Home About us Editorial board Ahead of print Current issue Search Archives Submit article Instructions Subscribe Contacts Login 


 
 Table of Contents  
ORIGINAL ARTICLE
Year : 2017  |  Volume : 13  |  Issue : 4  |  Page : 647-650

Utility of fluorescence in situ hybridization analysis for detecting upper urinary tract-urothelial carcinoma


Minimally Invasive Urology Center, Shandong Provincial Hospital, Shandong University, Jinan, Shandong, China

Date of Web Publication13-Sep-2017

Correspondence Address:
Jianjun Zhang
Minimally Invasive Urology Center, Shandong Provincial Hospital, 9677 Jingshi East Road, Jinan, Shandong
China
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jcrt.JCRT_74_17

Rights and Permissions
 > Abstract 

Objectives: The objective of this study was to evaluate the clinical utility of fluorescence in situ hybridization (FISH) in the detection of upper urinary tract-urothelial carcinoma (UUT-UC).
Methods: Between November 2011 and November 2015, voided urine specimens from 52 consecutive patients with UUT-UC and 26 controls were collected for both FISH test and cytology. Sensitivity and specificity of FISH test and cytology were determined and compared. The frequency of chromosomal aberrations was also analyzed.
Results: For FISH analysis, the sensitivity was 79.5% and specificity was 96.3%. For cytology, the sensitivity was 27.3% and specificity was 100%. The overall sensitivity for FISH was significantly higher than that of in single value-based urine cytology (79.5% vs. 27.3%, respectively, P < 0.001). The sensitivities of FISH and cytology by grade were 64.3% vs. 28.6% for low-grade urothelial carcinomas (P = 0.128) and 86.7% vs. 26.7% for high-grade urothelial carcinomas (P < 0.001), respectively. Twenty-seven (77.1%) cases were positive due to the gain of two or more chromosomes in five or more urinary cells, among which, 21 (60%) cases showed positivity in all the 4 chromosomes, 7 (20%) cases matched the criterion that 10 or more cells gained a single chromosome, whereas only 1 (2.9%) case was positive because of the homozygous deletion of 9p21 in 12 or more cells.
Conclusions: FISH has superior sensitivity and similar specificity in the detection of UUT-UC, compared with cytology. The present findings indicated that FISH can be applied as a noninvasive diagnostic tool for suspected UUT-UC patients.

Keywords: Cytology, fluorescence in situ hybridization, upper urinary tract-urothelial carcinoma


How to cite this article:
Yu J, Xiong H, Wei C, Cui Z, Jin X, Zhang J. Utility of fluorescence in situ hybridization analysis for detecting upper urinary tract-urothelial carcinoma. J Can Res Ther 2017;13:647-50

How to cite this URL:
Yu J, Xiong H, Wei C, Cui Z, Jin X, Zhang J. Utility of fluorescence in situ hybridization analysis for detecting upper urinary tract-urothelial carcinoma. J Can Res Ther [serial online] 2017 [cited 2018 Nov 16];13:647-50. Available from: http://www.cancerjournal.net/text.asp?2017/13/4/647/214482


 > Introduction Top


Upper urinary tract-urothelial carcinoma (UUT-UC) is a relatively uncommon disease and accounts for only 5% of all the urothelial tumors and 7% to 8% of all renal malignancies.[1] Greater than 90% of these tumors are of transitional cell origin. About 0.7–1.1/100,000 individuals would be affected, with a male-to-female incidence of 1.7:1.[2] More tumors occur in older individuals with a mean age of incidence of 65 years.

UUT-UC is a multifocal disease, and additional tumors may occur in the ureter or bladder or on the contralateral side. Patients with a history of transitional cell cancer (TCC) have a greater risk of developing UUT-UC. Evaluation of UUT-UC should involve imaging techniques, voided urine cytology, cystoscopy, and retrograde pyelography. Computed tomography and magnetic resonance imaging are applied to define the location of lesions and lymph node status. Cytologic analysis of voided urine samples provided a high specificity (>90%) but a low sensitivity (<50%), and it also depends on the skills of cytopathologists.[3],[4],[5],[6] Other marker systems, such as the fibrinogen/fibrin degradation products test, the bladder tumor antigen (BTA) test, and urinary nuclear matrix protein 22, have not achieved an equal or better performance than cytology analysis.[4],[7],[8],[9] Ureteroscopy has been considered the gold standard for the diagnosis and surveillance, but it is an invasive procedure that can be associated with severe complications such as bleeding, ureteral perforation, or stripping of the ureter.

Considering these shortcomings in the diagnosis of UUT-UC, a more reliable and noninvasive method is needed. Recently, the UroVysion fluorescence in situ hybridization (FISH) test has been shown to provide both high sensitivity and specificity in the detection of TCC, which has been approved by the U.S. Food and Drug Administration for use in the initial evaluation of hematuria.[10] FISH test applies fluorescent probes to detect chromosomal aberrations, specifically chromosomes 3, 7, 9, and 17. In this study, the clinical utility of FISH test in the detection of UUT-UC was evaluated.


 > Methods Top


Patients and specimen collection

Between November 2011 and November 2015, 52 consecutive patients suspected of UUT-UC were prospectively selected in this study. These patients underwent evaluation because of gross hematuria, atypical obstruction, and/or radiographic abnormalities. Finally, eight patients were excluded from the study for various reasons, including three patients had ureteral inflammatory stricture, two patients had simultaneous bladder cancer, one patient refused to accept further treatment because of personal reasons, and two patients showed normal signs. Forty-four patients (25 males and 19 females, mean age 72.3 years; range: 55–83 years) were included in this study. Tumor stage and grade were determined after therapeutic surgery; 35 patients underwent laparoscopic nephroureterectomy, two patients underwent endoscopic resection, and seven patients underwent partial ureterectomy. Tumors in the renal pelvis were observed in 13 patients, 29 tumors were observed in the ureter, and tumors in both sites were observed in two patients. The histopathologic classification was performed according to the Union Internationale Contre le cancer criteria. Finally, voided urine samples from 44 patients with UUT-UC (1 pTa-low-grade [LG], 1 pTa-high grade [HG], 5 pT1-LG, 1 pT1-HG, 7 pT2-LG, 17 pT2-HG, 1 pT3-LG, 6 pT3-HG, and 5 pT4-HG) were included and studied. Twenty-six voided urine samples from patients (mean age: 52.3 years; range: 25–71 years, with urinary calculi or benign prostate hyperplasia) with no evidence of UUT-UC were studied as controls. This study was approved by the Ethics Committee of our hospital and informed consent was obtained from all patients and controls before their participation in this study.

One hundred milliliter of voided urine was collected the day before treatment and divided into two aliquots for FISH analysis and cytologic evaluation.

Cytology analysis

Urinary cytology was performed by staining according to the Papanicolaou technique. The samples were evaluated by an experienced pathologist who was blinded to the clinical history of the patients. Results were assessed as positive only in cases with the detection of tumor cells. Suspicious and negative cases were assessed together as negative for the calculations of sensitivity and specificity. Specimens with mild atypia were categorized as negative whereas specimens with moderate-to-severe atypia were categorized as positive.

Fluorescencein situ hybridization analysis

FISH tests were performed following the manufacturer's instructions, as previously described.[11] A test result was defined as positive if it met one of the following criteria: 5 or more morphologically abnormal cells of 25 analyzed cells that had polysomy of 2 or more chromosomes (3, 7, or 17); 10 or more nuclei with the same polysomy in one chromosome (3, 7, or 17); and homozygous deletion of 9p21 in 12 or more cells.

Statistical analysis

To assess the significance of the differences between cytology and FISH analysis, as far as tumor grades and stages were concerned, Fisher's exact test (SAS software, v9.4, SAS Institute, Inc., Cary, NC, USA) was applied to determine the statistical difference. P < 0.05 was considered statistically significant.


 > Results Top


In total, voided urine specimens from 44 patients with UUT-UC and 26 controls were analyzed in this study. In the 44 patients with UUT-UC, the cytology was negative in 32 of the total 44 specimens (72.7%) and positive in 12 specimens (27.3%), whereas FISH obtained positive consequences in 35 specimens (79.5%) and negative in 9 specimens (20.5%). The overall sensitivity of cytology and FISH was 27.3% and 79.5%, respectively [Table 1]. There was no significant difference between renal pelvic UC and ureteric UC for the sensitivity of cytology and FISH [Table 2].
Table 1: Detailed comparison of sensitivity and specificity obtained by cytology and fluorescence in situ hybridization

Click here to view
Table 2: Comparison of sensitivity obtained by cytology and fluorescence in situ hybridization between tumors in renal pelvis and ureter

Click here to view


As far as tumor grades and stages were concerned, sensitivity of cytology and FISH by grade was 28.6% vs. 64.3% for LG (P = 0.128) and 26.7% vs. 86.7% for HG (P < 0.001), respectively. FISH was more sensitive for low- and high-grade lesions than that of in cytology. Sensitivity of cytology and FISH for nonmuscle-invasive tumors (pTa, pT1) was 25% and 12.5%, respectively, whereas for muscle-invasive tumors (pT2, pT3, pT4), it was 27.8% and 94.4%, respectively [Table 1].

Among the 26 controls, there was one false-positive result by FISH in a woman (aged 47 years) with left kidney stone disease and urinary tract infection (UTI). There was no evidence of any UC in this patient by image examinations. Thus, the specificity of cytology and FISH was 100% and 96.2%, respectively [Table 1]. The positive and negative predictive values (NPVs) of cytology for UUT-UC were 100% and 44.8%, respectively, whereas for FISH, they were 97.2% and 74.3%, respectively.

For all 35 positive FISH results, 27 (77.1%) cases were positive due to the gain of two or more chromosomes in five or more urinary cells, among which, 21 (60%) cases showed positivity of all the four chromosomes, seven (20%) cases matched the criterion that 10 or more cells gained a single chromosome, whereas only one (2.9%) case was positive because of the homozygous deletion of 9p21 in 12 or more cells.


 > Discussion Top


Suspected UUT-UCs are usually evaluated with imaging techniques (computed tomography, magnetic resonance imaging, and retrograde pyelography,), cytology, and ureteroscopy with biopsy. Patients suspected of UUT-UC by imaging examinations are usually confirmed by cytology or ureteroscopy with biopsy. However, the role of cytology in the diagnosis of UUT-UC has been controversial. The results of cytology depend on experiences and skills of the cytopathologist. Voided urine cytology has been shown to be of little value in the diagnosis of UUT-UC with a sensitivity of 35% to 50%, especially in low-grade tumors.[5] In this study, the sensitivity of cytology was only 28.6% in low-grade tumors. With ureteral catheterization (washing and brushing), the sensitivity of cytology can be increased from 35% to 88%; however, this procedure was intensive for patients and urologists.[4]

Ureteral endoscopy with biopsy has provided accuracy in grading 71% of low-grade and 80% high-grade urothelial carcinomas before surgery.[3],[12] However, this is an invasive procedure, which can be associated with severe complications such as bleeding, ureteral perforation, extravasation of tumor cells, disruption or stricture formation, or stripping of ureter, especially in proximal urothelial carcinomas of the urinary tract. Moreover, flat or small lesions can be missed in ureteroscopy. Therefore, a more reliable and less invasive method is urgently needed.

A few studies on urine-based markers have been reported for the diagnosis of UUT-UC. ImmunoCyt traces the monoclonal antibodies M344, LDQ10, and 19A211 against transitional cell carcinoma in exfoliated urothelial cells. Lodde et al. and Mian et al. reported the overall sensitivity as 86% to 91%, superior to that of in cytology.[4] However, some benign diseases (e.g., cystitis and benign prostatic hyperplasia) can produce false-positive results. The BTA test detects human complement factor H-related protein in voided urine samples. Walsh et al.[8] reported high sensitivity and specificity of the BTA test for the detection of UUT-UC; however, Siemens et al. reported that no satisfied results could be achieved.[7]

In this study, we focused on the utility of FISH in the diagnosis of UUT-UC. FISH is a cytogenetic-based technology that enables the analysis of multiple chromosomes in several cells. FISH analysis has been initially applied for detecting hematologic cancers and has later being applied for the diagnosis of solid tumors. Marín-Aguilera et al. first reported the utility of FISH as a noninvasive technique in the diagnosis of UUT-UC.[13] The sensitivity of FISH was significantly higher than that of in cytology (76.7% vs. 36%, respectively). In the present study, the obtained sensitivity has been 79.5%, a little higher than the sensitivity reported by Marín-Aguilera et al. This difference might be resulted from more high-grade tumors included in our study. Mian et al. reported a FISH analysis with 100% sensitivity; however, the tumor cohort was very small.[14]

The sensitivities of FISH were 12.5% and 94.4% in nonmuscle-invasive tumors and muscle-invasive tumors, respectively, while the sensitivities were 45.5% and 100%, respectively, in the Marin-Aguilera's study. Two patients with pT2 grade were FISH negative in our study; both patients had severe urinary obstruction caused by the tumor, which might prevent the dissemination of exfoliated tumor cells by urine. The sensitivity of FISH in nonmuscle-invasive tumors was lower than that of in Marin-Aguilera's study. FISH sensitivity in high-grade tumors was 86.7%, significantly higher than the sensitivity of cytology, similar to that of in the studies of Marin-Aguilera et al. (86.4%) and Xu et al. (90.6%).[15] Xu et al. developed a cyto-FISH modality, combining the cytology and FISH results, to detect UUT-UC, which reached an overall sensitivity and specificity of 85.9% and 97.8%, respectively.

The overall sensitivity of cytology was 27.3%, lower than the sensitivity reported by Akkad et al. (60%)[16] and Xu et al. (45.1%). The difference in the cytology results could be interpreted by the different statistical categorizations of atypia of any degree. In this study, specimens with mild atypia were categorized as negative. Furthermore, we truly believe that the results of cytology depend on the skills of cytopathologist.

The specificities of cytology and FISH in this study were 100% and 96.2%, respectively. One false-positive result by FISH was detected from a woman (aged 47 years) with left kidney calculus and UTI. There was no evidence of any UC for this patient by imaging techniques. Frequent follow-up visits were performed for 1 year after percutaneous nephrolithotomy, and no evidence of UC was observed. Marin-Aguilera et al. and Xu et al. have also reported false-positive results by FISH in healthy controls and UTI patients.

The positive predictive value (PPV) in this study was 97.2%, a little higher than the PPV reported by Mian et al., Akkad et al., and Marin-Aguilera et al., which were 83.3%, 87.5%, and 95.8%, respectively. Mian et al. reported that the NPV was 100%, in contrast to our low NPV of 74.3%. It can be interpreted that their tumor cohort was relatively small. Mian et al. believed that they could exclude a UUT-UC in case of FISH negativity and all other tests being negative.[14] However, the same recognition could not be obtained in this study because of the low NPV.

Our sample size has been relatively small compared to that of in FISH studies on bladder TCC. The sensitivity of FISH was generally superior to cytology and in high-grade tumors; however, the sensitivity of FISH in low-grade tumors was disappointed in this study. The longitudinal data on this patient set were continuously collected. As a noninvasive method, FISH is a good adjunct of cytology; however, FISH still cannot replace the utility of ureteroscopy with biopsy in the diagnosis of UUT-UC.


 > Conclusions Top


In this study, we showed that FISH provided superior sensitivity and similar specificity in the detection of UUT-UC, compared to that of in cytology. The present findings indicated that FISH can be used as a noninvasive diagnostic tool for suspected UUT-UC patients.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
 > References Top

1.
Flanigan RC. Urothelial tumors of upper urinary tract. In: Wein AJ, Novick AC, Partin AW, Peters CA, editors. Campbell-Walsh Urology. 9th ed. Philadelphia: Saunders Elsevier; 2007. p. 1638-52.  Back to cited text no. 1
    
2.
Munoz JJ, Ellison LM. Upper tract urothelial neoplasms: Incidence and survival during the last 2 decades. J Urol 2000;164:1523-5.  Back to cited text no. 2
    
3.
Guarnizo E, Pavlovich CP, Seiba M, Carlson DL, Vaughan ED Jr., Sosa RE. Ureteroscopic biopsy of upper tract urothelial carcinoma: Improved diagnostic accuracy and histopathological considerations using a multi-biopsy approach. J Urol 2000;163:52-5.  Back to cited text no. 3
    
4.
Lodde M, Mian C, Wiener H, Haitel A, Pycha A, Marberger M. Detection of upper urinary tract transitional cell carcinoma with ImmunoCyt: A preliminary report. Urology 2001;58:362-6.  Back to cited text no. 4
    
5.
Zincke H, Aguilo JJ, Farrow GM, Utz DC, Khan AU. Significance of urinary cytology in the early detection of transitional cell cancer of the upper urinary tract. J Urol 1976;116:781-3.  Back to cited text no. 5
    
6.
Chen GL, El-Gabry EA, Bagley DH. Surveillance of upper urinary tract transitional cell carcinoma: The role of ureteroscopy, retrograde pyelography, cytology and urinalysis. J Urol 2000;164:1901-4.  Back to cited text no. 6
    
7.
Siemens DR, Morales A, Johnston B, Emerson L. A comparative analysis of rapid urine tests for the diagnosis of upper urinary tract malignancy. Can J Urol 2003;10:1754-8.  Back to cited text no. 7
    
8.
Walsh IK, Keane PF, Ishak LM, Flessland KA. The BTA stat test: A tumor marker for the detection of upper tract transitional cell carcinoma. Urology 2001;58:532-5.  Back to cited text no. 8
    
9.
van Rhijn BW, van der Poel HG, van der Kwast TH. Urine markers for bladder cancer surveillance: A systematic review. Eur Urol 2005;47:736-48.  Back to cited text no. 9
    
10.
Sarosdy MF, Kahn PR, Ziffer MD, Love WR, Barkin J, Abara EO, et al. Use of a multitarget fluorescence in situ hybridization assay to diagnose bladder cancer in patients with hematuria. J Urol 2006;176:44-7.  Back to cited text no. 10
    
11.
Laudadio J, Keane TE, Reeves HM, Savage SJ, Hoda RS, Lage JM, et al. Fluorescence in situ hybridization for detecting transitional cell carcinoma: Implications for clinical practice. BJU Int 2005;96:1280-5.  Back to cited text no. 11
    
12.
El-Hakim A, Weiss GH, Lee BR, Smith AD. Correlation of ureteroscopic appearance with histologic grade of upper tract transitional cell carcinoma. Urology 2004;63:647-50.  Back to cited text no. 12
    
13.
Marín-Aguilera M, Mengual L, Ribal MJ, Musquera M, Ars E, Villavicencio H, et al. Utility of fluorescence in situ hybridization as a non-invasive technique in the diagnosis of upper urinary tract urothelial carcinoma. Eur Urol 2007;51:409-15.  Back to cited text no. 13
    
14.
Mian C, Mazzoleni G, Vikoler S, Martini T, Knüchel-Clark R, Zaak D, et al. Fluorescence in situ hybridisation in the diagnosis of upper urinary tract tumours. Eur Urol 2010;58:288-92.  Back to cited text no. 14
    
15.
Xu C, Zeng Q, Hou J, Gao L, Zhang Z, Xu W, et al. Utility of a modality combining FISH and cytology in upper tract urothelial carcinoma detection in voided urine samples of Chinese patients. Urology 2011;77:636-41.  Back to cited text no. 15
    
16.
Akkad T, Brunner A, Pallwein L, Gozzi C, Bartsch G, Mikuz G, et al. Fluorescence in situ hybridization for detecting upper urinary tract tumors – A preliminary report. Urology 2007;70:753-7.  Back to cited text no. 16
    



 
 
    Tables

  [Table 1], [Table 2]



 

Top
 
 
  Search
 
Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

  >Abstract>Introduction>Methods>Results>Discussion>Conclusions>Article Tables
  In this article
>References

 Article Access Statistics
    Viewed670    
    Printed18    
    Emailed0    
    PDF Downloaded50    
    Comments [Add]    

Recommend this journal


[TAG2]
[TAG3]
[TAG4]