Critical review of biomarkers for the early detection and surveillance of bladder cancer

Review

Keywords Cystoscopy Detection Biomarkers Prediction Diagnosis Urothelial carcinoma Bladder neoplasms

Critical review of biomarkers for the early detection and surveillance of bladder cancer Daher C. Chade, Shahrokh F. Shariat, Guilherme Godoy, Siegfried Meryn and Guido Dalbagni Abstract

Daher C. Chade, MD, PhD Urology Service, Memorial Sloan-Kettering Cancer Center, New York, New York, USA Shahrokh F. Shariat, MD, PhD Division of Urology, Sidney Kimmel Center for Prostate and Urologic Cancer, Memorial Sloan-Kettering Cancer Center, York Avenue, New York, NY, USA Guilherme Godoy, MD Urology Service, Memorial Sloan-Kettering Cancer Center, New York, New York, USA Siegfried Meryn, MD Medical University of Vienna, Vienna, Austria Guido Dalbagni, MD Urology Service, Memorial Sloan-Kettering Cancer Center, New York, New York, USA E-mail: [email protected] Online 7 October 2009

368

Increasing interest in the early diagnosis of urothelial carcinoma of the bladder (UCB) has led to a better understanding of bladder carcinogenesis and an explosion of new biomarkers for this disease. Current surveillance protocols after initial diagnosis include serial cystoscopy, which is invasive and expensive, and cytology, which has a low sensitivity and suffers from high variability. To date, the US Food and Drug Administration (FDA) has approved six urine-based biomarkers to complement cystoscopy in the monitoring of UCB patients. In addition, various promising tests are under investigation. In this review, we describe the rationale and address the most recent and relevant findings for the FDAapproved biomarkers (bladder tumor antigen (BTA) test, BTA stat, BTA TRAK, ImmunoCyt, NMP22, and UroVysion) and the most promising investigational biomarkers (urinary UCB test, BLCA-1, BLCA-4, hyaluronic acid, hyaluronidase, Lewis X antigen, microsatellite analysis, Quanticyt, soluble Fas, survivin, telomerase, and cytokeratin 20). Most of the comparative studies have shown that noninvasive biomarkers have equal or higher sensitivity for UCB detection than cytology, even in high-grade cancers. None of these tests, however, meets all of the criteria of an ideal tumor biomarker. For the investigational biomarkers, improved standardization and automation are still required, as well as prospective, large-scale assessment in heterogeneous patient populations. Moreover, despite the use of urine biomarkers in a variety of clinical situations, their role is not well defined at this time. Identifying an optimal marker that would replace, delay, or complement cystoscopy and/or cytology in the monitoring of patients with UCB is still ongoing. Urinary biomarkers may eventually be used to screen patients at high-risk, to help diagnose or even predict disease recurrence, and to decrease the need for invasive procedures. ß 2009 WPMH GmbH. Published by Elsevier Ireland Ltd.

Introduction Early detection of urothelial carcinoma of the urinary bladder (UCB) still represents a challenge to clinicians, considering that disease progression results in significant morbidity and mortality [1]. Most patients with UCB are diagnosed after they present with gross or microscopic hematuria. At initial diagnosis, about 30% of patients have muscle-invasive cancer. Of this population, 50% have distant

Vol. 6, No. 4, pp. 368–382, December 2009

metastasis within 2 years, and 60% die within 5 years despite treatment [2–4]. Approximately 70% of patients have cancers confined to the epithelium or subepithelial connective tissue at initial diagnosis. These cancers are usually managed with endoscopic resection and selective use of intravesical therapy. The recurrence rate for these tumors ranges from 50–70%, and 10–15% progress to muscle invasion over a 5year period [5,6]. Recurrence may be seen locally or in the upper urinary tract even after

ß 2009 WPMH GmbH. Published by Elsevier Ireland Ltd.

Review several years, requiring life-long surveillance. Diagnosis and surveillance of UCB consists of cystoscopy aided by cytology. Current followup protocols after initial presentation typically include flexible cystoscopy and urine cytology every 3 months for 1–3 years, every 6 months for an additional 2–3 years, and then annually, assuming no disease recurrence. Cystoscopy, a relatively short, minimally traumatic office procedure performed with local urethral anesthesia, identifies most papillary and sessile lesions. Nevertheless, it is still (minimally) invasive and causes discomfort and distress to patients. In addition, although considered the gold standard for diagnosis, cystoscopy may be inconclusive, falsely positive due to a grossly abnormal bladder mucosa (especially in patients with an indwelling catheter or active inflammation), or even falsely negative due to operator error or because small areas of tumor or carcinoma in situ are difficult to accurately recognize [7,8]. Although new cystoscopic technologies such as fluorescence or narrow-band imaging are emerging, with potentially increased accuracy in detecting tumor lesions, the invasiveness and cost of the procedures underscores the need for better urinary biomarkers in the management of patients with UCB. Urine cytology has a reasonable sensitivity and specificity for the detection of high-grade UCB, but it has a weak sensitivity for detecting low-grade tumors, ranging from 4–31% (median of 12%) [9]. The performance of urine cytology in predicting cancer recurrence may vary widely among institutions [10]. In a recent multi-institutional study that included 12 centers from nine countries, urine cytology was positive in 38–65% of patients with recurrent UCB [10]. Sensitivity for grade 3 recurrence was 33–95%, while sensitivity for tumor stage T2 and higher was 37–100%. Other disadvantages are that results are not available immediately and evaluation requires a highly trained cytopathologist, who may not be available in all areas. Moreover, cytology is expensive, difficult to standardize, and takes approximately 1 week to report. Unfortunately, because cytology misses up to 60% of high-grade tumors, it cannot replace or delay cystoscopy. An ideal bladder tumor marker would accurately monitor patients with a history of UCB, identify recurrence early, and prevent disease progression. Because of the relatively low pre-

valence of UCB in general, screening the whole population would probably not be cost-effective, although potentially feasible if a very low cost marker becomes available [11–13]. An accurate marker would also have the potential to replace, delay, or complement cystoscopy and/or cytology in the monitoring of patients with UCB. An ideal UCB monitoring test would be noninvasive, objective, easy to perform and interpret, with high sensitivity and specificity, and would provide an immediate or rapid result. While most of the urine-based tests remain investigational and are undergoing preclinical evaluation, several have already undergone clinical trials and have been approved for clinical use (Table 1).

FDA-approved UCB Biomarkers Bladder tumor antigen (BTA) tests The term BTA describes at least three distinct tests. The original BTA (Bard Diagnostic Sciences, Inc., Redmond, WA) had lower specificity than, and equivalent sensitivity to, urine cytology and was therefore taken off the market [14]. Subsequently, the BTA stat and BTA TRAK (Polymedco Inc., Cortlandt Manor, NY) tests were introduced. The BTA stat is a qualitative point-of-care test with an immediate result, whereas BTA TRAK is a quantitative test that requires trained personnel and a reference laboratory. These assays detect human complement factor H-related protein (as well as complement factor H), which is present in the urine of patients with UCB [15]. It is believed that complement factor H production by tumor cells may prevent tumor cell lysis by immune cells. The overall sensitivity and specificity for the BTA stat test are 57–83% [16–20] and 60–92% [17,21,22], respectively. The reported specificity, however, must be assessed critically. Many of the studies excluded patients who had other commonly occurring genitourinary problems, such as renal stones, infection, and hematuria. In healthy persons without genitourinary signs or symptoms, the specificity is 97%, but in patients with benign genitourinary conditions, the specificity is only 46% [22]. When patients without UCB present with hematuria for other causes, the blood in the urine also contains complement factor

Vol. 6, No. 4, pp. 368–382, December 2009

369

Review Table 1 Summary of sensitivity and specificity of FDA-approved and investigational tests Urinary Tests

Sensitivity (%)

Specificity (%)

References

FDA-approved BTA stat BTA TRAKa ImmunoCyt NMP22 test kitc NMP BladderChek UroVysion

57–83 51–91 50–100 47–100 50–90 74–100

60–92 24b–97 69–79 60–90 85–87 65–86d

[16–22] [27–34] [37–39] [16,18–20,30,46–52] [58,59] [63–66]

Investigational UBC BLCA–1 BLCA–4 Hyaluronic acid HA–HAase Lewis X antigen Microsatellite analysis Quanticyt Soluble Fas Survivin Telomerase Cytokeratin 20

12–66 80 89–96 92 91 80–94 58–97 45–69 75–100 64 7–100 85

90–97 87 95 93 70 83 73–100 70–93 NAe 93 24–90 94

[33,75] [79] [81–82] [85] [89] [92–94] [95–99] [16,100,103,105] [105] [109] [119–132] [133]

a b c d e

Cutoff value varied between 14 and 17.1 U/ml. Set sensitivity at 90%. Cutoff value varied between 3.6 and 13.7 U/ml. Include Duet scanning system and reflex FISH. Areas under the ROC curve for sFas = 0.757 (95% CI = 0.694–0.819).

H, which can react with the antibody in the test and lead to a false-positive result [23,24]. In a recent prospective, multicenter trial of over 500 patients, the reported sensitivity of BTA stat to monitor for UCB recurrence was greater than that of cytology, particularly in grade 1 lesions (47.9% vs 12.5%) [25]. However, prior intravesical treatment, benign prostatic hyperplasia, kidney stones, and urinary tract infections caused a high false-positive rate, and the BTA stat test by itself would have missed 46.6% of tumors detected on cystoscopy. The BTA TRAK test is a quantitative sandwich immunoassay that is performed in a reference laboratory [26]. The cutoff limit of human complement factor H-related protein to detect UCB is 14 U/ml as recommended by the manufacturer [27]. When this cutoff is used, the reported overall sensitivity is 62–91% [27–34]. However, with a set sensitivity of 90%, the specificity of BTA TRAK was only 24.8% [33]. As with the BTA stat test, benign genitourinary conditions, particularly noncancer-related hematuria, may yield false-positive results [27,28,30].

370

Vol. 6, No. 4, pp. 368–382, December 2009

Both tests have sensitivities comparable to that of cytology for high-grade tumors and better than cytology for low-grade tumors. These tests are approved by the US Food and Drug Administration (FDA) only in combination with cystoscopy for monitoring of UCB. Because of their high false-positive rate, they are not sufficiently accurate to be used for screening, early detection, or surveillance without cystoscopy of urothelial carcinoma, particularly in patients with other genitourinary symptoms.

ImmunoCyt ImmunoCyt (DiagnoCure Inc., Quebec) combines cytology with an immunofluorescence assay (immunocytochemistry) [35]. ImmunoCyt detects cellular biomarkers for UCB in exfoliated urothelial cells using fluorescent monoclonal antibodies to pinpoint a high molecular weight form of carcinoembryonic antigen and two bladder tumor cell-associated mucins. Because the test requires the use of a fluorescent microscope by trained personnel,

Review it is performed in a reference laboratory. Other disadvantages are the large number of exfoliated cells necessary to perform an accurate test and its high cost [36]. ImmunoCyt has an overall sensitivity of 50–100% [37–39]. Its specificity has been reported as 69–79%, with a higher false-positive rate in patients with benign prostatic hyperplasia or cystitis [37– 39]. Its advantage may be an improved sensitivity when compared to cytology, especially in low-grade tumors; but this appears to come at the cost of reduced specificity and reduced positive predictive values (PPV) [40,41]. A recent study found that ImmunoCyt has a sensitivity of 76% and a PPV of 43% [42]. The test may be useful as an adjunct to cystoscopy and should be used only for monitoring patients with UCB.

Nuclear matrix protein 22 tests The nuclear matrix protein 22 (NMP22) test detects a nuclear mitotic apparatus protein that is a component of the nuclear matrix. NMPs make up the framework of a cell’s nucleus and play an important role in gene expression [43–45]. NMP22 is a protein that localizes to the spindle poles during mitosis and thus regulates chromatid and daughter cell separation [46,47]. There is a substantially higher level of NMP22 in the urine of patients with bladder cancer. However, because this protein is released from dead and dying urothelial cells, many non-malignant conditions of the urinary tract, such as stones, infection, inflammation, and hematuria, as well as instrumentation (i.e., cytoscopy), can cause a false-positive test result. Both a laboratorybased quantitative microplate enzyme immunoassay and a qualitative point-of-care test are FDA-approved for use in bladder cancer surveillance. The latter is also approved for detection of bladder cancer in high-risk patients. The sensitivity of the original NMP22 immunoassay has ranged from 47–100% and its specificity from 60–90% depending on the cutoff value used [16,18–20,22,28,30,48–52]. The specificity may be significantly decreased in the presence of benign inflammatory or infectious conditions, renal or bladder calculi, a foreign body (stent, catheter), bowel interposition, other genitourinary cancer, or instrumentation [18]. When patients with these problems were excluded, the specificity increased to 99% [53]. Of interest, intravesical

bacillus Calmette-Gue´rin does not alter the performance characteristics of NMP22 [54]. Despite these promising data, the quantitative enzyme NMP22 immunoassay has not been widely used due to a high false-positive rate and the unclear clinical role of the marker. Also, the ideal cutoff point remains undetermined, with a recommended value of 10 U/ml or greater as a positive test result, but with studies suggesting threshold values between 3.6 and 13.7 U/ml [55,56]. Nomograms have incorporated urinary NMP22 levels, associated with urinary cytology, patient age, and gender (Fig. 1) [57]. Urinary levels of NMP22 improved the ability to predict UCB recurrence and progression by a statistically and clinically significant margin. However, there is a significant heterogeneity in performance characteristics of NMP22 that may affect its predictive accuracy when evaluating cancer recurrence and progression in different populations [55]. More recently, the point-of-care test NMP22 BladderChek was introduced. A multi-institutional trial revealed that the addition of the NMP22 BladderChek test to cystoscopy improved the detection rate of UCB in patients with risk factors for UCB [58]. The NMP22 BladderChek test sensitivities were 50% and 90% for noninvasive and invasive cancer, respectively, with an overall sensitivity of 55.7%. In contrast, cytology performed poorly, with comparable sensitivities of 16.7% and 22.2% in noninvasive and invasive UCBs, respectively, with an overall sensitivity of 15.8%. Overall specificity was still higher for cytology at 99.2% compared with 85.7% for NMP22 BladderChek. A subsequent study of 668 patients with a history of UCB found that NMP22 BladderChek detected 8 of 9 cancers not diagnosed by initial cystoscopy [59]. Urine cytology only detected 3 of the cancers missed by cystoscopy. The study has been criticized for the low sensitivity of cytology (12.2%), and questions regarding the impact on the therapeutic decision-making process remain open. A study that tried to address this issue through a risk-based analysis of the data reported that NMP22 had a negative predictive value (NPV) of 96.9% overall [60]. While the overall PPV was 20.3%, it increased in men with high-risk factors (smokers, gross hematuria, and older patients). For instance, the PPV

Vol. 6, No. 4, pp. 368–382, December 2009

371

Review

Figure 1

372

Nomograms for detection of bladder urothelial carcinoma recurrence after treatment of stage Ta, T1, and/or carcinoma in situ (CIS) urothelial carcinoma of the urinary bladder in 2681 patients who underwent office cystoscopy. (A) recurrence of any transitional cell carcinoma; (B) recurrence of grade 3 Ta or T1, or of CIS; (C) recurrence of T2 or higher stage urothelial carcinoma. Reprinted with permission from Shariat et al [57].

Vol. 6, No. 4, pp. 368–382, December 2009

Review in male smokers aged >65 years with gross hematuria (who had a cancer incidence of 46.2%) was 77.8%. The same group developed a predictive tool based on the data from 670 patients [61]. First, they showed that NMP22 BladderChek was superior to urinary cytology (accuracy: 76.0% vs 56.2%). Split-sample logistic regression analysis using an external validation cohort of 602 patients next showed that their NMP22-based predictive tool had an accuracy of 82.4%, which was significantly more accurate than a cytology-based nomogram (74.7%, P = 0.006). Adding cytology to NMP22 did not improve the predictive accuracy of the model (82.6%, P = 0.1).

UroVysion UroVysion (Abbott Molecular, Inc., Des Plaines, IL) is a multitarget fluorescence in situ hybridization (FISH) assay that detects aneuploidy in chromosomes 3, 7, and 17 as well as loss of the 9p21 locus of the P16 tumor suppressor gene

Figure 2

(Fig. 2) [62]. This test has been approved by the FDA both for monitoring patients with a history of UCB and for detection in patients with hematuria. The FISH test combines assessment of the morphologic changes of conventional cytology with molecular DNA changes. Each probe is a fluorescently labeled, single-stranded DNA fragment (nucleic acid sequence) complementary to specific target sequences of cellular DNA that are denatured to allow hybridization with the probe. Fluorescence microscopy allows visualization of the hybridized, labeled probe. The kit contains a mixture of unlabeled blocking DNA to suppress sequences contained within the target loci that are common to other chromosomes. A minimum of 25 morphologically abnormal cells is viewed. If 4 or more cells exhibit polysomy, the case is considered positive for tumor. However, no uniform criterion exists for a positive UroVysion assay at this time. In all published comparative studies, FISH outperforms cytology across all stages and grades of UCB [63–66]. The overall sensitivity

Positive florescence in situ hybridization (FISH) images. (A) FISH-positive nucleus; (B) chromosome 3, SpectrumRed; (C) chromosome 7, SpectrumGreen; (D) chromosome 17, SpectrumAqua; (E) locus 9p21, SpectrumGold. Reprinted with permission from Moonen et al [104].

Vol. 6, No. 4, pp. 368–382, December 2009

373

Review of FISH is superior to that of cytology (74% vs 48%), especially in high-grade disease such as carcinoma in situ (100% for FISH vs 67% for cytology). Cumulative data from comparative studies showed that the sensitivity of cytology is lower than that of FISH across all tumor grades: 19% versus 58% for grade 1 tumors, 50% versus 77% for grade 2 tumors, and 71% versus 96% for grade 3 tumors. Similarly, FISH outperformed cytology across all tumor stages: 64% (FISH) versus 35% for Ta, 83% versus 66% for T1, and 94% versus 76% for muscle-invasive carcinoma. A recent meta-analysis of FISH showed that overall performance of FISH was better than that of cytology (area under the curve: 87% vs 63%) [67]. This difference, however, was almost entirely attributable to the difference in performance in diagnosing pTa patients; indeed, the higher performance disappeared when pTa patients were excluded from the analysis (area under the curve: 94% vs 91%). Combining morphology with FISH may prove an alternative modality of cancer detection [68,69]. Using the Duet automatic scanning system (BioView, Ltd., Rehovat, Israel), which examines both morphology and FISH scoring, an examination of voided urine specimens from 115 patients with negative or aty-

Figure 3

374

pical cytology found that 44% of these patients did not have a prior diagnosis of UCB. The combination of morphology and FISH resulted in 100% sensitivity and 65% specificity [68]. Atypical findings are problematic because they raise concerns with the patient and the physician about the possible presence of cancer. One of the possible applications of FISH is to clarify the therapeutic dilemma associated with an atypical cytologic result. To address this issue, investigators prospectively tested the value of reflex FISH in atypical cytology patients with (n = 70) and without (n = 50) a prior history of UCB. Reflex FISH detected all visible tumors, and had 83–100% sensitivity and 78–86% specificity for equivocal or negative cystoscopies [70]. However, the PPV was only between 43% and 62%. The authors concluded that FISH assay was unnecessary in patients with obvious tumors on cystoscopy but was beneficial in those with equivocal or negative cystoscopy findings. The authors proposed a management algorithm using reflex FISH assay (Fig. 3). Several limitations have precluded a wider use of this test by urologists: the high cost and the necessity of large urine volume and/or tumor burden as well as exfoliation of tumor cells. Bladder washing may help increase the number of cells available for inspection with

Management algorithm using reflex florescence in situ hybridization (FISH) assay. In patients with a negative lesion and positive FISH, one should strongly consider performing a biopsy of the lesion and possibly random biopsies of the bladder and prostatic urethra and an upper tract evaluation. In patients with equivocal cystoscopy and positive FISH, one should ensure that there has been a recent evaluation of the upper tract and close monitoring. Reprinted with permission from Lotan et al [70].

Vol. 6, No. 4, pp. 368–382, December 2009

Review FISH, thereby increasing the diagnostic yield [71]. Finally, another limitation is that the FISH assay does not detect diploid cells without 9p21 deletions. Although there is a relatively high rate of false-positive results due to a low PPV of the test, findings may suggest an anticipatory positive result, in which a premalignant change would precede the discovery of a recurrent malignancy [64,72,73]. One study [64] found that 89% of the patients that had a false-positive test had a positive bladder biopsy within 12 months of the test, while another found that FISH preceded tumor recurrence in 85% of patients [72]. Nonetheless, the real role of an anticipatory positive result is still unclear as most patients with non-muscleinvasive UCB eventually experience disease recurrence.

Investigational UCB Biomarkers Urinary UBC test IDL Biotech AB (Bromma, Sweden) developed the UBC, a point-of-care qualitative assay, and the UBC enzyme-linked immunosorbent assay (UBC II ELISA), a quantitative assay that measures cytokeratins 8 and 18 in the urine [74,75]. Cytokeratins are intermediate filament proteins that are characteristic of epithelial cells [76]. A study measuring UBC in the urine of 180 patients found an overall sensitivity of 66% and specificity of 90% [75]. In a comparative study, however, BTA stat and BTA TRAK outperformed the UBC Rapid Test, particularly with regards to sensitivity [77]. Similarly, cytology had a better sensitivity and specificity than either UBC or UBC II ELISA in a somewhat smaller study group [78]. More recently, investigators compared the sensitivity and specificity of cytology (19.8% and 99%), BTA (53.8% and 83.9%), and UBC (12.1% and 97.2%) [33]. For carcinoma in situ, UBC had a higher sensitivity (100%) compared to cytology (66.6%) and BTA (0%). The performance of this UBC test, in sum, is not superior to cytology or other current biomarkers and is therefore unlikely to supplant or replace them.

BLCA-1 and BLCA-4 BLCA-1 and BLCA-4 are nuclear transcription factors present in UCB. BLCA-1 is not expressed in nonmalignant urothelium [79], while BLCA-4

is expressed in both the tumor and adjacent benign areas of the bladder but not in noncancerous bladders [80]. BCLA-4 is measured in the urine using an ELISA assay; its sensitivity ranges from 89–96% and its specificity reaches 100% [81,82]. Similarly, in a small study, BLCA-1 demonstrated good performance with 80% sensitivity and 87% specificity [79]. Tumor grade did not seem to affect the expression of these markers. However, BCLA-4 levels are elevated in up to 19% of patients with spinal cord injuries [83]. These biomarkers are in their discovery phase, awaiting assay refinement and validation.

Hyaluronic acid and hyaluronidase Hyaluronic acid (HA), a nonsulfated glycosaminoglycan, is associated with tumor metastasis and interferes with immune surveillance [84]. At a cutoff value of 100 ng/ml, urine hyaluronic acid has been shown to yield 92% sensitivity and 93% specificity for detecting UCB [85]. Hyaluronidase (HAase), an endoglycosidase, degrades HA into small fragments that promote angiogenesis [86]. There is a correlation between the secretion of hyaluronidase by UCB cells and their invasive potential. An analysis of 139 urine specimens detected a 5- to 8-fold elevation of HAase in the urine of patients with grade 2 or 3 UCB [87]. The levels of HA and HAase can be combined in the HA–HAase test [88]. In a study of 225 urine samples from 70 patients with known UCB, the HA–HAase test performed better than the BTA stat test, yielding a sensitivity greater than 90% across all tumor grades [89]. However, the accuracy of HA–HAase for detecting low-grade tumors is poor and may even be less than that of routine voided-urine cytology [90]. Further refinement of the assay and evaluation in larger clinical trials would help define the clinical applicability of this marker.

Lewis X antigen Lewis-related antigens are cell-surface molecules divided into four subclasses, of which only the Lewis X group is associated with UCB [76]. The Lewis X antigen is expressed in epithelium from UCB cells, regardless of the tumor grade or stage [91]. Overall, the sensitivity ranges from approximately 80–94% with a specificity of approximately 83% [92–94]. The sensitivity increases when two consecutive urine samples are examined. Nonetheless,

Vol. 6, No. 4, pp. 368–382, December 2009

375

Review further studies are necessary before recommending this test for clinical care.

Microsatellite analysis One of the most common genetic changes in UCB is loss of heterogeneity in chromosome 9 [76,95]. Chromosomes 4p, 8p, 9p, 11p, and 17p also often display loss of heterogeneity in patients with UCB [96,97]. Several studies have analyzed voided urine with 17–20 microsatellite biomarkers [95,98]. The overall sensitivity from these studies ranged from 72–97%, and overall specificity ranged from 80–100%. Microsatellite biomarkers outperformed cytology in low-grade, low-stage tumors. In a study of 228 patients, the authors showed that the sensitivity of the test was 58% and the specificity was 73% [99]. While promising, the routine clinical use of microsatellite analysis is still questionable.

Figure 4

Quanticyt An altered number of chromosomes is a characteristic of many types of tumors. Quanticyt (Gentian Scientific Software, Niawier, The Netherlands) aims to detect aneuploid cells using flow cytometry on bladder washings [100]. Initial reports for this assay were promising, but subsequent studies demonstrated that the sensitivity of the test was only marginally better than that of urine cytology with decreased specificity [101,102]. Overall sensitivity of the Quanticyt assay ranges from 45–69% [16,100,103], and its specificity ranges from 70–93% [16,100,103,104]. Moreover, the availability of the test may be restricted due to high cost and the need for a specialized laboratory.

Soluble Fas As an apoptotic-related mediator, Fas has been associated with UCB biologic behavior. A conventional ELISA assay showed that higher urinary levels of soluble Fas (sFas) are independently associated with UCB recurrence and progression to invasive tumor stage, even after adjusting for the effects of cytology, NMP22, and patient age [105]. This association remained significant in patients with a normal cytology. While the overall performance of urinary sFas was not significantly different from NMP22, at sensitivity values above 75% sFas had a consis-

376

Vol. 6, No. 4, pp. 368–382, December 2009

Receiver operating curves of sFas and NMP22 for prediction of bladder urothelial carcinoma presence. Abbreviations: solid line, soluble Fas (sFas); dotted line, nuclear mitotic apparatus protein (NMP22); AUC, area under the receiver operating curve; CI, confidence interval. Reprinted with permission Svatek et al [105].

tently higher specificity than NMP22 (Fig. 4). This may give sFas an important advantage over other urinary markers, since false-positive results are a main concern for clinical use. Nevertheless, further assay improvements are needed in addition to external validation in large studies.

Survivin Survivin is a novel member of the inhibitor-ofapoptosis gene family that counteracts cell death, controls mitotic progression, and induces changes in gene expression that are associated with tumor-cell invasiveness [106]. Survivin messenger ribonucleic acid (mRNA) is overexpressed in human cancers and can be detected in urine using a bio-dot immunoassay incorporating a rabbit polyclonal antisurvivin antibody [107]. Urinary levels of survivin gene activation, both at the protein and the mRNA level, are associated with UCB presence, higher grade, and advanced pathologic stage [108–111]. In the first study to evaluate the diagnostic potential of survivin in UCB using both protein and mRNA detection methods, survivin protein and mRNA were detected in all of 47 patients with new, recurrent, or active UCB, but in only 3 of 35 patients with a negative

Review cystoscopic evaluation [112]. Another study found that urinary levels of survivin correlate with increased risk of UCB presence and higher grade, but not tumor invasion [109]. In that study, survivin sensitivity was 64% and specificity was 93%. Another study reported that survivin mRNA copy number (obtained from bladder washings) correlated with recurrencefree survival [110]. Similarly, survivin mRNA was detected in urine from 24 of 35 (68%) patients with UCB [111]. None of the 33 healthy patients had detectable urinary survivin mRNA. The authors reported a sensitivity of 68.6% and a specificity of 100% for urinary survivin mRNA, compared to 31.4% and 97.1%, respectively, for voided cytology. The urinary detection of survivin seems an accurate diagnostic test for UCB, regardless of tumor stage and grade. Although these results are promising, the lack of assay standardization and cutoff value are important issues to be resolved before survivin can be used in the clinical setting [109].

Telomerase Telomerase is a ribonucleoprotein enzyme that acts in chromosomal instability by synthesizing telomeres [113–116]. In normal somatic tissue, cells do not produce telomerase. Malignant neoplasms, including UCB [117], have been shown to produce telomerase and thus to regenerate telomeres and prevent cell death [118]. The standard technique to measure telomerase activity is the telomeric repeat amplification protocol (TRAP) assay [118]. Another telomerase-based assay detects the catalytic subunit of telomerase, human telomerase reverse transcriptase, using polymerase chain reaction (PCR). When compared with TRAP, human telomerase reverse transcriptase PCR has higher sensitivity than the TRAP assay, ranging between 75% and 100% [119,120]. The overall sensitivity of telomerase testing for detection of UCB is between 7% and 100%, with the results of most studies between 70% and 86% [20,52,121–129]. The overall specificity of telomerase for UCB varies from 24–90%, mostly in the range of 60–70% [20,52,121,123– 128,130,131]. In 2005, researchers who examined a range of TRAP cutoffs found good overall performance characteristics [132]. Using these various cutoffs, they calculated an area under the receiver operating characteristic curve of

0.951 for the assay, and with an arbitrary cutoff of 50 enzyme units, the test had a sensitivity of 90% with a specificity of 88%, which persisted across all grades of tumor. However, because many UCB patients have other urological and nonurological comorbidities, the clinical applicability of the telomerase assay may be limited. Another possible limitation of this test is the potential for inactivation of the telomerase enzyme in urine, leading to extremely low sensitivity (7% in one study) [129]. Considering the need for trained personnel in a reference laboratory and the wide range of results from different studies, telomerase assays are not yet ready for clinical use.

Cytokeratin 20 Recent data has shown an association of cytokeratin 20 mRNA expression level with UBC [133]. When quantitative real-time PCR was performed in 95 urine samples, cytokeratin 20 expression diagnosed 51 cases (85%) with UBC yielding a specificity of 94.3%. The authors also showed a correlation between cytokeratin 20 expression and the clinicopathologic features of UBC, such as tumor stage and grade. This preliminary data needs further testing.

Conclusions In summary, the published data on urinary markers have shown a wide range of efficacy for bladder cancer detection, depending on various factors and selection criteria. BTA stat, BTA TRAK and Immuncyt showed an improved role for detecting low-grade tumors, while the latter may better be reserved for patients previously diagnosed with UCB, due to its low PPV. Similarly, NMP22 is also suitable for surveillance. Additionally, NMP22 is approved for UCB detection in high-risk patients, although a high false-positive rate may preclude its overall superiority over the other tests. UroVision has shown to be superior to cytology in all stages and is significantly better in pTa tumors. Moreover, a possible anticipatory detection of premalignant changes may be addressed in future studies. In conclusion, noninvasive UCB tests may significantly improve the management of nonmuscle-invasive UCB. These tests may be used

Vol. 6, No. 4, pp. 368–382, December 2009

377

Review to screen patients at high risk, help diagnose or even predict disease recurrence, decreasing the need for invasive procedures. They may initially act as an adjunct to current tests, but eventually replace them. Most of the comparative studies have shown that noninvasive biomarkers have equal or higher sensitivity for UCB detection than cytology, even in highgrade cancers. None of these tests, however, meets all of the criteria of an ideal tumor biomarker. Improved standardization and automation of the methods are still required, as well as prospective, large-scale validation in heterogeneous patient populations. Some of the FDA-approved UCB urine biomarkers, such as NMP22 and UroVysion, should be gradually incorporated into clinical practice as adjunctive tests to cystoscopy in the follow-up of patients with UCB. In selected cases of lowgrade, stage Ta UCB, a reduced frequency of cystoscopies should be applied [134]. The decision-making process for follow-up may be also influenced by the patient perception of efficacy. A recent study [135] found that patients chose flexible cystoscopy over a bladder biomarker when the accuracy of the biomarker was less than 90%. While biomarkers are intended to provide an accurate, less inva-

sive alternative, biomarkers that do not have a high specificity could lead to more invasive tests than would otherwise be recommended, due to false-positive results. Great interest in this topic has generated significant improvement in the search for an ideal urinary biomarker. Protein-expression profiling of UCB offers an alternative means of distinguishing aggressive tumor biology and may improve the accuracy of outcome prediction. Moreover, a direct measure of therapy response may be acquired with promising new tests. Towards identifying and validating biomarkers for cancer care, we need to establish formal, systematic research phases such as those used for drugs [136]. Until then, the accepted standard for early detection and monitoring of UCB remains cystoscopy complemented with one of the FDA-approved tests such as cytology, FISH, or NMP22.

Acknowledgement This work was supported by The Sidney Kimmel Center for Prostate and Urologic Cancers. Research activities are also supported in part by a NIH T32 training grant.

References [1] Jemal A, Siegel R, Ward E, Hao Y, Xu J, Murray T, et al. Cancer statistics, 2008. CA Cancer J Clin 2008;58(2):71–96. [2] Shariat SF, Karakiewicz PI, Palapattu GS, Lotan Y, Rogers CG, Amiel GE, et al. Outcomes of radical cystectomy for transitional cell carcinoma of the bladder: a contemporary series from the bladder cancer research consortium. J Urol 2006;176(6): 2414–22. [3] Stein JP, Lieskovsky G, Cote R, Groshen S, Feng AC, Boyd S, et al. Radical cystectomy in the treatment of invasive bladder cancer: long-term results in 1,054 patients. J Clin Oncol 2001;19(3):666–75. [4] Madersbacher S, Hochreiter W, Burkhard F, Thalmann GN, Danuser H, Markwalder R, et al. Radical cystectomy for bladder cancer today – a homogeneous series without neoadjuvant therapy. J Clin Oncol 2003;21(4):690–6. [5] Prout Jr GR, Barton BA, Griffin PP, Friedell GH. The National Bladder Cancer Group. Treated history of noninvasive grade 1 transitional cell carcinoma. J Urol 1992;148(5):1413–9.

378

Vol. 6, No. 4, pp. 368–382, December 2009

[6] Pagano F, Bassi P, Galetti TP, Meneghini A, Milani C, Artibani W, et al. Results of contemporary radical cystectomy for invasive bladder cancer: a clinicopathological study with an emphasis on the inadequacy of the tumor, nodes and metastases classification. J Urol 1991;145(1):45–50. [7] van der Poel HG, Debruyne FM. Can biological markers replace cystoscopy? An update Curr Opin Urol 2001;11(5):503–9. [8] Herr HW. The natural history of a T1 bladder cancer: life-long tumour diathesis. BJU Int 1999;84(9):1102–3. [9] Lotan Y, Roehrborn CG. Sensitivity and specificity of commonly available bladder tumor markers versus cytology: results of a comprehensive literature review and metaanalyses. Urology 2003;61(1):109–18: discussion 118. [10] Karakiewicz PI, Benayoun S, Zippe C, Ludecke G, Boman H, Sanchez-Carbayo M, et al. Institutional variability in the accuracy of urinary cytology for predicting recurrence of transitional cell carcinoma of the bladder. BJU Int 2006;97(5):997– 1001.

[11] Lokeshwar V, Soloway M. Current bladder tumor tests: does their projected utility fulfill clinical necessity? J Urol 2001;165: 1067–77. [12] Svatek RS, Sagalowsky AI, Lotan Y. Economic impact of screening for bladder cancer using bladder tumor markers: a decision analysis. Urol Oncol 2006;24(4):338–43. [13] Lotan Y, Svatek RS, Sagalowsky AI. Should we screen for bladder cancer in a high-risk population?: A cost per life-year saved analysis Cancer 2006;107(5):982–90. [14] Johnston B, Morales A, Emerson L, Lundie M. Rapid detection of bladder cancer: a comparative study of point of care tests. J Urol 1997;158:2098–101. [15] Kinders R, Jones T, Root R, Bruce C, Murchison H, Corey M, et al. Complement factor H or a related protein is a marker for transitional cell cancer of the bladder. Clin Cancer Res 1998;4(10):2511–20. [16] Wiener HG, Mian C, Haitel A, Pycha A, Schatzl G, Marberger M. Can urine bound diagnostic tests replace cystoscopy in the management of bladder cancer? J Urol 1998;159(6):1876–80.

Review [17] Takashi M, Schenck U, Kissel K, Leyh H, Treiber U. Use of diagnostic categories in urinary cytology in comparison with the bladder tumour antigen (BTA) test in bladder cancer patients. Int Urol Nephrol 1999;31(2):189–96. [18] Sharma S, Zippe CD, Pandrangi L, Nelson D, Agarwal A. Exclusion criteria enhance the specificity and positive predictive value of NMP22 and BTA stat. J Urol 1999;162(1):53–7. [19] Gutierrez Banos JL, Martin Garcia B, Hernandez Rodriguez R, Portillo Martin JA, Correas Gomez MA, del Valle Schaan JI, et al. Utilidad del BTA Stat Test (Bard) en el diagno´stico del ca´ncer vesical. Resultados preliminares y comparacio´n con citologı´a y cistoscopia. Arch Esp Urol 1998;51(8): 778–82. [20] Landman J, Chang Y, Kavaler E, Droller MJ, Liu BC. Sensitivity and specificity of NMP22, telomerase, and BTA in the detection of human bladder cancer. Urology 1998;52(3):398–402. [21] Leyh H, Mazeman E. Bard BTA test compared with voided urine cytology in the diagnosis of recurrent bladder cancer. Eur Urol 1997;32:425–8. [22] Leyh H, Marberger M, Conort P, Sternberg C, Pansadoro V, Pagano F, et al. Comparison of the BTA stat test with voided urine cytology and bladder wash cytology in the diagnosis and monitoring of bladder cancer. Eur Urol 1999;35(1):52–6. [23] Nasuti JF, Gomella LG, Ismial M, Bibbo M. Utility of the BTA stat test kit for bladder cancer screening. Diagn Cytopathol 1999;21(1):27–9. [24] Oge O, Kozaci D, Gemalmaz H. The BTA stat test is nonspecific for hematuria: an experimental hematuria model. J Urol 2002;167(3):1318–9: discussion 1319-20. [25] Raitanen MP. The role of BTA stat test in follow-up of patients with bladder cancer: results from FinnBladder studies. World J Urol 2008;26(1):45–50. [26] Malkowicz SB. The application of human complement factor H-related protein (BTA TRAK) in monitoring patients with bladder cancer. Urol Clin of North Am 2000;27(1):63–73. [27] Thomas L, Leyh H, Marberger M, Bombardieri E, Bassi P, Pagano F, et al. Multicenter trial of the quantitative BTA TRAK assay in the detection of bladder cancer. Clin Chem 1999;45(4):472–7. [28] Mahnert B, Tauber S, Kriegmair M, Schmitt UM, Hasholzner U, Reiter W, et al. BTATRAK – a useful diagnostic tool in urinary bladder cancer? Anticancer Res 1999;19(4A):2615–9. [29] Irani J, Desgrandchamps F, Millet C, Toubert ME, Bon D, Aubert J, et al. BTA stat and BTA TRAK: a comparative evaluation

[30]

[31]

[32]

[33]

[34]

[35]

[36]

[37]

[38]

[39]

[40]

[41]

of urine testing for the diagnosis of transitional cell carcinoma of the bladder. Eur Urol 1999;35(2):89–92. Heicappell R, Wettig IC, Schostak M, Muller M, Steiner U, Sauter T, et al. Quantitative detection of human complement factor H-related protein in transitional cell carcinoma of the urinary bladder. Eur Urol 1999;35(1):81–7. Ellis WJ, Blumenstein BA, Ishak LM, Enfield DL. The Multi Center Study Group. Clinical evaluation of the BTA TRAK assay and comparison to voided urine cytology and the Bard BTA test in patients with recurrent bladder tumors. Urology 1997;50(6): 882–7. Abbate I, D’Introno A, Cardo G, Marano A, Addabbo L, Musci M, et al. Comparison of nuclear matrix protein 22 and bladder tumor antigen in urine of patients with bladder cancer. Anticancer Res 1998;18(5B):3803–5. Babjuk M, Soukup V, Pesl M, Kostirova M, Drncova E, Smolova H, et al. Urinary cytology and quantitative BTA and UBC tests in surveillance of patients with pTapT1 bladder urothelial carcinoma. Urology 2008;71(4):718–22. Tsui KH, Chen SM, Wang TM, Juang HH, Chen CL, Sun GH, et al. Comparisons of voided urine cytology, nuclear matrix protein-22 and bladder tumor associated antigen tests for bladder cancer of geriatric male patients in Taiwan, China. Asian J Androl 2007;9(5):711–5. Fradet Y, Lockhart C. Performance characteristics of a new monoclonal antibody test for bladder cancer: ImmunoCyt trade mark. Can J Urol 1997;4:400. Greene KL, Berry A, Konety BR. Diagnostic utility of the ImmunoCyt/uCyt+ test in bladder cancer. Rev Urol 2006;8(4):190–7. Olsson H, Zackrisson B. ImmunoCyt a useful method in the follow-up protocol for patients with urinary bladder carcinoma. Scand J Urol Nephrol 2001;35:280–2. Mian C, Pycha A, Wiener H, Haitel A, Lodde M, Marberger M. Immunocyt: a new tool for detecting transitional cell cancer of the urinary tract. J Urol 1999;161(5):1486–9. Vriesema JL, Atsma F, Kiemeney LA, Peelen WP, Witjes JA, Schalken JA. Diagnostic efficacy of the ImmunoCyt test to detect superficial bladder cancer recurrence. Urology 2001;58(3):367–71. Mian C, Lodde M, Comploj E, Negri G, Egarter-Vigl E, Lusuardi L, et al. Liquidbased cytology as a tool for the performance of uCyt+ and Urovysion Multicolour-FISH in the detection of urothelial carcinoma. Cytopathology 2003;14(6): 338–42. Pfister C, Chautard D, Devonec M, Perrin P, Chopin D, Rischmann P, et al. Immunocyt

[42]

[43]

[44]

[45]

[46]

[47]

[48]

[49]

[50]

[51]

[52]

[53]

test improves the diagnostic accuracy of urinary cytology: results of a French multicenter study. J Urol 2003;169(3):921–4. Sullivan PS, Nooraie F, Sanchez H, Hirschowitz S, Levin M, Rao PN, et al. Comparison of ImmunoCyt, UroVysion, and urine cytology in detection of recurrent urothelial carcinoma: a ‘‘split-sample’’ study. Cancer Cytopathol 2009;117(3):167–73. Gordon JN, Shu WP, Schlussel RN, Droller MJ, Liu BCS. Altered extracellular matrices influence cellular processes and nuclear matrix organisations of overlying human bladder urothelial cells. Cancer Res 1993;53:4971–7. Berezney R, Coffey DS. Identification of a nuclear protein matrix. Biochem Biophys Res Commun 1974;60(4):1410–7. Pardoll DM, Vogelstein B, Coffey DS. A fixed site of DNA replication in eukaryotic cells. Cell 1980;19:527–36. Compton DA, Cleveland DW. NuMA is required for the proper completion of mitosis. J Cell Biol 1993;120:947–57. Shelfo SW, Soloway MS. The role of nuclear matrix protein 22 in the detection of persistent or recurrent transitional-cell cancer of the bladder. World J Urol 1997;15:107–11. Hughes JH, Katz RL, Rodriguez-Villanueva J, Kidd L, Dinney C, Grossman HB, et al. Urinary nuclear matrix protein 22 (NMP22): a diagnostic adjunct to urine cytologic examination for the detection of recurrent transitional-cell carcinoma of the bladder. Diagn Cytopathol 1999;20(5):285–90. Miyanaga N, Akaza H, Tsukamoto T, Ishikawa S, Noguchi R, Ohtani M, et al. Urinary nuclear matrix protein 22 as a new marker for the screening of urothelial cancer in patients with microscopic hematuria. Int J Urol 1999;6(4):173–7. Stampfer D, Carpinito G, Rodriguez-Villanueva J, Willsey L, Dinney C, Grossman H, et al. Evaluation of NMP22 in the detection of transitional cell carcinoma of the bladder. J Urol 1998;159:394–8. Soloway MS, Briggman JV, Carpinito GA, Chodak GW, Church PA, Lamm DL, et al. Use of new tumor marker, urinary NMP22, in detection of occult or rapidly recurring transitional cell carcinoma of the urinary tract following surgical treatment. J Urol 1996;156(2 Pt 1):363–7. Ramakumar S, Bhuiyan J, Besse JA, Roberts SG, Wollan PC, Blute ML, et al. Comparison of screening methods in the detection of bladder cancer. J Urol 1999;161(2): 388–94. Ponsky LE, Sharma S, Pandrangi L, Kedia S, Nelson D, Agarwal A, et al. Screening and monitoring for bladder cancer: refining the use of NMP22. J Urol 2001;166:75–8.

Vol. 6, No. 4, pp. 368–382, December 2009

379

Review [54] Mansoor I, Calam RR, Al-Khafaji B. Role of urinary NMP-22 combined with urine cytology in follow-up surveillance of recurring superficial bladder urothelial carcinoma. Anal Quant Cytol Histol 2008;30(1):25–32. [55] Shariat SF, Marberger MJ, Lotan Y, Sanchez-Carbayo M, Zippe C, Ludecke G, et al. Variability in the performance of nuclear matrix protein 22 for the detection of bladder cancer. J Urol 2006;176(3): 919–26. [56] Shariat SF, Casella R, Wians Jr FH, Ashfaq R, Balko J, Sulser T, et al. Risk stratification for bladder tumor recurrence, stage and grade by urinary nuclear matrix protein 22 and cytology. Eur Urol 2004;45(3):304–13: author reply 313. [57] Shariat SF, Zippe C, Ludecke G, Boman H, Sanchez-Carbayo M, Casella R, et al. Nomograms including nuclear matrix protein 22 for prediction of disease recurrence and progression in patients with Ta, T1 or CIS transitional cell carcinoma of the bladder. J Urol 2005;173(5):1518–25. [58] Grossman HB, Messing E, Soloway M, Tomera K, Katz G, Berger Y, et al. Detection of bladder cancer using a point-of-care proteomic assay. JAMA 2005;293(7):810–6. [59] Grossman HB, Soloway M, Messing E, Katz G, Stein B, Kassabian V, et al. Surveillance for recurrent bladder cancer using a point-of-care proteomic assay. JAMA 2006;295(3):299–305. [60] Lotan Y, Shariat SF. Impact of risk factors on the performance of the nuclear matrix protein 22 point-of-care test for bladder cancer detection. BJU Int 2008;101(11): 1362–7. [61] Lotan Y, Capitanio U, Shariat SF, Hutterer GC, Karakiewicz PI. Impact of clinical factors, including a point-of-care nuclear matrix protein-22 assay and cytology, on bladder cancer detection. BJU Int 2009;103(10):1368–74. [62] Junker K, Boerner D, Schulze W, Utting M, Schubert J, Werner W. Analysis of genetic alterations in normal bladder urothelium. Urology 2003;62(6):1134–8. [63] Halling KC. Vysis UroVysion for the detection of urothelial carcinoma. Expert Rev Mol Diagn 2003;3(4):507–19. [64] Skacel M, Fahmy M, Brainard JA, Pettay JD, Biscotti CV, Liou LS, et al. Multitarget fluorescence in situ hybridization assay detects transitional cell carcinoma in the majority of patients with bladder cancer and atypical or negative urine cytology. J Urol 2003;169(6):2101–5. [65] Friedrich MG, Hellstern A, Toma MI, Hammerer P, Huland H. Are false-positive urine markers for the detection of bladder carcinoma really wrong or do they predict tumor recurrence? Eur Urol 2003;43(2): 146–50: discussion 150-1.

380

Vol. 6, No. 4, pp. 368–382, December 2009

[66] Halling KC, King W, Sokolova IA, Karnes RJ, Meyer RG, Powell EL, et al. A comparison of BTA stat, hemoglobin dipstick, telomerase and Vysis UroVysion assays for the detection of urothelial carcinoma in urine. J Urol 2002;167(5):2001–6. [67] Hajdinjak T. UroVysion FISH test for detecting urothelial cancers: meta-analysis of diagnostic accuracy and comparison with urinary cytology testing. Urol Oncol 2008;26(6):646–51. [68] Daniely M, Rona R, Kaplan T, Olsfanger S, Elboim L, Freiberger A, et al. 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. Cancer 2007;111(6):517–24. [69] Daniely M, Rona R, Kaplan T, Olsfanger S, Elboim L, Zilberstien Y, et al. Combined analysis of morphology and fluorescence in situ hybridization significantly increases accuracy of bladder cancer detection in voided urine samples. Urology 2005;66(6):1354–9. [70] Lotan Y, Bensalah K, Ruddell T, Shariat SF, Sagalowsky AI, Ashfaq R. Prospective evaluation of the clinical usefulness of reflex fluorescence in situ hybridization assay in patients with atypical cytology for the detection of urothelial carcinoma of the bladder. J Urol 2008;179(6):2164–9. [71] Bergman J, Reznichek RC, Rajfer J. Surveillance of patients with bladder carcinoma using fluorescent in-situ hybridization on bladder washings. BJU Int 2008;101(1):26– 9. [72] Gofrit ON, Zorn KC, Silvestre J, Shalhav AL, Zagaja GP, Msezane LP, et al. The predictive value of multi-targeted fluorescent insitu hybridization in patients with history of bladder cancer. Urol Oncol 2008;26(3):246–9. [73] Yoder BJ, Skacel M, Hedgepeth R, Babineau D, Ulchaker JC, Liou LS, et al. Reflex UroVysion testing of bladder cancer surveillance patients with equivocal or negative urine cytology: a prospective study with focus on the natural history of anticipatory positive findings. Am J Clin Pathol 2007;127(2):295–301. [74] Heicappell R, Schostack M, Mu¨ller M, Miller K. Evaluation of urinary bladder cancer antigen as a marker for diagnosis of transitional cell carcinoma of the urinary bladder. Scand J Clin Lab Invest 2000;60(4):275–82. [75] Mian C, Lodde M, Haitel A, Egarter Vigl E, Marberger M, Pycha A. Comparison of two qualitative assays, the UBC rapid test and the BTA stat test, in the diagnosis of urothelial cell carcinoma of the bladder. Urology 2000;56(2):228–31. [76] Saad A, Hanbury DC, McNicholas TA, Boustead GB, Woodman AC. The early

[77]

[78]

[79]

[80]

[81]

[82]

[83]

[84]

[85]

[86]

[87]

[88]

detection and diagnosis of bladder cancer: a critical review of the options. Eur Urol 2001;39:619–33. Babjuk M, Kostirova M, Mudra K, Pecher S, Smolova H, Pecen L, et al. Qualitative and quantitative detection of urinary human complement factor H-related protein (BTA stat and BTA TRAK) and fragments of cytokeratins 8, 18 (UBC rapid and UBC IRMA) as markers for transitional cell carcinoma of the bladder. Eur Urol 2002;41(1):34–9. Hakenberg OW, Fuessel S, Richter K, Froehner M, Oehlschlaeger S, Rathert P, et al. Qualitative and quantitative assessment of urinary cytokeratin 8 and 18 fragments compared with voided urine cytology in diagnosis of bladder carcinoma. Urology 2004;64(6):1121–6. Myers-Irvin JM, Landsittel D, Getzenberg RH. Use of the novel marker BLCA-1 for the detection of bladder cancer. J Urol 2005;174(1):64–8. Konety BR, Nguyen TS, Dhir R, Day RS, Becich MJ, Stadler WM, et al. Detection of bladder cancer using a novel nuclear matrix protein, BLCA-4. Clin Cancer Res 2000;6(7):2618–25. Van Le TS, Myers J, Konety BR, Barder T, Getzenberg RH. Functional characterization of the bladder cancer marker, BLCA4. Clin Cancer Res 2004;10(4):1384–91. Van Le TS, Miller R, Barder T, Babjuk M, Potter DM, Getzenberg RH. Highly specific urine-based marker of bladder cancer. Urology 2005;66(6):1256–60. Konety BR, Nguyen TS, Brenes G, Sholder A, Lewis N, Bastacky S, et al. Clinical usefulness of the novel marker BLCA-4 for the detection of bladder cancer. J Urol 2000;164(3 Pt 1):634–9. Knudson W. Tumor-associated hyaluronan. Providing an extracellular matrix that facilitates invasion. Am J Pathol 1996;148(6):1721–6. Lokeshwar VB, Obek C, Soloway MS, Block NL. Tumor-associated hyaluronic acid: a new sensitive and specific urine marker for bladder cancer. Cancer Res 1997;57(4): 773–7. Lokeshwar VB, Block NL. HA-HAase urine test. A sensitive and specific method for detecting bladder cancer and evaluating its grade. Urol Clin North Am 2000;27(1): 53–61. Pham HT, Block NL, Lokeshwar VB. Tumorderived hyaluronidase: a diagnostic urine marker for high-grade bladder cancer. Cancer Res 1997;57(4):778–83. Hautmann SH, Lokeshwar VB, Schroeder GL, Civantos F, Duncan RC, Gnann R, et al. Elevated tissue expression of hyaluronic acid and hyaluronidase validates the HAHAase urine test for bladder cancer. J Urol 2001;165(6 Pt 1):2068–74.

Review [89] Lokeshwar VB, Schroeder GL, Selzer MG, Hautmann SH, Posey JT, Duncan RC, et al. Bladder tumor markers for monitoring recurrence and screening comparison of hyaluronic acid-hyaluronidase and BTAStat tests. Cancer 2002;95(1):61–72. [90] Hautmann S, Toma M, Lorenzo Gomez MF, Friedrich MG, Jaekel T, Michl U, et al. Immunocyt and the HA-HAase urine tests for the detection of bladder cancer: a side-by-side comparison. Eur Urol 2004;46(4):466–71. [91] Decenzo JM, Howard P, Irish CE. Antigenic deletion and prognosis of patients with stage A transitional cell bladder carcinoma. J Urol 1975;114(6):874–8. [92] Golijanin D, Sherman Y, Shapiro A, Pode D. Detection of bladder tumors by immunostaining of the Lewis X antigen in cells from voided urine. Urology 1995;46(2):173–7. [93] Pode D, Golijanin D, Sherman Y, Lebensart P, Shapiro A. Immunostaining of Lewis X in cells from voided urine, cytopathology and ultrasound for noninvasive detection of bladder tumors. J Urol 1998;159(2):389– 92: discussion 393. [94] Friedrich MG, Hellstern A, Hautmann SH, Graefen M, Conrad S, Huland E, et al. Clinical use of urinary markers for the detection and prognosis of bladder carcinoma: a comparison of immunocytology with monoclonal antibodies against Lewis X and 486p3/12 with the BTA STAT and NMP22 tests. J Urol 2002;168(2):470–4. [95] van Rhijn BW, Lurkin I, Kirkels WJ, van der Kwast TH, Zwarthoff EC. Microsatellite analysis – DNA test in urine competes with cystoscopy in follow-up of superficial bladder carcinoma: a phase II trial. Cancer 2001;92(4):768–75. [96] Czerniak B, Chaturvedi V, Li L, Hodges S, Johnston D, Roy JY, et al. Superimposed histologic and genetic mapping of chromosome 9 in progression of human urinary bladder neoplasia: implications for a genetic model of multistep urothelial carcinogenesis and early detection of urinary bladder cancer. Oncogene 1999;18(5):1185–96. [97] Knowles MA, Elder PA, Williamson M, Cairns JP, Shaw ME, Law MG. Allelotype of human bladder cancer. Cancer Res 1994;54(2):531–8. [98] von Knobloch R, Hegele A, Brandt H, Olbert P, Heidenreich A, Hofmann R. Serum DNA and urine DNA alterations of urinary transitional cell bladder carcinoma detected by fluorescent microsatellite analysis. Int J Cancer 2001;94(1):67–72. [99] van der Aa MN, Zwarthoff EC, Steyerberg EW, Boogaard MW, Nijsen Y, van der Keur KA, et al. Microsatellite analysis of voidedurine samples for surveillence of low-grade non-muscle-invasive urothelial carcinoma:

[100]

[101]

[102]

[103]

[104]

[105]

[106]

[107]

[108]

[109]

[110]

[111]

feasibility and clinical utility in a prospective multicenter study (Cost-Effectiveness of Follow-Up of Urinary Bladder Cancer Trial [CEFUB]). Eur Urol 2009;55(3):659–68. van der Poel HG, van Balken MR, Schamhart DH, Peelen P, de Reijke T, Debruyne FM, et al. Bladder wash cytology, quantitative cytology, and the qualitative BTA test in patients with superficial bladder cancer. Urology 1998;51(1):44–50. Gregoire M, Fradet Y, Meyer F, Tetu B, Bois R, Bedard G, et al. Diagnostic accuracy of urinary cytology, and deoxyribonucleic acid flow cytometry and cytology on bladder washings during followup for bladder tumors. J Urol 1997;157(5):1660–4. Cajulis RS, Haines 3rd GK, Frias-Hidvegi D, McVary K, Bacus JW. Cytology, flow cytometry, image analysis, and interphase cytogenetics by fluorescence in situ hybridization in the diagnosis of transitional cell carcinoma in bladder washes: a comparative study. Diagn Cytopathol 1995;13(3):214–23: discussion 224. Witjes JA, van der Poel HG, van Balken MR, Debruyne FM, Schalken JA. Urinary NMP22 and karyometry in the diagnosis and follow-up of patients with superficial bladder cancer. Eur Urol 1998;33(4):387–91. Moonen PM, Merkx GF, Peelen P, Karthaus HF, Smeets DF, Witjes JA. UroVysion compared with cytology and quantitative cytology in the surveillance of non-muscleinvasive bladder cancer. Eur Urol 2007;51(5):1275–80: discussion 1280. Svatek RS, Herman MP, Lotan Y, Casella R, Hsieh JT, Sagalowsky AI, et al. Soluble Fas – a promising novel urinary marker for the detection of recurrent superficial bladder cancer. Cancer 2006;106(8):1701–7. Altieri DC. Survivin, versatile modulation of cell division and apoptosis in cancer. Oncogene 2003;22(53):8581–9. Margulis V, Lotan Y, Shariat SF. Survivin: a promising biomarker for detection and prognosis of bladder cancer. World J Urol 2008;26(1):59–65. Smith SD, Wheeler MA, Plescia J, Colberg JW, Weiss RM, Alterieri DC. Urine detection of survivin and diagnosis of bladder cancer. JAMA 2001;285(3):324–8. Shariat SF, Casella R, Khoddami SM, Hernandez G, Sulser T, Gasser TC, et al. Urine detection of survivin is a sensitive marker for the noninvasive diagnosis of bladder cancer. J Urol 2004;171(2 Pt 1):626–30. Schultz IJ, Kiemeney LA, Karthaus HF, Witjes JA, Willems JL, Swinkels DW, et al. Survivin mRNA copy number in bladder washings predicts tumor recurrence in patients with superficial urothelial cell carcinomas. Clin Chem 2004;50(8):1425–8. Weikert S, Christoph F, Schrader M, Krause H, Miller K, Mu¨ller M. Quantitative analysis

[112]

[113] [114] [115] [116]

[117]

[118]

[119]

[120]

[121]

[122]

[123]

[124]

[125]

of survivin mRNA expression in urine and tumor tissue of bladder cancer patients and its potential relevance for disease detection and prognosis. Int J Cancer 2005;116(1):100–4. Smith SD, Wheeler MA, Plescia J, Colberg JW, Weiss RM, Altieri DC. Urine detection of survivin and diagnosis of bladder cancer. JAMA 2001;285(3):324–8. Greider CW. Telomere length regulation. Annu Rev Biochem 1996;65:337–65. Blackburn EH. Telomeres and their synthesis. Science 1990;249:489–90. Blackburn EH. Structure and function of telomeres. Nature 1991;350:569–73. Morin GB. The human telomere terminal transferase enzyme is a ribonucleoprotein that synthesizes TTAGGG repeats. Cell 1989;59:521–9. Ito H, Kyo S, Kanaya T, Takakura M, Inoue M, Namiki M. Expression of human telomerase subunits and correlation with telomerase activity in urothelial cancer. Clin Cancer Res 1998;4(7):1603–8. Kim NW, Piatyszek MA, Prowse KR, Harley CB, West MD, Ho PL, et al. Specific association of human telomerase activity with immortal cells and cancer. Science 1994;266:2011–5. De Kok JB, Schalken JA, Aalders TW, Ruers TJ, Willems HL, Swinkels DW. Quantitative measurement of telomerase reverse transcriptase (hTERT) mRNA in urothelial cell carcinomas. Int J Cancer 2000;87(2): 217–20. Melissourgos N, Kastrinakis NG, Davilas I, Foukas P, Farmakis A, Lykourinas M. Detection of human telomerase reverse transcriptase mRNA in urine of patients with bladder cancer: evaluation of an emerging tumor marker. Urology 2003;62(2):362–7. Wu X, Kakehi Y, Takahashi T, Habuchi T, Ogawa O. Telomerase activity in urine after transurethral resection of superficial bladder cancer and early recurrence. Int J Urol 2000;7:210–7. Cheng CW, Chueh SC, Chern HD. Diagnosis of bladder cancer using telomerase activity in voided urine. J Formos Med Assoc 2000;99(12):920–5. Cassel A, Rahat MA, Lahat N, Lindenfield N, Mecz Y, Stein A. Telomerase activity and cytokeratin 20 as markers for the detection and followup of transitional cell carcinoma: an unfulfilled promise. J Urol 2001;166: 841–4. Kavaler E, Landman J, Chang Y, Droller MJ, Liu BC. Detecting human bladder carcinoma cells in voided urine samples by assaying for the presence of telomerase activity. Cancer 1998;82(4):708–14. Lee DH, Yang SC, Hong SJ, Chung BH, Kim IY. Telomerase: a potential marker of

Vol. 6, No. 4, pp. 368–382, December 2009

381

Review bladder transitional cell carcinoma in bladder washes. Clin Cancer Res 1998;4(3): 535–8. [126] Dalbagni G, Han W, Zhang ZF, CordonCardo C, Saigo P, Fair WR, et al. Evaluation of the telomeric repeat amplification protocol (TRAP) assay for telomerase as a diagnostic modality in recurrent bladder cancer. Clin Cancer Res 1997;3(9):1593–8. [127] Yokota K, Kanda K, Inoue Y, Kanayama H, Kagawa S. Semi-quantitative analysis of telomerase activity in exfoliated human urothelial cells and bladder transitional cell carcinoma. Br J Urol 1998;82(5):727–32. [128] Mayfield MP, Shah T, Flannigan GM, Hamilton Stewart PA, Bibby MC. Telomerase activity in malignant and benign bladder conditions. Int J Mol Med 1998;1(5): 835–40.

382

[129] Mu¨ller M, Krause H, Heicappell R, Tischendorf J, Shay JW, Miller K. Comparison of human telomerase RNA and telomerase activity in urine for diagnosis of bladder cancer. Clin Cancer Res 1998;4(8): 1949–54. [130] Lee MY, Tsou MH, Cheng MH, Chang DS, Yang AL, Ko JS. Clinical application of NMP22 and urinary cytology in patients with hematuria or a history of urothelial carcinoma. World J Urol 2000;18:401–5. [131] Weikert S, Krause H, Wolff I, Christoph F, Schrader M, Emrich T, et al. Quantitative evaluation of telomerase subunits in urine as biomarkers for noninvasive detection of bladder cancer. Int J Cancer 2005;117(2): 274–80. [132] Sanchini MA, Gunelli R, Nanni O, Bravaccini S, Fabbri C, Sermasi A, et al. Relevance

Vol. 6, No. 4, pp. 368–382, December 2009

[133]

[134]

[135]

[136]

of urine telomerase in the diagnosis of bladder cancer. JAMA 2005;294(16): 2052–6. Guo B, Luo C, Xun C, Xie J, Wu X, Pu J. Quantitative detection of cytokeratin 20 mRNA in urine samples as diagnostic tools for bladder cancer by real-time PCR. Exp Oncol 2009;31(1):43–7. Soloway MS. Bladder tumor markers, intravesical therapy and systemic chemotherapy (Editorial). J Urol 2001;166: 488–9. Yossepowitch O, Herr HW, Donat SM. Use of urinary biomarkers for bladder cancer surveillance: patient perspectives. J Urol 2007;177(4):1277–82: discussion 1282. Bensalah K, Montorsi F, Shariat SF. Challenges of cancer biomarker profiling. Eur Urol 2007;52(6):1601–9.