Transitional Cell Carcinoma Patients Research Articles

Quantitation of rare circulating tumor cells by folate receptor α ligand-targeted PCR in bladder transitional cell carcinoma and its potential diagnostic significance.

Numerous attempts for detection of circulating tumor cells (CTC) have been made to develop reliable assays for early diagnosis of cancers. In this study, we validated the application of folate receptor α (FRα) as the tumor marker to detect CTC through tumor-specific ligand PCR (LT-PCR) and assessed its utility for diagnosis of bladder transitional cell carcinoma (TCC). Immunohistochemistry for FRα was performed on ten bladder TCC tissues. Enzyme-linked immunosorbent assay (ELISA) for FRα was performed on both urine and serum specimens from bladder TCC patients (n = 64 and n = 20, respectively) and healthy volunteers (n = 20 and n = 23, respectively). Western blot analysis and qRT-PCR were performed to confirm the expression of FRα in bladder TCC cells. CTC values in 3-mL peripheral blood were measured in 57 bladder TCC patients, 48 healthy volunteers, and 15 subjects with benign urologic pathologies by the folate receptor α ligand-targeted PCR. We found that FRα protein was overexpressed in both bladder TCC cells and tissues. The levels of FRα mRNA were also much higher in bladder cancer cell lines 5637 and SW780 than those of leukocyte. Values of FRα were higher in both serum and urine specimens of bladder TCC patients than those of control. CTC values were also higher in 3-mL peripheral blood of bladder TCC patients than those of control (median 26.5 Cu/3 mL vs 14.0 Cu/3 mL). Area under the receiver operating characteristic (ROC) curve for bladder TCC detection was 0.819, 95 % CI (0.738-0.883). At the cutoff value of 15.43 Cu/3 mL, the sensitivity and the specificity for detecting bladder cancer are 82.14 and 61.9 %, respectively. We concluded that quantitation of CTCs through FRα ligand-PCR could be a promising method for noninvasive diagnosis of bladder TCC.

Efficacy of radical cystectomy plus adjuvant intraarterial chemotherapy with gemcitabine and cisplatin on locally advanced bladder cancer.

Bladder cancer is the ninth most common cancer in the world; fewer than 15% of transitional-cell carcinoma patients survive 2 years if left untreated. Although radical cystectomy is the standard treatment of choice, much of them relapse and the necessity of adjuvant chemotherapy is still under debate. The aim of the study was to evaluate the efficacy of adjuvant intraarterial chemotherapy (IAC) with gemcitabine and cisplatin (GC) on locally advanced bladder cancer. This is a retrospective study on 60 patients with locally advanced bladder carcinoma who underwent radical cystectomy between May 2000 and June 2011. Patients were studied in two groups based on IAC and followed up for up to 5 years. Among 60 patients, there were 25 patients who underwent IAC (GC) after radical cystectomy (the IAC group) and 35 patients who underwent radical cystectomy alone (the control group). Although not significant, the relapse rates were slightly reduced in the IAC group than in the control group. Patients with IAC had a reduction in mortality compared with patients without IAC over 5 years. Specifically, IAC significantly reduced about 82% of mortality within the first year (hazard ratio = 0.18, 95% CI 0.03-0.97, P = 0.04). Additionally, IAC was well tolerated and safe. The most common adverse effect was transient myelosuppression (10/25, 40%), which was resolved by various medical treatments. Compared with radical cystectomy alone, radical cystectomy in combination with adjuvant IAC moderately but significantly reduces 1-year mortality. Our preliminary data showed only marginal benefit for the early survival. However, a randomized clinical study is needed to determine the long-term survival benefit.

Intravesical thermo-chemotherapy based on conductive heat: a first pharmacokinetic study with Mitomycin C in superficial transitional cell carcinoma patients

To evaluate, for the first time, the mitomycin C (MMC) pharmacokinetics during intravesical hyperthermia treatment based on conductive heat and the stability and recovery of the drug at the end of the instillation period. Eleven patients with recurrent intermediate-risk superficial transitional cell carcinoma of the bladder were treated weekly for six cycles with intravesical MMC (40 mg MMC in 50 ml) in local hyperthermia (45 °C) with Unithermia(®) system. Each instillation lasted 45 min, with the solution being replaced after the first 22 min. The MMC recovery at the end of the two instillation period and the plasmatic pharmacokinetics of MMC were evaluated by high-pressure liquid chromatography. Nine patients completed all the six planned cycles, whereas two patients missed the last cycle because of allergic reactions. No other systemic toxicity was observed, and the local toxicities were mild. Median MMC concentration in the instillation residual solution decreases from the initial 0.8 to 0.22 mg/ml for the 0-22-min instillation period and to 0.38 mg/ml for the 22-45-min instillation period; the median percentage of MMC recovered after instillation was 66.2 and 99.6, respectively. In all patients, MMC plasmatic C max resulted considerably lower than the toxic threshold (400 ng/ml). The MMC is stable during the instillation, and its absorption occurs mainly during the first minutes of the treatment. The plasmatic MMC concentration is always well below the threshold level for myelosuppression, as confirmed by the total lack of hematological toxicity evidenced by the patients. In order to evaluate the efficacy of the treatment performed with UniThermia(®) in reducing the disease recurrence rate in short- and long-term follow-up, we are currently carrying out a clinical multicentric study involving a larger number of patients.

A population-based study on the association between urinary calculi and kidney cancer

Using a nationwide population-based dataset and case-control study design, we investigate the association between urinary calculi (UC) and kidney cancer (KC) in Taiwan. The data for this case-control study were sourced from the Taiwan National Health Insurance program. The cases included 1308 incident patients pathologically diagnosed with KC. This study also used 6540 randomly selected subjects as controls. Conditional logistic regression was used to examine the associations between KC and patients previously diagnosed with UC. Of the sampled patients, 1262 (16.1 %) had previously been diagnosed with UC; 415 (31.7 % of the patients with KC) and 847 controls (13.0 % of patients without KC). After adjusting for monthly income, geographic location, urbanization level, hypertension, diabetes, renal disease, obesity, cystic kidney disease, tobacco use disorder, and alcohol abuse, we found that patients with KC were likely to have been previously diagnosed with UC than controls (odds ratio [OR] 3.18, 95% confidence interval [CI] 2.75-3.68, p < 0.001). In addition, the magnitude of the observed associations were stronger among females (females OR 3.59; 95% CI 2.87-4.48 vs. males OR 2.93, 95% CI 2.42-3.55) and transitional cell carcinoma patients (transitional cell carcinoma, OR 3.96; 95% CI 3.23-4.86 vs. renal cell carcinoma OR 2.76, 95% CI 2.31-3.29). We conclude that there is an association between KC and prior UC, especially in females and patients with transitional cell carcinoma.

Abstract B52: Targeting epigenetic dysregulation in a canine model of bladder cancer

Abstract Bladder cancer, which consists mainly of Transitional Cell Carcinoma (TCC), is the fifth most common cancer in the United States with over 70,000 cases and 15,000 deaths expected in 2013. Despite the successful development of molecularly targeted therapeutics for many cancers, there has been little change in the standard of care for TCC patients over the last few decades. New studies have revealed that aberrant epigenetic events, such as DNA hypermethylation and altered histone modifications, are common in TCC and are thought to promote tumorigenesis. Epigenetic therapies targeting these dysregulated pathways may reverse these effects and can have a profound influence on tumor growth. One such epigenetic modifier, EZH2, the catalytic subunit of polycomb repressive complex 2 (PRC2), is overexpressed in a variety of cancers and was found to enhance tumor progression by silencing several tumor suppressor genes. Recently, we have verified that EZH2 is upregulated in human TCC through an FGF2-NDY1/KDM2B-dependent suppression of miR-101. These data suggest that FGF2-NDY1/KDM2B-EZH2 pathway may be important in transitional cell carcinogenesis and may provide relevant epigenetic targets for the treatment of TCC. Spontaneous canine TCC also exhibits activation of FGF2 signaling and has been suggested as a model for bladder cancer. Therefore, we hypothesized that the development of aberrant epigenetic mechanisms downstream of FGF2 would be conserved between humans and dogs and that canine TCC could serve as a relevant large animal model for the clinical targeting of the FGF2-NDY1-EZH2 axis. To test our hypothesis, we employed canine TCC cell lines that we developed from dogs of various breeds and found that both EZH2 and NDY1/KDM2B are upregulated in canine TCC when compared to normal tissue. In order to target the pathway, we inhibited EZH2 both pharmacologically, using the commercially available inhibitor DZNep (3-deazaneplanocin), and through targeted knockdown. We determined that both DZNep and shRNA knockdown of EZH2 inhibited proliferation and induced apoptosis in canine TCC cells in anchorage-dependent (2D) and anchorage-independent (3D) cultures. By conducting these experiments in parallel with human TCC cell lines, we showed that sensitivity to DZNep is comparable between human and canine TCC cell lines. Western blot analysis and qRT-PCR demonstrated that, upon DZNep treatment, there is reduction of EZH2 at the protein level and inhibition of its activity, as evidenced by decreased global trimethylation of Histone H3 at lysine 27 (H3K27me3). Furthermore, inhibition of EZH2 was associated with upregulation of a set of genes that are commonly silenced by H3K27me3. Human and canine bladder cancer share histopathologic characteristics and clinical behavior. With this study, we confirm that the molecular underpinnings of spontaneous TCC are also shared between dogs and humans. Therefore canine TCC provides a means to understand epigenetic dysregulation in the urothelium as well as to investigate its clinical targeting. Citation Format: Parthena Foltopoulou, Monica Betancur-Boissel, Philip N. Tsichlis, Elizabeth A. McNiel. Targeting epigenetic dysregulation in a canine model of bladder cancer. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr B52.

Epithelial-mesenchymal transition markers in metastatic transitional cell carcinoma (mTCC) patients under vinflunine treatment.

e15532 Background: Vinflunine (VFL) demonstrated in vitro antiproliferative effects in human tumour cell lines, and in vitro antitumor activities against subcutaneous tumour xenografts. Recently, we demonstrated that VFL causes these effects through tubulin instability, which is one of the leading intracellular molecular process that drives epithelial-mesenchymal transition (EMT). In addition, VFL may induce up-regulation in expression levels of E-cadherin/N-cadherin cocient. Methods: The study included a retrospective analysis of 18 patients with mTCC treated with carboplatin-based chemotherapy. The patients who demonstrated objective response (38.8%) continued with vinflunine 320 mg/m2 one day every three weeks until progression. EMT protein expression (E-cadherin, beta-catenin, vimentin, CK18) was evaluated by Western blotting and also by immunohistochemistry (IHQ). Results: Overall mean age of patients was 71 years. The median overall survival was 3.5 years for the carboplatin-based chemotherapy (95% CI, 0.377-3.623). In patients treated with VFL, it was demonstrated a correlation of performance status and overall response with the histological type and age. We also found a relationship between different EMT markers in patients that received VFL. Conclusions: The above data provide evidence for an intrincate mechanism by which VFL could stop the transition from epithelial cells to the invasive mesenchymal phenotype. Molecular mechanism of Snail, Slug, Twist and Hakai will be reported.

Predictive markers in metastatic transitional cell carcinoma (mTCC) patients treated with carboplatin plus pemetrexed regimen.

e15558 Background: Treatment with platinum derivatives plus pemetrexed has shown a benefit in metastatic transitional cell bladder carcinoma (mTCC); however, the subset of patients most likely to benefit has not yet been identified. Predictive markers of pemetrexed response can be inferred from its mechanism of action in inhibiting nucleotide synthesis. In the present study we evaluated the predictive role of BRCA1, RAP80, TS, REV3 and AEG1 mRNA levels in a series of mTCC samples from patients treated with carboplatin plus pemetrexed (CP) and correlated with pathological response to chemotherapy and median survival. Methods: We retrospectively analyzed 18 patients with mTCC. Eligible patients received pemetrexed 500 mg/m2 in combination with carboplatin at an area under the concentration curve of 5. Total treatment dose was given on day 1 of each cycle and repeated every 21 days for 6 cycles. Gene expression levels were quantified by real-time quantitative PCR from paraffin-embedded pre-treatment tumor samples obtained by transurethral resection. Results: Patients were male, mean age 71 years (range 51-83). Median overall survival was 3.5 years (95% CI, 0.377-3.623). Non-responders had average survival of 2.2 years (SE=0.436) compared to an average of 5.5 years for responders (SE=0.661) (Pearson X2=4.667, p=0.031). We found an association between non-responders and elevated RAP80 expression (paired t (df)=-2.69, p=0.028) (paired t (df)=2.372, p=0.039). In the multivariate analysis elevated TS level was the variable most predictive of shorter survival (OR=1.158, p=0.047). Conclusions: CP regimen demonstrated moderate activity in patients with mTCC. Our data indicates that RAP80 and TS could be predictors of poor outcome. Also, RAP80 expression may predict efficacy of CP as first-line treatment and could be a useful tool to customize therapy in bladder cancer patients. A prospective study is warranted to validate these observations.

Long-term results of retroperitoneoscopic nephroureterectomy for upper urinary tract transitional cell carcinoma in China

Objective: We compared long-term clinical outcomes of upperurinary tract transitional cell carcinoma (TCC) patients treated byretroperitoneoscopic nephroureterectomy (RNU) or open radicalnephroureterectomy (ONU).Methods: Upper urinary tract TCC patients were treated with RNU(n = 86) or ONU (n = 72) and followed-up for more than threeyears. Demographic and clinical data, including preoperativeindexes, intraoperative indexes and long-term clinical outcomes,were retrospectively compared to determine long-term efficacy ofthe two procedures.Results: The RNU and ONU groups were statistically similar inage, gender, previous bladder cancer history, tumour location,pathologic tumour stage, pathologic node metastasis or tumourpathologic grade. The original surgery time required for both RNUand ONU was statistically similar, but RNU was associated witha significantly smaller volume of intraoperative estimated bloodloss and shorter length of postoperative hospital stay. Follow-up(average: 42.4 months, range: 3-57) revealed that the RNU 3-yearrecurrence-free survival rate was 62.8% and the 3-year cancerspecific survival rate was 80.7%. In the ONU group, the 3-yearrecurrence-free survival and the three-year cancer-specific survivalrates were 59.2% and 80.3%, respectively. Neither of the survivalrates were statistically different between the two groups. T stage,grade, lymph node metastasis and bladder tumour history wererisk factors for tumour recurrence; the operation mode and thebladder cuff incision mode had no correlation with the recurrencefreesurvival.Conclusion: The open surgery strategy and the retroperitoneoscopicnephroureterectomy strategy are equally effective for treating upperurinary tract TCC. However, the RNU procedure is less invasive,and requires a shorter duration of postoperative hospitalized care;thus, RNU is recommended as the preferred strategy.

Is sparing the prostate still considered radical cystectomy? The case against prostate-sparing cystectomy for bladder cancer

Radical cystoprostatectomy with bilateral pelvic lymphadenectomy and urinary diversion remains the gold standard therapy for localized bladder cancer in men.1 The procedure has been refined over several decades with the current application of nerve-sparing techniques.2,3 Unfortunately, the procedure carries significant risks of urinary and sexual dysfunction resulting in bladder-sparing approaches being popularized for selected early invasive bladder cancers. More recently, some authors have suggested that prostate-sparing cystectomy may be appropriate in selected men to improve postoperative sexual and urinary function.4–6 They propose that leaving the prostate, or part of it, intact will result in less damage to pelvic innervation and better preservation of sexual function and urinary sphincter control in cases of orthotopic diversion. These potential functional improvements must be weighed against any risks for oncologic outcomes. One potential oncologic pitfall is the assumption that the prostate does not harbour transitional cell carcinoma (TCC) of the bladder. The incidence of occult prostatic involvement with TCC in patients undergoing radical cystectomy ranges between 20% and 48%,7–9 raising significant concern about sparing the prostate. In a large study, Pettus and colleagues10 concluded that patients with tumours involving the trigone and bladder neck or associated carcinoma in situ of the bladder should be considered at high risk for developing prostatic urethral carcinoma. Moreover, the absence of these factors did not conclusively rule out the presence of carcinoma in the prostatic urethra. Another significant risk is the potential for prostatic adenocarcinoma. Many argue that prostate cancer is not a major problem in this population, since men undergoing cystectomy for invasive bladder cancer are often elderly, with a poor prognosis for their bladder cancer. However, prostate-sparing surgery is usually offered to men whose pelvic disease is not advanced and in whom sexual function is normal, thus younger men. It is precisely in these younger men with early forms of invasive bladder cancer that occult prostate cancer could pose problems in the future. Occult prostate cancer has a reported incidence of 23% to 47% in men undergoing radical cystectomy. Furthermore, about 50% of these cancers are potentially significant based on histologic criteria.11–14 Revelo and colleagues11 detected that 41% of cases had unsuspected prostatic cancer, of which 48% were clinically significant and 16% were extracapsular. Of importance, 60% involved the apex, including 79% of significant cases, suggesting that recommending apical sparing as a compromise could potentially be equally hazardous. These findings were confirmed by another study in which 27% of patients undergoing cystectomy for invasive TCC of the bladder were found to have prostatic carcinoma (PCa), with 33% involving the apex of the prostate.15 The authors suggested that the high likelihood of apical prostate cancer requires careful excision of the apical margins. Kablain and colleagues13 previously reported that 38% of patients undergoing cystectomy for bladder cancer and who harbour prostate carcinoma had a distinct apical predominance of the tumours. In a recent study, Hautmann and colleagues16 addressed the probability of overlooking clinically significant prostate cancer using sextant biopsy. They analyzed specimens from 133 consecutive patients undergoing cystoprostatectomy for bladder cancer. Patients were included in the study if they had serum prostate-specific antigen (PSA) 25%. Systematic sextant biopsy was performed in the operating room immediately following cystoprostatectomy and prostates were subsequently step-sectioned (3 mm). The threshold defined to distinguish between significant and insignificant cancer was 0.5 cm3. Incidental prostatic cancer was found in 58 of the 133 bladder cancer patients (44%), with presence of significant cancer in many. Sextant biopsy detected 7 cancers, including 4 of 47 (9%) that were insignificant and 3 of 11 (27%) that were significant. Thus, sextant biopsy missed 8 (73%) of 11 of significant prostate cancers. These data suggest that despite careful screening for prostate cancer for cystoprostatectomy patients, it remains regularly overlooked in a significant proportion of clinically relevant cases. In a recent prospective study by Saad and colleagues,17 425 men had radical cystoprostatectomy for invasive bladder cancer and entire mounted prostates were step-sectioned at 2 to 3 mm intervals. Of 425 cases examined, there was histological evidence of prostatic adenocarcinoma in 90 cases (21.2%). Prostatic TCC was found in 66 patients (15.6%). There was also significant benign prostatic hyperplasia in 175 (41.2%). They concluded that the marginal reported functional advantage of prostate-sparing cystectomy is not worth the potential oncological risks. In a prospective study, Koraitim and colleagues18 studied urinary continence and voiding differences between men undergoing classic radical cystoprostatectomy and men with prostate-sparing cystectomy. They found no statistical difference between both groups with respect to urinary continence, but found a significant difference in peak flow rate. They concluded that preservation of prostatic apex may present an element obstructing the evacuation of ileal bladders.

Role of flexible cystoscopy and ultrasound in the detection of recurrent Bladder Tumour

Background: Rigid cystoscopy under anaesthesia for the surveillance of recurrent bladder tumour creates a large urological workload. Recently, flexible cystoscopy became a popular alternative and an easy, safe and effective means of check cystoscopy in following-up patients of superficial transitional cell carcinoma (TCC) of the urinary bladder. Because of the frequency of tumor recurrences and the necessity of finding, whether a less invasive, easily repeatable investigation is capable of providing precise information about the bladder cavity, and, could decrease the frequency of repeated rigid cystoscopy under anaesthesia and inpatient admission; we have decided to perform this study. Methods: This is a prospective comparative study that involved 85 patients (70 male and 15 female) with a mean age of 61 years (41-80 years), who had undergone one or more transurethral resections for TCC of bladder (stage pTa and pT1; grade I and II.) in the department of Urology, BSMMU between July 2005 -Feb 2007. Ultrasonography(USG) of the bladder was performed one week before check cystoscopy. We have calculated sensitivity and accuracy of USG and flexible in comparison to rigid cystoscopy.Results: Eighty five (85) sessions of follow-up investigations- Ultrasound and flexible cystoscopy showed 31 recurrences confirmed by rigid cystoscopy and biopsy. In over 85 rigid cystoscopies, 54 were negative and 31 were positive for tumour. Sensitivity, the most important parameter, was 97% for the two examinations together. Each method separately had the following sensitivity: ultrasound 77%; flexible cystoscopy 90%.Conclusion: Considering that there was only one false-negative result of combined abdominal ultrasound and flexible cystoscopy; with this follow-up scheme we could have saved our patient from rigid cystoscopies, reducing the cost of in-patient admission and anaesthesia.DOI: http://dx.doi.org/10.3329/kyamcj.v3i1.13655 KYAMC Journal Vol. 3, No.-1, June 2012 pp.214-219

GSTM1-null and GSTA1-low activity genotypes are associated with enhanced oxidative damage in bladder cancer

ObjectivesTo examine the association between gene variants of the detoxifying and antioxidant enzymes glutathione transferase M1 (GSTM1) and glutathione transferase A1 (GSTA1) and the extent of oxidative damage in patients with transitional cell carcinoma (TCC) of the urinary bladder.MethodsGSTM1 deletion polymorphism was identified by polymerase chain reaction, and the restriction fragment length polymorphism method was used for the single nucleotide polymorphism of GSTA1. Enzyme immunoassay was used to determine markers of DNA (8-hydroxy-2′-deoxyguanosine, 8-OHdG) and lipid (8-epiprostaglandin F2α) oxidative damage in the urine of 80 TCC patients and 60 age-matched controls.ResultsUrinary 8-OHdG and 8-epi-prostaglandin F2α concentrations in TCC patients were significantly higher than in controls (P = 0.043 and 0.001, respectively). GSTM1 and GSTA1 polymorphisms influence vulnerability to both DNA and lipid oxidation, with the GSTM1-null gene variant having a more pronounced effect. A significant effect of combined GSTM1 and GSTA1 genotypes on the extent of oxidative damage was found only for 8-OHdG (P = 0.018). In addition, TCC patients with the most malignant tumors exhibited significantly higher frequencies of GSTM1-null or GSTA1-low activity genotypes, associated with a twofold increase in urinary 8-OHdG concentration (P = 0.044).ConclusionsOur results suggest that absent GSTM1 or reduced GSTA1 antioxidant activity may increase the accumulation of oxidative DNA damage, thereby contributing to the malignant potential of TCC.

Endoscopic Versus Laparoscopic Management of Noninvasive Upper Tract Urothelial Carcinoma: 20-Year Single Center Experience

We compare the outcomes of endoscopic surgery to laparoscopic nephroureterectomy for the management of specifically noninvasive upper tract urothelial carcinoma. A retrospective database review identified consecutive patients with clinically noninvasive upper tract urothelial carcinoma who underwent endoscopic surgery (59, via ureteroscopic ablation or percutaneous resection) or laparoscopic nephroureterectomy (70) at a single center during 20 years (1991 to 2011). Overall survival, upper tract urothelial carcinoma specific survival, upper tract recurrence-free survival, intravesical recurrence-free survival, progression-free survival and renal unit survival were estimated using Kaplan-Meier methods, with differences assessed using the log rank test. Median age and followup were 74.8 years and 50 months, respectively. Overall renal preservation in the endoscopic group was high (5-year renal unit survival 82.5%), although this came at a cost of high local recurrence (endoscopic surgery 5-year recurrence-free survival 49.3%, laparoscopic nephroureterectomy 100%, p <0.0001). For G1 upper tract urothelial carcinoma, endoscopic surgery 5-year disease specific survival (100%) was equivalent to that of laparoscopic nephroureterectomy (100%). However, laparoscopic nephroureterectomy demonstrated superior disease specific survival to endoscopic surgery for G2 disease (91.7% vs 62.5%, p = 0.037) and superior progression-free survival for G3 disease (88.9% vs 55.6%, p = 0.033). For G1 upper tract urothelial carcinoma, endoscopic management can provide effective oncologic control and renal preservation. However, endoscopic management should not be considered for higher grade disease except in compelling imperative cases or in patients with poor life expectancy as oncologic outcomes are inferior to those of laparoscopic nephroureterectomy.

Long-term results of retroperitoneoscopic nephroureterectomy for upper urinary tract transitional cell carcinoma in China

We compared long-term clinical outcomes of upper urinary tract transitional cell carcinoma (TCC) patients treated by retroperitoneoscopic nephroureterectomy (RNU) or open radical nephroureterectomy (ONU). Upper urinary tract TCC patients were treated with RNU (n = 86) or ONU (n = 72) and followed-up for more than three years. Demographic and clinical data, including preoperative indexes, intraoperative indexes and long-term clinical outcomes, were retrospectively compared to determine long-term efficacy of the two procedures. The RNU and ONU groups were statistically similar in age, gender, previous bladder cancer history, tumour location, pathologic tumour stage, pathologic node metastasis or tumour pathologic grade. The original surgery time required for both RNU and ONU was statistically similar, but RNU was associated with a significantly smaller volume of intraoperative estimated blood loss and shorter length of postoperative hospital stay. Follow-up (average: 42.4 months, range: 3-57) revealed that the RNU 3-year recurrence-free survival rate was 62.8% and the 3-year cancer specific survival rate was 80.7%. In the ONU group, the 3-year recurrence-free survival and the three-year cancer-specific survival rates were 59.2% and 80.3%, respectively. Neither of the survival rates were statistically different between the two groups. T stage, grade, lymph node metastasis and bladder tumour history were risk factors for tumour recurrence; the operation mode and the bladder cuff incision mode had no correlation with the recurrence-free survival. The open surgery strategy and the retroperitoneoscopic nephroureterectomy strategy are equally effective for treating upper urinary tract TCC. However, the RNU procedure is less invasive, and requires a shorter duration of postoperative hospitalized care; thus, RNU is recommended as the preferred strategy.

Expression of CD147, BIGH3 and Stathmin and their potential role as diagnostic marker in patients with urothelial carcinoma of the bladder

BackgroundUrothelial carcinoma of the bladder is characterised by very high recurrence rate, followed up by cystoscopy which being invasive technique makes the need for non-invasive markers important for Transitional Cell Carcinoma (TCC) detection. CD147 is a transmembrane protein highly expressed in tumour cells which aids in tumour invasion and growth. BIGH3, an Extracellular matrix protein (ECM) which interacts with various ECM component in different tissue system and Stathmin(STMN1) is cytosolic microtubule destabilising protein also called as Oncoprotein18 due to its role in tumour promotion. So far the expression of BIGH3 and STMN1 remains undetermined in cancer subjects including TCC. We therefore studied the levels and molecular expression of these molecules in TCC patients, to evaluate their usefulness as diagnostic markers. MethodsThirty consecutive TCC patients and two sets of control- 15 Benign prostatic hyperplasia (BPH) patient and 15 healthy were taken. Serum and urine levels of these molecules were estimated by ELISA and relative mRNA expression by Q-PCR from tumour and normal urothelium. Post-Hoc analysis and ROC curve were determined to evaluate the significance and sensitivity and specificity. ResultsThe mean concentrations of these molecules were found to be significantly increased (p<0.001) in the serum and urine of TCC patients, with varying significance in each grade for different molecules. The urinary levels of CD147 (67pg/ml) and serum STMN1 concentration (1.38ng/ml) showed a specific increase as compared to the controls, while BIGH3 was elevated in both serum and urine samples. Molecular (mRNA) expression was elevated in the high grade (Muscle Invasive) stage of the disease for all the molecules, with a significant 3-fold increase that correlated with disease severity being observed for STMN1. ROC analysis gave optimal combination of sensitivity and specificity for diagnosis of the disease in urine and serum sample for STMN1. ConclusionOf CD147, BIGH3 and STMN1, significant results were obtained for STMN1 and it could serve as the best possible diagnostic marker for TCC detection in future.

Abstract LB-478: MiRNA as biomarkers to predict de novo chemoresistance to neoadjuvant chemotherapy in patients with muscle invasive bladder cancer

Abstract The primary goal of this study was to determine whether miRNA expression profiling can predict de novo chemoresistance to treatment with neoadjuvant chemotherapy in patients with muscle invasive bladder cancer (MI-BC). The relative expression of 9 miRNAs in 41 annotated, diagnostic MI-BC patient samples was determined by quantitative real-time PCR (qPCR) and correlated with patient response to neoadjuvant chemotherapy, p53 status (SSCP/sequencing) and pRb, p21 expression (IHC). These 9 miRNA were chosen based on a known link to either transitional cell carcinoma (TCC) progression and/or chemoresistance. Expression of these miRNA was also assessed in 8 paired pre- versus post-chemotherapy patient samples from non-responders. Patients with higher let-7c expression levels had significantly higher odds of responding to neoadjuvant chemotherapy (p = 0.023, OR 2.493, 95% CI 1.121, 5.546). The odds of response was not significantly different between patients with vs. without p53 mutations (p = 0.948, OR 0.955, 95% CI 0.231, 3.945). Post-chemotherapy samples expressed significantly higher levels of let-7c (geometric mean ratio = 3.5, P = 0.006). Lower let-7c expression levels appear to predict chemoresistance in TCC patients, prospective studies will be needed to validate this finding. Pre- versus post-chemotherapy patient sample analyses indicate increased let-7c expression plays a role in mediating disease progression following treatment with chemotherapy. Future studies will focus on determining the functional role played by let-7c in chemoresistance and disease progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-478. doi:1538-7445.AM2012-LB-478

Abstract LB-86: Metabolic presentation of bladder cancer in urine

Abstract Introduction and Objective Recently it has been shown that many oncogenes regulate metabolism, revitalizing interest in Warburg's observation of aerobic glycolysis by tumors. We and others have demonstrated the utility of metabolic signatures for the diagnosis of cancer in minimally invasive clinical samples and the differential diagnosis of cancer by stage and grade using biopsy samples. Here we apply state of the art metabolomic analysis to the diagnosis of bladder cancer from urine samples. Methods A case controlled study of ten transitional cell carcinoma (TCC) patients and matched non-TCC controls was performed. Briefly, clarified urine was extracted into aqueous methanol and characterized using three analytical platforms (gas chromatography mass spectrometry (MS) and ultrahigh performance liquid chromatography tandem MS). Spectral data were compared against an in-house library of over 2500 authentic standards and the relative abundance of each compound within the data set was calculated. Two-group statistics (R) and multivariate analysis (Array Studio, OmicSoft) were performed to identify differentially abundant metabolites. Results Three hundred and sixty-seven compounds were identified in this study and polypharmacy did not compromise the quality of the signal obtained for endogenous metabolites. Thirteen compounds, arising from diverse metabolic pathways, including: energetics, oxidative stress and amino acid catabolism, exhibited statistically significant differences between TCC and non-TCC cases. Principal component (PC) analysis demonstrated that the majority of the information in these metabolites (PC 1) separated TCC from non-TCC cases. Conclusions These data indicate the feasibility of a novel, non-invasive, urine-based diagnostic test to detect the presence of bladder tumors. Such a test could be positioned for use in conjunction with the current standard of care to improve patient management. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-86. doi:1538-7445.AM2012-LB-86

LASP-1, a Novel Urinary Marker for Detection of Bladder Cancer

ObjectivesThe LIM and SH3 (LASP)-1 protein is a focal adhesion protein that has been linked to oncogenesis. The aim of this study was to evaluate the diagnostic use of the detection of LASP-1 in tumor specimens and in urine for noninvasive detection of transitional cell carcinoma (TCC). Materials and methodsImmunohistochemical staining for LASP-1 was performed on 72 archived bladder tumor specimens, and LASP-1 content was measured in 132 spontaneous urine sample sediments by Western blot analysis. ResultsIn the histologic specimens, immunohistochemical staining for LASP-1 showed abundant expression throughout the urothelium of the bladder and ureter. However, modest overexpression of LASP-1 was observed in the TCC specimens. Measurement of the LASP-1 protein concentrations in urinary cell pellets from healthy donors and bladder cancer patients was highly sensitive for the presence of TCC. The cut-off value was determined by receiver operating characteristic (ROC) analysis. When a cut-off value of 1 ng LASP-1/500 μl of urine was used, the sensitivity, specificity, and positive and negative predictive values of the assay were 83.1%, 85.3%, 83.1%, and 80.6%, respectively.The increased urinary LASP-1 content in TCC patients was attributable in part to a specific increase in cell shedding presumably caused by changes in cell adhesion, as confirmed by LASP-1 knockdown. Contamination with erythrocytes above 250 cells/μl and urinary infection gave false positive results and are therefore sample exclusion criteria. ConclusionsIn the absence of urinary infection or gross hematuria, urinary LASP-1 level is a promising marker for transitional cell carcinoma.

Prognostic impact of expression Notch-1 in invasive bladder transitional cell carcinoma.

299 Background: Notch signal pathway plays a fundamental role in mediator of tumorigenesis, either as a tumor promoter or suppressor, depending on cellular context and expression levels; we investigated the prognosis of invasive bladder transitional cell carcinoma patients with over-expression of Notch-1. Methods: Tumor tissue samples from 78 resected patients with invasive bladder transitional cell carcinoma were obtained, and immunohistochemistry was used to evaluate the expression of the molecular markers Notch-1 and Jagged-1. To further confirm the immunohistochemical results tissue samples were subjected to RT-PCR and Western blot analysis, staining evaluation results were analyzed statistically in relation to various clinicopathological characters. Kaplan–Meier and Cox proportional hazards regression methods were used to assess associations of Notch-1 expression with invasive bladder transitional cell carcinoma survival. Results: Survival time in patients with high expression of Notch-1 and Jagged-1 was significantly shorter than that in patients with low expression p=0.009 and more recurrence p=0.01, in multivariate analysis, Notch1 expression was proved to be an independent predictor of prognosis; (hazard ratio Notch-1 was 1.93 (95% CI, 1.02-3.03; P =0.014), expression was not related to patient sex or age. Conclusions: Notch-1 expression is associated with tumor progression, indicating that Notch-1 may be involved in the carcinogenesis and progression of invasive bladder transitional cell carcinoma. These findings also suggest that Notch-1 may be a potential diagnostic marker and therapeutic target in patients.

Antioncogenic Effects of Transient Receptor Potential Vanilloid 1 in the Progression of Transitional Urothelial Cancer of Human Bladder

The progression of normal cells to a tumorigenic and metastatic state involves the accumulation of mutations in multiple key signaling proteins, encoded by oncogenes and tumor suppressor genes. Recently, members of the TRP channel family have been included in the oncogenic and tumor suppressor protein family. TRPM1, TRPM8, and TRPV6 are considered to be tumor suppressors and oncogenes in localized melanoma and prostate cancer, respectively. Herein, we focus our attention on the antioncogenic properties of TRPV1. Changes in TRPV1 expression occur during the development of transitional cell carcinoma (TCC) of human bladder. A progressive decrease in TRPV1 expression as the TCC stage increases triggers the development of a more aggressive gene phenotype and invasiveness. Finally, downregulation of TRPV1 represents a negative prognostic factor in TCC patients. The knowledge of the mechanism controlling TRPV1 expression might improve the diagnosis and new therapeutic strategies in bladder cancer.

Fluorescence in situ hybridization assay detects upper urinary tract transitional cell carcinoma in patients with asymptomatic hematuria and negative urine cytology

We evaluated the performance of a multiprobe FISH (fluorescence in situ hybridization) assay for noninvasive detection of upper urinary tract transitional cell carcinoma (UUT-TCC) in patients with asymptomatic hematuria and negative urine cytology. Voided urine samples from 285 patients with asymptomatic hematuria and negative urine cytology were prospectively analyzed by FISH technique. FISH assays were performed to detect chromosomal changes frequently associated with TCC, including aneuploidy of chromosomes 3, 7 and 17, and loss of the 9p21 locus. Eleven (3.9%) had a positive FISH result. Of the 11 patients, nine (81.8%) were found to have a TCC of the upper urinary tract, while no patients with negative FISH findings were found to have UUT-TCC. In this selected cohort, the sensitivity and specificity of FISH for the detection of UUT-TCC was 100% and 99.3%, respectively. Our preliminary data suggest that the clinical utility of FISH assay of chromosomes 3, 7, 9, and 17 as a noninvasive ancillary test for the diagnosis of UUT-TCC in a selected patient population with asymptomatic hematuria and negative urine cytology and by significant high sensitivity and specificity may be a reliable diagnostic approach for early detection of UUT-TCC patients. Further larger prospective and multicenter trials are needed to confirm our results.