided cytology for detecting bladder cancer in patients evaluated for gross or microscopic hematuria for all grades and stages. Based on these data UroVysion was approved by the Food and Drug Administration for use in patients with hematuria.

The UroVysion fluorescence in situ hybridization assay is an effective tool for monitoring recurrence of bladder cancer

The newly developed UroVysion fluorescence in situ hybridization (FISH) probe was applied to urine specimens from 19 patients being monitored for recurrence of bladder cancer. The results for the multi-target DNA FISH assay were compared with independent analyses of urine cytology and flexible cystoscopy. Patients with tumors identified through the cystoscopy exam were biopsied and/or underwent surgery. In 12 patients with normal cytoscopy, cytology and FISH were also normal. Therefore, the specificity of these two tests was 100%. In 7 patients, a tumor was diagnosed by cystoscopy, and 3 of them had abnormal urine cytology while 6 of them had an abnormal result in the FISH assay. Accordingly, the sensitivity was 43% for the cytology and 87% for the FISH test. Interestingly, a pT1G3 tumor in a bladder diverticulum was not detected by cytology or the FISH test. These results agreed with a large series previously published using similar FISH probes and support the proposal for a multicenter trial to confirm the usefulness of the UroVysion probe as a screening tool to select patients for cystoscopy.

Clinical Evaluation of a Multi-target Fluorescent in Situ Hybridization Assay for Detection of Bladder Cancer

The UroVysion fluorescence in situ hybridization assay (UroVysion Bladder Cancer Recurrence Kit, Vysis, Inc., Downers Grove, Illinois) is a multi-target assay that detects aneuploidy of chromosomes 3, 7 and 17, and loss of the 9p21 band in exfoliated cells in urine from patients with transitional cell carcinoma. We performed 2 multicenter trials. In 1 trial we compared the sensitivity of the FISH assay to the BTA Stat test (Bion Scientific, Redmond, Washington) and voided cytology in the detection of transitional cell carcinoma. In a separate study of healthy volunteers and patients with other (nontransitional cell carcinoma) conditions we determined the specificity of the FISH assay. A total of 176 patients with transitional cell carcinoma in the previous 9 months provided voided urine before cystoscopy. Each specimen was split, preserved and shipped to a central laboratory where all 3 tests were performed. All sites were blinded to results. Sensitivity calculations were based on central pathology review of resected tissue. Specificity was determined by testing 275 volunteers who were healthy and with nontransitional cell carcinoma conditions. The 21 sites enrolled 176 patients with a history of transitional cell carcinoma, with 62 recurrences while undergoing surveillance. Overall sensitivities (with 95% CI) were FISH 71% (95% CI 58 to 82), BTA Stat test 50% (37 to 63) and cytology 26% (16 to 39). FISH was negative in 260 of the 275 healthy volunteers or patients with no history of transitional cell carcinoma (specificity 94.5%). Sensitivity of the FISH assay is superior to that of cytology and at least equivalent to the BTA Stat test in detecting recurrent transitional cell carcinoma. Its specificity approaches that of cytology. Further testing of its clinical use is warranted.