Steven A. Seelig

A COMPARISON OF CYTOLOGY AND FLUORESCENCE IN SITU HYBRIDIZATION FOR THE DETECTION OF UROTHELIAL CARCINOMA

PURPOSE We determine the relative sensitivities of cytology and fluorescence in situ hybridization (FISH) for the detection of urothelial carcinoma. MATERIALS AND METHODS A mixture of fluorescent labeled probes to the centromeres of chromosomes 3, 7 and 17, and band 9p21 (P16/CDKN2A gene) was used to assess urinary cells for chromosomal abnormalities indicative of malignancy. A total of 280 urine specimens from 265 patients, including 150 with a history of urothelial carcinoma and 115 without a history of urothelial carcinoma, were analyzed. FISH analysis was performed without prior knowledge of clinical findings, that is biopsy, cystoscopy and cytology results. A positive result was defined as 5 or more urinary cells with gains of 2 or more chromosomes. RESULTS A total of 75 biopsies showed urothelial carcinoma at FISH analysis among the 265 patients. The sensitivity of urine cytology for pTa (36 cases), pTis (18) and pT1-pT4 (15) tumors was 47%, 78% and 60%, respectively, for an overall sensitivity of 58%. The sensitivity of FISH for pTa (37 cases), pTis (17) and pT1-pT4 (19) tumors was 65%, 100% and 95%, respectively, for an overall sensitivity of 81%. FISH was significantly more sensitive than cytology for pTis (p = 0.046), pT1-pT4 (p = 0.025), grade 3 (p = 0.003) and all tumors (p = 0.001). The specificity of cytology and FISH among patients without cystoscopic evidence of urothelial carcinoma and no history of urothelial carcinoma was 98% and 96%, respectively (p = 0.564). CONCLUSIONS The sensitivity of FISH for the detection of urothelial carcinoma is superior to that of cytology, and the specificity of FISH and cytology for urothelial carcinoma are not significantly different. Further prospective studies are required but FISH has the potential to improve significantly the management of urothelial carcinoma.

The development of a multitarget, multicolor fluorescence in situ hybridization assay for the detection of urothelial carcinoma in urine

The purpose of this study was to develop a multitarget, multicolor fluorescence in situ hybridization (FISH) assay for the detection of urothelial carcinoma (UC) in urine specimens. Urinary cells obtained from voided urine specimens of 21 patients with UC and 9 normal donors were analyzed with nine different centromere enumeration probes and a single locus-specific indicator probe to determine an optimal set of FISH probes for UC detection. The four probes with the greatest sensitivity for UC detection were then labeled with a unique fluorophore and combined into a single probe set. The probes with the greatest combined sensitivity for UC detection were CEP3, CEP7, CEP17, and the 9p21 (P16) LSI. This probe set was used to evaluate urine specimens acquired from 179 patients for prospective testing (46 with biopsy-proven UC). FISH slides were evaluated by scanning the slide for cells with nuclear features suggestive of malignancy and assessing the FISH signal pattern of these cells for polysomy (ie, gains of two or more different chromosomes). A receiver operator characteristic curve revealed that a cutoff of 5 cells with polysomy as the positive criterion for cancer resulted in an overall sensitivity of 84.2% for patients with biopsy-proven UC and a specificity of 91.8% among patients with genitourinary disorders but no evidence of UC. This study demonstrates that a multitarget, multicolor FISH assay containing centromeric probes to chromosomes 3, 7, and 17 and a locus-specific probe to band 9p21 has high sensitivity and specificity for the detection of UC in voided urine specimens.

A Comparison of BTA Stat, Hemoglobin Dipstick, Telomerase and Vysis Urovysion Assays for the Detection of Urothelial Carcinoma in Urine

PURPOSE We determine the sensitivity and specificity of various assays for the detection of urothelial carcinoma. MATERIALS AND METHODS A total of 280 voided urine specimens from 265 patients were obtained immediately before cystoscopy for BTA stat, (Bard Diagnostic, Redmond, Washington) hemoglobin dipstick, (Bayer, Elkhart, Indiana) telomerase and UroVysion (Vysis, a wholly owned subsidiary of Abbott Laboratories, Abbott Park, Illinois) analysis. RESULTS Of the 265 patients 75 had biopsy proven urothelial carcinoma, and the sensitivity of the assays was determined from these patients. From most sensitive to least sensitive, the overall sensitivity of UroVysion (73 cases), BTA stat (72), hemoglobin dipstick (73) and telomerase (70) was 81%, 78%, 74%, and 46%, respectively. Each of the first 3 tests was statistically significantly more sensitive than the telomerase assay (p <0.05). However, the differences in overall sensitivity of UroVysion, BTA stat and hemoglobin dipstick were not statistically significant. The specificity of the tests was calculated for 80 of the 265 patients in this study who had no history of urothelial carcinoma and negative cystoscopy findings despite common urological complaints. From most specific to least specific, the specificity of UroVysion, telomerase, BTA stat and hemoglobin dipstick was 96%, 91%, 74% and 51%, respectively. UroVysion and telomerase were statistically significantly (p <0.01) more specific than the BTA stat and hemoglobin dipstick assays, and all of the assays were more specific than hemoglobin dipstick testing (p <0.001). CONCLUSIONS Our study reveals that UroVysion is the most sensitive and specific assay among those tested for the detection of urothelial carcinoma. Telomerase testing had good specificity but poor sensitivity. The BTA stat and hemoglobin dipstick tests had good sensitivity but relatively poor specificity. UroVysion is a promising new assay for the detection of urothelial carcinoma in urine specimens. However, further studies are needed to explore the role of the various assays in the treatment of patients with superficial urothelial carcinoma.

A fluorescence in situ hybridization-based assay for improved detection of lung cancer cells in bronchial washing specimens

Interphase fluorescence in situ hybridization (FISH) is a powerful tool for detecting chromosome and locus‐specific changes in tumor cells. We developed a FISH‐based assay to detect genetic changes in bronchial washing specimens of lung carcinoma patients.

The role of fluorescence in situ hybridization technologies in molecular diagnostics and disease management.

Large genomic changes, such as aneuploidy, deletions, and other chromosomal rearrangements, have long been associated with pregnancy loss, congenital abnormalities, and malignancy. These genomic changes are quantitative, unambiguous, and fundamental in the transition of normal cells to abnormal ones. Detection of these large genetic changes has an increasingly important role in determining patient diagnosis and care, including therapeutic selection. We have developed two major product platforms that assess genomic changes at various levels of resolution. Fluorescence in situ hybridization (FISH) techniques and the related technology of array-based comparative genomic hybridization (CGH) allow detection of genesized or larger alterations in the genome. FISH is a robust DNA probe technology that can measure both balanced and unbalanced genomic changes on a cell-by-cell basis. In most instances, it is not dependent on metaphase chromosomes, and it is widely used in clinical diagnostics. Array-based CGH has much greater multiplexing capabilities than FISH. This technology has the potential to examine many regions of the genome simultaneously for changes in DNA copy number and identify complex patterns of gains and losses within the genome. In this article, we review several of the current medical applications of FISH and discuss such advanced techniques as CGH and array-based CGH.