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Objective: To evaluate whether differences exist between men and women in response to intravesical BCG treatments. The incidence of urothelial carcinoma of the bladder is lower in women but they tend to present with more aggressive and advanced disease. Some prior studies also suggest there are sex-based differences in response to treatment for non-muscle invasive bladder tumors., Methods: In this retrospective study, we reviewed all consecutive patients who received BCG at the CHU de Québec - Laval University from 2009-2019. Men and women were treated with intravesical BCG therapy following pathologic confirmation of urothelial carcinoma. Outcomes evaluated include recurrence, progression, and treatment tolerability. Recurrence was defined as a pathology confirmed cancer whereas progression was the new development of high-grade (recurrence) pathology or an increase of stage. Tolerability was defined according to the proportion of prescribed BCG received. All clinical details were obtained through review of the medical records, collaborated by pharmacy records for BCG administration. Competing-risk analysis was used to compare outcomes., Results: Among 613 patients who received BCG at our institution between 2009-2019, 472 (77.0%) were men and 141 (23.0%) were women. The recurrence rate was not different between sexes, with a 5-year recurrence risk of 52% (95% CI: 36.93-65.4) among women compared to 57.5% (CI 95%: 51.9-62.6) among men. The overall non-progression rate at 1,3 and 5 years was 97.3% (95% CI: 95.6%-98.3%), 93.6% (95% CI: 91.2%-95.4%), and 91.7% (95% CI: 88.4%-94.1%), respectively. The completion of ≥5 induction BCG instillations and maintenance BCG use was similar in both genders., Conclusions: We report a contemporary NMIBC cohort treated with BCG and find no clear evidence for sex-based differences in response to BCG treatment in regard of progression, recurrence, and tolerability., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Purpose: The impact of sex hormones on cancer immunotherapy remains controversial. Androgens, via the androgen receptor (AR), may impact the success of immune checkpoint blockade. This study characterizes AR and programmed death ligand-1 (PD-L1) expression in bladder tumors with long clinical follow-up., Methods: AR and PD-L1 expression was analyzed using immunohistochemistry on 143 transurethral resection (TUR) and 203 radical cystectomy (RC) specimens. Descriptive statistics and survival analyses assessed the relationship of AR and PD-L1 staining with clinical outcomes of tumor recurrence, progression, and overall survival., Results: AR expression was observed in a higher proportion of TUR than RC specimens (59% vs 35%, p < 0.001). High immune cell (IC) PD-L1 expression was associated with higher stage and grade. Patients with the combination of an absence of AR expression and the highest (> 10%) IC PD-L1 expression in TUR tumors had an increased risk of recurrence and progression. In RC specimens, the expression of AR increased the risk of local recurrence (adjusted hazard ratio (HR) 2.09, 95% CI 0.98-4.45), which was even higher among patients who also had IC PD-L1 expression (HR 4.16, 95% CI 1.28-13.52). For 28 paired metastatic lymph nodes among RC patients, tumor cell PD-L1 expression was significantly correlated (r = 0.48, p = 0.01), while no relationship with IC PD-L1 expression was observed., Conclusions: The expression of AR and its relationship to clinical outcomes appears to vary between non-muscle invasive and muscle-invasive bladder cancer. Our results support the role of IC PD-L1 expression as an independent risk factor for bladder cancer outcomes.

Purpose: To assess whether receipt of 5-alpha reductase inhibitors (5-ARIs) influences the findings on surgical pathology at the time of radical cystectomy (RC) and subsequent clinical outcomes. 5-ARIs may slow the progression of non-muscle-invasive bladder cancer., Patients and Methods: We retrospectively reviewed all patients who underwent RC at our institution between 2009 and 2017. Men were included who had urothelial cancer in the RC specimen. Patients with nonurothelial pathology or who had no cancer in the specimen were excluded. Odds ratios for pathologic features and hazard ratios for survival were adjusted for baseline patient characteristics and disease stage., Results: Our study cohort included 338 men; 48 patients (14%) were receiving dutasteride or finasteride at time of RC, 58 (17%) metformin, and 195 (58%) statins. Among patients receiving 5-ARIs, there was a lower proportion of positive margins (P = .08) and lymphovascular invasion (P = .05). This was statistically significant when patients with urothelial carcinoma variants were excluded. Multivariable logistic regression analysis demonstrated that 5-ARI receipt was associated with a lower odds ratio (OR) for the presence of lymphovascular invasion (OR = 0.49; 95% confidence interval, 0.24-1.00; P = .049) and positive surgical margins (OR = 0.30; 95% confidence interval, 0.09-1.07; P = .063). Further, 5-ARI receipt was associated with better overall survival, with an adjusted hazard ratio of 0.40 (95% confidence interval, 0.19-0.83; P = .015). No similar tendencies were observed with metformin or statins., Conclusion: 5-ARIs may exert a protective biologic effect on the invasive properties of high-grade urothelial carcinoma. Further research is needed to understand the therapeutic implications., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: To assess the rate of urinary tract infection (UTI), the characteristics of the bacterial aetiological agents involved, the type and duration of antibiotics used, and the clinical risk factors of UTI in a multi-institutional cohort of patients who had undergone radical cystectomy (RC)., Patients and Methods: The pre- and postoperative characteristics of patients who had undergone open RC at 1 of 3 institutions between 2009 and 2015 were analyzed by means of the patient charts. Patients were classified according to the presence or absence of UTI. Analysis of the severity of UTI was based on the EAU/EAU Section of Infections in Urology (ESIU) classification system. The bacterial aetiological agents and their antibiotic susceptibility were also assessed. Factors predicting postoperative UTI were identified using univariable and multivariable logistic regression analysis., Results: Of 217 patients, 42 (19.4%) had developed postoperative UTI, of whom 50% had urosepsis or uroseptic shock. Multivariable analysis showed continent urinary derivation as the only significant predictor of UTI with an odds ratio of 5.03 (95% confidence interval 2.12-11.9, P < 0.001). The duration of perioperative antibiotic prophylaxis was not associated with an increased risk of UTI. Enterococcus was the most commonly isolated bacteria (25.7%), but this species is not covered by the recommended antibiotic prophylaxis., Conclusion: Patients with continent urinary diversion after RC have a significantly higher risk of developing UTI. Prolonged perioperative administration of antibiotics does not seem to reduce the risk of UTI. Enterococcus as the most commonly isolated bacteria is not covered by most recommended antibiotic prophylaxis regimens. Therefore different antibiotic regimens should be considered for high-risk patients., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Survival in patients with bladder cancer has only moderately improved over the past 2 decades. A potential reason for this is nonadherence to clinical guidelines and best practice, leading to wide variations in care. Common quality indicators (QIs) are needed to quantify adherence to best practice and provide data for benchmarking and quality improvement., Objective: To produce an evidence- and consensus-based list of QIs for the management of bladder cancer., Methods: A modified Delphi method was used to develop the indicator list. Candidate indicators were extracted from the literature and rated by a 27-member Canadian expert panel in several rounds until consensus was reached on the final list of indicators. In rounds with numeric ratings, a frequency analysis was performed., Results: A total of 86 indicators were rated, 52 extracted from the literature and 34 suggested by the panel. After iterative rounds of ratings and discussion, a final list of 60 QIs spanning several disciplines and phases of the cancer care continuum was developed., Conclusions: This is the first study to comprehensively produce common QIs representing structure, process, and outcome measures in bladder cancer management. Though developed in Canada, these indicators can be used in other countries with slight modifications to track performance and improve care., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Cigarette smoking is the most important known risk factor for urinary bladder cancer. Selected arylamines in cigarette smoke are recognized human bladder carcinogens and undergo biotransformation through several detoxification pathways, such as the glutathione S-transferases (GST), and uridine-diphospho-glucuronosyltransferases (UGT) pathways. GSTM1 deletion status and UGT1A1*28 rs8175347 genotypes were assessed in 189 non-muscle-invasive bladder cancers (NMIBC) patients with pTa (77.2%) and pT1 (22.8%) tumors and a mean follow-up of 5.6 years, to investigate whether two common functional polymorphisms in GSTM1 and UGT1A1 genes and smoking history are associated with recurrence-free survival of patients with NMIBC. Most patients were current (48.7%) or previous (35.4%) cigarette smokers and 15.9% never smoked. Tumor recurrence occurred in 65.1% of patients, at a median time of 12.9 months. Upon multivariate analysis, previous and current smokers approximately tripled their risk of recurrences [HR = 2.76; 95% confidence interval (CI), 1.03-7.40 and HR = 2.93; 95% CI, 1.08-7.94, respectively]. When adjusted for age, smoking status, stage, grade, gender, and presence of carcinoma in situ, carriers of GSTM1 (+/- and -/-) and UGT1A1*28/*28 alleles were significantly at risk of NMIBC recurrence (HR = 10.05; 95% CI, 1.35-75.1 and HR = 1.91; 95% CI, 1.01-3.62, respectively). Compared with nonsmokers with UGT1A1*1/*1 and *1/*28 genotypes, previous and current smokers homozygous for the UGT1A1*28 allele demonstrated a risk of recurrence of 4.95 (95% CI, 1.02-24.0) and 5.32 (95% CI, 2.07-13.7), respectively. This study establishes a connection between GSTM1, UGT1A1, and tobacco exposure as prognostic markers of NMIBC recurrence in bladder cancer patients. These findings warrant validation in larger cohorts., (©2015 American Association for Cancer Research.)

Aim: We determined whether the Y-box binding protein-1 (YB-1) and its binding partner, the X-linked ribosomal protein S4 (RPS4X), are associated with clinical outcome in bladder cancer., Materials & Methods: A population of 167 patients with muscle-invasive bladder tumor without evidence of metastasis at time of cystectomy was analyzed retrospectively. YB-1 and RPS4X expressions were evaluated immunohistochemically in tumors and analyzed for association with clinical variables and survival., Results: Kaplan-Meier and multivariate Cox regression analyses indicated that low expression of RPS4X was associated with a higher risk of death or disease recurrence. In contrast, YB-1 was not significantly associated with either recurrence-free or overall survival., Conclusion: Low RPS4X expression is associated with poor disease-specific and recurrence-free survival in bladder cancer.

Purpose: To evaluate the impact of concomitant carcinoma in situ (CIS) on upstaging and outcome of patients treated with radical cystectomy with pelvic lymph node dissection., Methods: We collected and pooled a database of 1,968 patients who have undergone radical cystectomy between 1998 and 2008 in eight academic centers across Canada. Collected variables included patient's age, gender, tumor grade, histology and the presence of concomitant CIS with either cTa-1 or cT2 disease, dates of recurrence and death., Results: In the presence of concomitant CIS, upstaging following radical cystectomy occurred in 48 and 55 % of patients with cTa-1 and cT2 disease, respectively. On univariate analysis, the presence of concomitant CIS with cT2 disease was associated with upstaging (p < 0.0001), and the presence of concomitant CIS with cTa-1 disease was also associated with upstaging but did not reach statistical significance (p = 0.0526). On multivariate analyses, the presence of concomitant CIS with either cTa-1 or cT2 tumors was independently prognostic of disease upstaging (p = 0.0001 and 0.0186, respectively). However, on multivariate analysis that incorporates pathologic stage, concomitant CIS was not significantly associated with worse overall, recurrence-free or disease-specific survival., Conclusion: These results demonstrate that while the presence of concomitant CIS on cystectomy specimens does not independently affect outcomes, its presence is significantly predictive of a higher rate of upstaging at radical cystectomy.

Objective: To evaluate risk factors for bladder cancer recurrence in a cohort of patients treated with radical nephroureterectomy (RNU)., Patients and Methods: At 10 Canadian University Centers, we retrospectively evaluated data, between 1990 and 2010, from 743 patients who were free from bladder cancer and were previously treated with RNU for upper tract urothelial cancer., Results: Of 743 patients, 167 (22.5%) developed bladder tumors after a median time of 17.2 months after RNU. Multivariable analysis detected age (hazard ratio [HR] = 1.028; 95% CI: 1.010-1.046; P = 0.0018), tumor location in both the renal pelvis and the ureter (HR = 2.205; 95% CI: 1.355-3.589; P = 0.0015), the use of adjuvant systemic chemotherapy (HR = 2.309; 95% CI: 1.439-3.705; P = 0.0005), and laparoscopic surgery (HR = 1.876; 95% CI: 1.226-2.87; P = 0.0037) as risk factors for bladder cancer recurrence. Open excision of a bladder cuff (HR = 0.661; 95% CI: 0.453-0.965; P = 0.0319) and transurethral resection of the intramural ureter (HR = 0.548; 95% CI: 0.306-0.981; P = 0.0429) on comparison with extravesical resection decreased the risk of bladder cancer recurrence significantly. Major limitations were the retrospective design and partially missing data, although the significance of variables did not change in the imputation analysis., Conclusion: Older patients, those with tumor location in both the renal pelvis and the ureter, and those treated with adjuvant systemic chemotherapy were found at higher risk for intravesical recurrence, as were those having undergone extravesical ureterectomy or laparoscopic RNU., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Purpose: To compare clinical and pathologic outcomes of radical cystectomy for muscle invasive bladder cancer in relation to prior history of non-invasive urothelial carcinoma., Materials and Methods: Retrospective data collected from 1,150 patients managed by radical cystectomy for urothelial carcinoma of the bladder from the Canadian Bladder Cancer Network were analysed. Patients with clinical stage T2 or more were included and divided into two groups: (Group 1) patients with prior history of non-invasive urothelial carcinoma (N = 365) and (Group 2) patients with clinical muscle invasive cancer de novo (N = 785). Variables analysed included patient age, gender, pathologic stage, adjuvant chemotherapy, recurrence and mortality., Results: Both groups were nearly equal in mean age and gender distribution, with mean ages of 67.2 and 66.7 years, and 79.7 and 79.5%, respectively (P = 0.4 and 0.9, respectively). The presence of preoperative hydronephrosis was 20.8 and 32.6% (P = 0.0007) for groups 1 and 2, respectively. The rate of higher pathological stage (T3 or T4) was 36.3 and 58% (P < 0.0001), positive lymph nodes were 20.1 and 28.8% (P = 0.002), and lymphovascular invasion was 31.7 and 46.2% (P = 0.0001) for groups 1 and 2, respectively. The rate of adjuvant chemotherapy was 15.5 and 23.3% (P = 0.002) for groups 1 and 2, respectively. None of the sampled patients received neoadjuvant chemotherapy. The overall survival (OS) and disease-specific survival (DSS) rates at 5 years were 62 and 70% for group 1 and 51 and 60% for group 2, respectively, while at 10 years, OS and DSS were 46 and 66% for group 1 and 35 and 49% for group 2, respectively (P = 0.0001 and 0.0002, respectively). Using multivariate analysis examining factors affecting recurrence and survival, we found that previous non-invasive bladder tumour history was associated with a significantly reduced risk of mortality and recurrence (Hazard ratio of 0.7 for all risks, P = 0.0002)., Conclusion: Our retrospective study suggests that patients with non-invasive urothelial carcinoma of the bladder that progress to muscle invasion and require radical cystectomy appear to have better pathologic and clinical outcome than patients presenting with clinical muscle invasive disease de novo.

Objectives: The role of advanced age as an independent prognostic factor for clinical outcomes after radical cystectomy is controversial. The objective of the current study was to assess the associations between age and clinical outcomes in a large, multi-institutional series of patients treated with radical cystectomy for bladder cancer., Materials and Methods: Institutional radical cystectomy databases containing detailed information on bladder cancer patients treated between 1993 and 2008 were obtained from 8 academic centers in Canada. Data were collected on 2,287 patients and combined into a relational database formatted with patient characteristics, pathologic characteristics, recurrence status, and survival status. Patient age was coded as <60 years, 60-69 years, 70-79 years, or ≥ 80 years. Clinical outcomes were 30-day mortality, 90-day mortality, overall survival (OS), disease-specific survival (DSS), and recurrence-free survival (RFS). Logistic regression and Cox proportional hazards regression analysis were used to analyze survival data., Results: Five hundred fifty-seven (24.6%), 679 (30.0%), 846 (37.4%), and 181 (8.0%) patients were <60 years, 60-69 years, 70-79 years, and ≥ 80 years, respectively. Increased age was associated with decreased utilization rates of neoadjuvant chemotherapy (P = 0.0143), adjuvant chemotherapy (P < 0.0001), and continent urinary diversion (P < 0.0001) as well as advanced pathologic tumor stage (P = 0.0003), increased positive surgical margins (P < 0.0001), and lymphovascular invasion (P = 0.0335). Compared with patients < 60 years, multivariate regression analysis showed that age ≥ 80 years was independently associated with 90-day mortality (OR 2.98, 95% CI 1.22-7.30), OS (HR 2.03, 95% CI 1.51-2.75), DSS (HR 1.56, 95% CI 1.09-2.24), and RFS (HR 2.06, 95% CI 1.57-2.70)., Conclusions: Age ≥ 80 years at the time of radical cystectomy was independently associated with adverse survival outcomes. These data suggest that increased chronologic age should be considered in clinical trial design and in nomograms predicting survival., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Unlabelled: Study Type - Prognosis (cohort) Level of Evidence 2a. What's known on the subject? and What does the study add? Radical cystectomy with pelvic lymph node dissection is recognized as the standard of care for carcinoma invading bladder muscle and for refractory non-muscle-invasive bladder cancer. Owing to high recurrence and progression rates, a two-pronged strict surveillance regimen, consisting of both functional and oncological follow-up, has been advocated. It is also well recognized that more aggressive tumours with extravesical disease and node-positive disease recur more frequently and have worse outcomes. This study adds to the scant body of literature available regarding surveillance strategies after radical cystectomy for bladder cancer. In the absence of any solid evidence supporting the role of strict surveillance regimens, this extensive examination of recurrence patterns in a large multi-institutional project lends further support to the continued use of risk-stratified follow-up and emphasizes the need for earlier strict surveillance in patients with extravesical and node-positive disease., Objectives: • To review our data on recurrence patterns after radical cystectomy (RC) for bladder cancer (BC). • To establish appropriate surveillance protocols., Patients and Methods: • We collected and pooled data from a database of 2287 patients who had undergone RC for BC between 1998 and 2008 in eight different Canadian academic centres. • Of the 2287 patients, 1890 had complete recurrence information and form the basis of the present study., Results: • A total of 825 patients (43.6%) developed recurrence. • According to location, 48.6% of recurrent tumours were distant, 25.2% pelvic, 14.5% retroperitoneal and 11.8% to multiple regions such as pelvic and retroperitoneal or pelvic and distant. • The median (range) time to recurrence for the entire population was 10.1 (1-192) months with 90 and 97% of all recurrences within 2 and 5 years of RC, respectively. • According to stage, pTxN+ tumours were more likely to recur than ≥pT3N0 tumours and ≤pT2N0 tumours (5-yr RFS 25% vs. 44% vs. 66% respectively, P < 0.001). Similarly, pTxN+ tumours had a shorter median time to recurrence (9 months, range 1-72 months) than ≥pT3N0 tumours (10 months, range 1-70 months) or ≤pT2N0 tumours (14 months, range 1-192 months, P < 0.001)., Conclusions: • Differences in recurrence patterns after RC suggest the need for varied follow-up protocols for each group. • We propose a stage-based protocol for surveillance of patients with BC treated with RC that captures most recurrences while limiting over-investigation., (© 2012 BJU INTERNATIONAL.)

Unlabelled: Study Type -Therapy (case series)., Level of Evidence: 4., Objective: To evaluate data obtained from a large, multi-institutional, contemporary series of patients who underwent radical cystectomy (RC) in a universal healthcare system aiming to assess outcome and identify novel prognostic variables., Materials and Methods: Data were collected and pooled from 2287 patients treated with RC between 1998 and 2008 by urological oncologists from eight Canadian academic centres. Collected variables included various clinicopathological parameters, recurrence and death. Survival and prognostic variables were analyzed using the Kaplan-Meier method and Cox regression analysis., Results: The median age of patients was 68 years with a mean (median) follow-up time of 35 (29) months. The 30, 60 and 90-day postoperative mortality rates were 1.3%, 2.6% and 3.2%, respectively. The 5-year overall, recurrence-free and cancer-specific survival was 57%, 48% and 67%, respectively, with a local recurrence rate of 6%. Pathological stage distribution was

Unlabelled: Study Type - Prognosis (case series)., Level of Evidence: 4 OBJECTIVE: To determine whether the expression of cyclo-oxygenase (COX)-2 has an influence on survival and on the response to chemotherapy in invasive bladder cancer., Patients and Methods: A population of 266 patients from a tertiary university centre with carcinoma invading bladder muscle without evidence of metastasis at time of cystectomy was analyzed retrospectively. COX-2 expression was evaluated immunohistochemically with a monoclonal anti-COX-2 antibody. All pertinent clinical and pathological parameters were reviewed and correlated with risk factors influencing outcome, including disease-specific and overall survival, as well as COX-2 expression. Immunoreactivity was categorized as positive if COX-2 staining was present in >5% tumour cells., Results: The expression of COX-2 was not influenced by tumour stage, grade or nodal status, nor any other parameters. The risk factors that influenced disease-specific survival in carcinoma invading bladder muscle on multivariate analysis were lymph node status (hazards ratio, HR = 2.46 for N1, P = 0.001, HR = 2.90 for N2, P < 0.001, HR = 5.19 for N3, P = 0.012), use of neoadjuvant chemotherapy (HR = 3.54; P= 0.004) or adjuvant chemotherapy (HR = 0.57, P = 0.014) and COX-2 expression (HR = 0.64 if >5% cells had positive expression; P = 0.025). Kaplan-Meier analysis showed an increased disease-specific survival (P = 0.0063), as well as longer recurrence-free survival (P = 0.003), in patients with muscle-invasive bladder tumours expressing COX-2 in >5% of the cells. A tendency was also observed in a subgroup with positive nodes treated with adjuvant chemotherapy (P = 0.093)., Conclusions: The overexpression of COX-2 is associated with a better recurrence-free and disease-specific survival in a large cohort of 266 patients with carcinoma invading bladder muscle treated by cystectomy. A trend for increased disease-specific survival was also observed for patients with COX-2 overexpression and positive nodes who received adjuvant chemotherapy. Potential of COX-2 as a prognostic marker in bladder cancer should be considered., (© 2010 THE AUTHORS; BJU INTERNATIONAL © 2010 BJU INTERNATIONAL.)

Objective: To investigate the role of (18) F-fluorodeoxyglusose positron-emission tomography (FDG-PET), combined with computed tomography (CT) and forced diuresis, in the staging and follow-up of urothelial carcinoma (UC)., Patients and Methods: We recruited 44 patients with muscle-invasive urothelial bladder cancer (UBC) before radical cystectomy (RC), 19 under follow-up after RC and seven after systemic chemotherapy. For those who had RC, histopathology was used as the reference standard to compare the sensitivity and specificity of FDG-PET/CT and standard CT in detecting UBC and pelvic lymph node metastasis. Furthermore, 36 patients with ≥ 6 months of follow-up imaging were considered to describe the progression of UC and extrapelvic positive FDG-PET/CT images., Results: For the detection of primary UBC, FDG-PET/CT was slightly more sensitive than CT (85% vs 77%) but less specific (25% vs 50%). For the detection of pelvic node metastasis FDG-PET/CT was more sensitive than CT (57% vs 33%) with a specificity of 100% for both imaging techniques. In 20 patients, extrapelvic FDG-PET/CT images showed suspected disease at the first evaluation. UC progressed in nine of the 10 patients who had synchronous multiple PET-positive retroperitoneal or mediastinal lymph nodes, and in only two of the nine with unique hyperactive lesions in the lung. FDG-PET/CT also detected a pT1G3 UC of the renal pelvis and all bone metastases detected by bone scintigraphy., Conclusions: FDG-PET/CT could replace standard CT and bone scintigraphy in the presurgical staging and monitoring of patients with UC after surgery or chemotherapy., (© 2010 THE AUTHORS. JOURNAL COMPILATION © 2010 BJU INTERNATIONAL.)

Cancer-testis (CT) genes encode proteins that are ideal targets for cancer immunotherapy because of their restricted expression in normal tissues and frequent expression in cancers. We previously observed that MAGE-A9 was one of the CT genes most frequently expressed in bladder tumors. To confirm that observation and evaluate the potential prognostic value of MAGE-A9 protein, we analyzed its expression by immunohistochemistry in 493 primary bladder tumors and 33 lymph node metastases, in comparison with MAGE-A4 protein, also frequently expressed in bladder tumors. Overall, MAGE-A4 and MAGE-A9 were observed, respectively, in 38% and 63% of nonmuscle-invasive tumors, 48% and 57% of muscle-invasive tumors, 65% and 84% of carcinomas in situ and in 73% and 85% of lymph node metastases. Expression was associated with higher grade (MAGE-A4, p = 0.007; MAGE-A9, p = 0.012). In multivariate Cox regression analyses, expression of MAGE-A9 in pTa tumors was associated with recurrence (HR = 1.829; p = 0.010). In univariate analyses, MAGE-A4 expression in these same tumors was associated with progression to muscle-invasive cancer (HR = 7.417, p = 0.013). MAGE-A9 expression was even more predictive of progression as all tumors that progressed expressed this antigen. In muscle-invasive bladder tumors, no association was found between expression of either MAGE and bladder cancer-specific death. In conclusion, MAGE-A9 is a target of choice for bladder cancer immunotherapy as it is expressed in 60% of bladder tumors, predominantly high-grade tumors, and at higher frequency in pTis and metastatic tumors. Moreover, in pTa tumors, an immunotherapy targeting MAGE-A9 could be protective against recurrence and progression to more advanced cancer., (2009 UICC)

Background: Carcinomas of the upper urinary tract are uncommon tumors that usually occurred in elderly patients. Competing causes of mortality should be considered when treating these patients., Methods: All patients with upper urinary tract tumors who were treated surgically at Centre Hospitalier Universitaire de Québec and affiliated hospitals from 1978 to 2001 were retrospectively reviewed. Clinical and pathologic variables were assessed from both the preoperative and postoperative periods of management, and clinical outcomes were tracked. Competing risks regression, Cox proportional hazards modeling, and multiple imputation were used to assess predictors of cancer-related and competing risks-related mortality in both preoperative and postoperative settings., Results: Competing risks were responsible for 46% of deaths in this cohort of 168 patients. Preoperatively, the most important predictor of cancer-related mortality was a clinically invasive tumor (hazards ratio [HR], 3.97; P < .001), whereas increasing age (HR, 1.07; P < .001) was found to be the most important predictor of competing mortality. Postoperatively, tumor grade was the most important predictor of cancer-related mortality (HR, 3.92; P < .001) whereas constitutional symptoms (HR, 1.91; P = .015) and increasing age (HR, 1.06; P < .001) were found to be predictive of competing mortality., Conclusions: In the current study, stage and grade were found to be the 2 most important independent predictors of survival in patients with tumors of the upper urinary tract and were highly correlated. Pain or weight loss was found to be a novel predictor of survival in this cancer. Although a survival disadvantage was not noted for women, nephron-sparing surgery, ureteral tumors, or older patients with respect to cancer, competing causes of mortality were found to be responsible for greater than one-third of observed deaths and age was the best predictor of this occurrence.

Background: The clinical significance of tumor-infiltrating dendritic cells (TIDCs) and tumor-associated macrophages (TAMs) as markers of immune response has been reported for many cancers., Objective: To measure tumor infiltration by CD83(+) dendritic cells (DCs) and CD68(+) macrophages in non-muscle-invasive urothelial cancer (NMIUC) prior to bacillus Calmette-Guérin (BCG) immunotherapy and to evaluate their significance in the response to immunotherapy., Design, Setting, and Participants: Patients with NMIUC at high risk of recurrence and progression were recruited for a study on markers of the response to BCG., Intervention: Patients were treated by transurethral resection followed by maintenance BCG., Measurements: Immunohistochemical staining with anti-CD83 and anti-CD68 monoclonal antibodies on 53 and 46 NMIUC tumors, respectively, prior to BCG treatment. A scoring index was calculated based on the average density of positive cells within the papillary axis, the stroma, lymphoid aggregates, and infiltration into tumors., Results and Limitations: CD83(+) TIDCs were observed mostly within lymphoid aggregates. Multivariate Cox regression analysis showed that maintenance BCG (more than one maintenance cycle) was highly effective in patients with a low level of CD83(+) TIDCs at time of resection (hazard ratio [HR]: 0.035; p=0.002) but showed reduced efficacy in patients with a high level of CD83(+) TIDCs (HR: 0.87; p=0.810). A high level of infiltration by CD83(+) TIDCs slightly decreased the risk of recurrence in patients treated with one or no maintenance BCG cycle (HR: 0.4; p=0.117). In the same population, a strong infiltration of CD68(+) TAMs was associated with an increased risk of recurrence (HR: 3.8; p=0.013)., Conclusions: These results suggest that patients with a high level of infiltration by CD83(+) TIDCs or CD68(+) TAMs do not respond as well to BCG immunotherapy. If confirmed in larger cohorts, the pretreatment level of infiltration by these cells may be useful to influence the choice of treatment strategy.

Background: Intravesical bacillus Calmette-Guérin (BCG) immunotherapy is effective in preventing recurrence and progression in nonmuscle-invasive bladder cancer but the dosing schedule and duration of treatment remain empirical. The outcome of BCG therapy was prospectively evaluated on patients according to the number of maintenance cycles received., Methods: Between 1997 and 2002, 111 patients with nonmuscle-invasive bladder cancer at high risk of recurrence and progression underwent transurethral resection followed by intravesical instillations of BCG. After an induction treatment of 6 weekly instillations, patients were scheduled to receive a 3-weekly maintenance treatment at the 3rd, 6th, 12th, 18th, 24th, 30th, and 36th month. At each visit a clinical assessment was obtained., Results: Over a median clinical follow-up of 31 months, 43% of patients experienced recurrence and 8.1% progressed to muscle-invasive disease or metastasis. Only 1 patient received all scheduled instillations. Patients who received at least 3 maintenance BCG cycles had a significantly reduced risk of recurrence (hazard ratio [HR] = 0.23, P = .0064, adjusted for gender, age, and stage) compared with patients receiving only induction BCG. Twelve months after the end of maintenance, the estimated Kaplan-Meier recurrence-free survival was 89% for patients who received at least 3 maintenance BCG cycles, 67% for those who received 2 maintenance BCG cycles, and 41% for those who received only the induction BCG or 1 maintenance cycle (P = .0003)., Conclusions: The results of this study suggest that a minimum of 3 cycles of maintenance BCG is required to significantly reduce the recurrence rate. It also suggests that more cycles may result in further improvements but the benefits may be outweighed by increasing side effects in some patients., (2008 American Cancer Society)

Tissue factor (TF), a transmembrane glycoprotein responsible for initiating the extrinsic pathway of blood coagulation plays a key role in cancer growth, metastasis and angiogenesis. Various studies have demonstrated the prognostic potential of TF expression in several cancers. However, its role in bladder cancer is unclear. This study evaluated the prognostic potential of TF expression in muscle-invasive bladder tumors from patients treated with radical cystectomies. Immunohistochemical staining using a monoclonal antibody (mAb) anti-TF was carried out on sections of tissue microarray blocks containing cores of muscle-invasive bladder tumors (4 cores/tumor) from 218 patients. The intensity of the staining was evaluated on a scale from 0 to 3 by two independent observers who were both unaware of the clinicopathological characteristics of the samples. TF was expressed in 77.6% of tumors, independently from baseline characteristics (age, gender, stage and grade) as assessed using the chi(2) and Student t tests. During follow-up (median: 2.6 years), 45.4% of the patients died from the progression of their cancer. Kaplan-Meier survival showed that among the 103 patients with node-negative (N0) transitional cell carcinoma (TCC), those with TF-positive tumors had shorter bladder cancer-specific survival (p = 0.0276). Moreover, multivariate Cox regression analysis showed they had a 3.15-fold greater risk of dying from bladder cancer (95% CI: 1.1-9.0; p = 0.032). In conclusion, TF expression was an independent predictor of disease-specific survival in N0 muscle-invasive TCCs treated by radical cystectomy and therefore, might help identify patients at higher risk of disease progression. These patients could potentially benefit from adjuvant chemotherapy., ((c) 2007 Wiley-Liss, Inc.)

Background: PD-L1 (programmed death ligand 1, B7-H1) is a cell surface glycoprotein that can impair T-cell function. PD-L1 is aberrantly expressed by multiple human malignancies and has been shown to carry a highly unfavorable prognosis in patients with kidney cancer. The role of PD-L1 was evaluated as a mechanism for local stage progression in urothelial carcinoma (UC) of the bladder., Methods: Using immunohistochemistry, PD-L1 expression was evaluated in a cohort of 280 high-risk UCs of the bladder. PD-L1 was modeled as a predictor of bladder cancer stage using ordinal logistic regression. Other covariates evaluated as potential confounders included age, gender, tumor grade, and lymphocytic infiltration. Further, PD-L1 was evaluated as a potential mechanism of bacillus Calmette-Guerin (BCG) failure in the subset of high-risk nonmuscle-invasive tumors that received this treatment., Results: PD-L1 expression was observed in 7% of pTa, 16% of pT1, 23% of pT2, 30% of pT3/4, and 45% of carcinoma in situ (CIS) tumors. PD-L1 expression was associated with high-grade tumors (odds ratio [OR] = 2.4, P = .009) and tumor infiltration by mononuclear cells (OR = 5.5, P = .004). We observed that the key determinants of stage progression in this cohort were World Health Organization/International Society of Urologic Pathology (WHO/ISUP) high-grade tumor pathology (OR = 4.77, 95% confidence interval [CI]: 2.73-8.34; P < .001) and PD-L1 expression (OR = 2.20, P = .012). PD-L1 expression was found to be extremely abundant in the BCG-induced bladder granulomata in 11 of 12 patients failing BCG treatment., Conclusions: Collectively, these data indicate that tumor PD-L1 may facilitate localized stage-advancement of UC and attenuate responses to BCG immunotherapy by neutralizing T cells that normally guard against cancer invasion from the epithelium into the bladder musculature.

Purpose: Superficial bladder tumors have a high recurrence rate and 10% to 20% of recurrences progress to invasive cancer. Recurrence and progression can best be prevented by nonspecific immunotherapy using intravesical BCG instillations. The NRAMP1 gene has been implicated in susceptibility to infectious and autoimmune diseases, and in response to BCG in murine models. We evaluated the association of 5 NRAMP1 gene polymorphisms with the risk of superficial bladder cancer recurrence and response to immunotherapy., Materials and Methods: The (GT)n, 274 C/T, 469 + 14 G/C, 1465-85 G/A and D534N polymorphisms were tested on peripheral blood DNA of individuals from three cohorts: 37 bladder tumor patients treated by transurethral resection and without recurrence after up to eight years, 67 patients at high risk of recurrence of their bladder tumors and treated with BCG and 109 controls, using restriction fragment length polymorphisms or microsatellite analysis following PCR amplification., Results: The D543N G:A genotype was found more frequently in patients at high risk of recurrence (8 of 67 or 12%) than in controls (2 of 109 or 2%) (p = 0.007). Patients with nonrecurrent tumors showed no difference with controls (1 of 37 or 3%) (p = 1.0). Moreover, in multivariate and survival analyses, both D543N and (GT)n polymorphisms showed association with recurrence-free survival in the cohort of patients at high risk of recurrence, following BCG treatment., Conclusions: These data suggest the implication of the NRAMP1 gene in bladder cancer recurrence and response to BCG immunotherapy.

Biological markers that are predictive of recurrence and progression of superficial bladder tumors must provide additional information to that provided by multiplicity, size and grade. Field anomalies in normal appearing urothelium of patients with papillary superficial transitional cell carcinoma have been associated with tumor antigens and chromosome 9 deletions. Also, primary tumors with chromosome 9 deletions are associated with a higher risk of recurrence. Abnormal expression of p53, p21 and Ki-67 cell cycle markers have little predictive value for recurrence. However, p53 overexpression or mutation and decreased expression of p27 are associated with cancer progression and survival. New markers, such as mutations in the fibroblast growth factor receptor 3 gene (found in 30% of tumors), anomalies of the PTEN gene and vascular endothelial growth factor expression, may have potential and require further evaluation. Molecular fingerprints of superficial tumors with distinct clinical behavior are being rapidly unravelled. Large-scale clinical studies are urgently needed to provide supportive evidence for their incorporation in clinical management.

Our aim was to determine whether the pattern of expression of the interrelated proteins p53, MDM2 and p21 could shed light on the etiopathogenic mechanisms of superficial bladder tumors. Protein expression was detected by immunohistochemistry (IHC) using monoclonal antibodies (MAbs) Pab 1801 for p53, IF2 for MDM2 and EA10 for p21 on 269 newly diagnosed bladder tumors (214 pTa and 55 pT1; 93 grade I, 145 grade 2 and 31 grade 3). While no p21 immunoreactivity was found in normal urothelium, 85% of tumors were strongly p21-positive. MDM2 was overexpressed in 44% of tumors, almost all being also positive for p21. p53 was overexpressed in 20% of cases: 66% of p53-positive tumors were also MDM2 positive, compared with only 38% of p53-negative tumors. p53 mutations were studied by direct DNA sequencing in a subset of 24 high-grade tumors. Both MDM2 and p21 were less frequently expressed in p53 mutated tumors compared with tumors with a wild-type gene. Distinct phenotypes were correlated with the frequency of poorly differentiated (grade 3) tumors. The most common phenotypes were p21+/MDM2-/p53- and p21+/MDM2+/p53- observed in 38% and 29% of tumors, respectively. Grade 3 tumors were found in 4% and 8% of these 2 groups, in contrast with 30% frequency in p21+/p53+ tumors (p = 0.002) regardless of their MDM2 phenotype. Four of the 5 (80%) tumors that were p53-positive but negative for p21 were grade 3. Our data suggest that several tumorigenic pathways for superficial bladder tumors can be reflected by the expression pattern of these 3 proteins.

The aim of the study was to determine whether the expression of the cell cycle markers p53, MDM2, p21, and Ki-67 was predictive of superficial bladder cancer recurrence and to compare the relative predictive power for tumor recurrence of a cell cycle index based on the number of abnormally expressed cell cycle markers with a clinicopathological index based on primary clinical tumor characteristics. The expression of p53, MDM2, and p21 proteins and the value of the Ki-67 index were analyzed for 244 patients. One hundred ninety-four lesions were determined to be superficial papillary tumors (pTa), whereas 50 tumors invaded the lamina propria (pT1). Tumor grade was noted low (grade 1) in 83 cases and high (grades 2-3) in 161 cases. An avidin-biotin peroxidase method was performed using monoclonal antibodies against p53, MDM2, p21, and Ki-67 antigens after antigen retrieval treatment of formalin-fixed specimens. The cell cycle marker index was created using the number of abnormally expressed cell cycle markers according to the following cutoff points: p53 (>5%), MDM2 (>20%), p21 (<5%), and Ki-67 (>10%). The clinicopathological index was created using the following adverse tumor characteristics: grades G2-G3, stage pT1, multifocality, and diameter of tumors > 3 cm. Cox regression models were used to calculate the relative risks and their 95% confidence intervals associated with disease recurrence for the clinicopathological index and the cell cycle marker index. The chi2 test was performed to describe the correlation between the Ki-67 index and p53, MDM2, and p21 protein expression. Kaplan-Meier survival curves were generated to demonstrate the disease-free survival according to these two prognostic indexes. The clinicopathological index was a strong, independent predictor of disease recurrence where tumors with three or four adverse tumor characteristics at initial resection had over four times the risk of recurrence than tumors with no risk factors (P for trend = 0.0001). A strong correlation was observed between the Ki-67 index >10% and both MDM2 and p21 proteins. MDM2 was overexpressed in 106 tumors (43%), and p53 was overexpressed in 47 (19%); Ki-67 was >10% in 171 cases (70%). Thirty-nine tumors (16%) were p21 negative. The risk of recurrence increased slightly with the number of abnormally expressed cell cycle markers, but when the clinicopathological index was taken into account in multivariate analysis, the cell cycle marker index was not predictive of disease recurrence (P for trend = 0.72). The cell cycle markers studied provided no added prognostic information on disease recurrence after initial resection of papillary superficial tumors when the clinicopathological parameters were taken into account.

The biological behavior of urothelial carcinomas remains unpredictable. The objective of this study was to determine the prognostic value of Ki-67 index in superficial papillary bladder tumors and to correlate it with the S-phase fraction (SPF) measured by flow cytometry. Three hundred nineteen patients with newly diagnosed superficial (pTa, pT1) bladder tumors were included between September 1990 and April 1992. Patients with bladder carcinoma in situ alone were excluded. We observed 255 pTa tumors and 64 pT1 tumors, whereas 111 lesions were classified as grade G1 and 208 as grade G2-G3. Ki-67 immunostaining was performed on paraffin-embedded material using a 3-step immunoperoxidase procedure with the murine monoclonal antibody MiB1. The relation between Ki-67 expression and prognostic variables (stage, grade, tumor size, multifocality, age, and sex) was investigated by the chi-square test. Cox regression was used to describe the association between Ki-67 and tumor recurrence in 308 patients with follow-up while adjusting for potentially confounding prognostic variables. The frequency of high Ki-67 expression (> or =10%) increased with stage (P = .005) and grade (P = .001), but not with tumor size or multifocality. Two hundred one patients experienced tumor recurrence in a median follow-up of 68 months. Stage, grade, tumor size, and multifocality were all independent predictors of recurrence. Ki-67 index greater than 10% was found to be an independent predictor of tumor recurrence among patients with tumors larger than 3 cm in diameter (HR = 2.05, CI = 1.18-3.55), but not those with smaller size tumors. With regards to the DNA index, a significant but weak correlation was observed between Ki-67 expression and the SPF (Spearman's correlation coefficient = 0.23, P = .004). In addition, aneuploid tumors had significantly higher expression of Ki-67 (22.5%) than diploid tumors (10.1%) (P = .0006). Moreover, patients with DNA aneuploid bladder tumors were more likely to have more than 10% Ki-67-positive cells than those with diploid tumors. In patients with newly diagnosed pTa or pT1 bladder tumors, a Ki-67 index above 10% is an independent predictor of shorter time to recurrence only in those with tumors larger than 3 cm.

The INK4A and the INK4B genes map to chromosome 9p21, an area frequently deleted in bladder neoplasms. In addition to the p16 protein, the INK4A encodes for a second product, termed p19(ARF). We analyzed tissues from 121 patients with initial Ta and T1 tumors. Deletions of the INK4A gene were observed in 17 of 121 (14.1%) cases. Point mutations were identified in 2 of 64 (3.1%) tumors. The INK4A-exon 1beta and the INK4B gene were codeleted with INK4A in all of the homozygously deleted cases analyzed. The p16 promoter underwent de novo methylation in 7 of 47 (14.9%) evaluable cases. The p16-positive phenotype was observed in 18 of 56 (32%) evaluable cases. p16 negative phenotype correlated with deletion and methylation status. A statistically significant association between INK4A homozygous deletions and tumor size was observed (P = 0.003). Patients bearing tumors with INK4A homozygous deletions had a lower recurrence-free survival (P = 0.040) than those with wild type INK4A. In conclusion, deletions and methylation of the INK4A gene occur frequently in superficial bladder tumors. However, only those deletions that affect both the p16 and the p19(ARF), deregulating both the pRb and p53 pathways, correlated with clinicopathological parameters of worse prognosis.

The ABO(H) histoblood group genes have been mapped by linkage analysis to 9q34.1-34.2, an area of common deletions in bladder cancer. Lack of ABO(H) antigen expression in bladder tumors is a frequent and well-documented event. In bladder neoplasms the loss of A and B transferase activity is due to down-regulation of the ABO gene transcripts. Our study was undertaken in order to determine the presence of structural alterations of the ABO(H) gene-encoding locus in primary bladder tumors, to estimate the extent of allelic deletions and to characterize further the pattern of histoblood group antigen expression. Fifty-three primary bladder tumors were analyzed by immuno-histochemistry (IHC) using a panel of well-characterized antibodies and fresh frozen tissue sections. Normal and tumor DNA also were analyzed by PCR coupled with restriction enzyme analysis in order to establish the ABO genotype. Results obtained from these analyses were then compared to allelotyping data at the 9q34.1-4 region by Southern blotting. IHC data showed undetectable levels of antigen expression on neoplastic cells in 59% of informative cases. PCR-based genotypic results revealed allelic losses in 27% of heterozygous cases. Four of the 16 pheno- and/or genotypically altered cases (25%) presented loss of heterozygosity at D9S10 or D9S7 loci. Our data indicate that the lack of ABO antigen expression in certain bladder tumors is due to the allelic loss of the ABO glycosyltransferase-encoding genes and that in some of these tumors the loss involves the surrounding chromosomal region at 9q34.1-4.

Epidemiological studies suggest that bladder cancer may be caused by carcinogens in tobacco and certain occupational exposures. Molecular studies have shown that chromosome 9 alterations and TP53 mutations are the most frequent events in bladder cancer. To date, the relationships between epidemiological risk factors and genetic alterations have not been fully explored in bladder cancer. The purpose of this study was to explore the association between smoking and chromosome 9 aberrations in bladder cancer cases. Seventy-three patients with bladder cancer at Memorial Sloan-Kettering Cancer Center were evaluated for smoking history, occupational history, and chromosome 9 alterations. The epidemiological data were abstracted from medical charts. Patients' tumor tissues were analyzed using RFLP and microsatellite polymorphism assays for detection of chromosome 9 alterations. Elevated odds ratios (ORs) were found for chromosome 9 alterations in smokers compared to those in nonsmokers (OR = 4.2; 95% confidence interval, 1.02-17.0) after controlling for age, sex, race, occupational history, and stage of disease. The ORs were 3.6 for those smoking < or = 20 cigarettes per day and 5.8 for those smoking > 20 cigarettes per day. No association was found between occupational history and chromosome 9 alterations. This study supplies evidence suggestive of the link between smoking and chromosome 9 alterations in the etiology of bladder cancer and indicates that potential tumor suppressor genes on chromosome 9 may be involved in smoking-related bladder carcinogenesis.

The proliferative rate of tumor cells is frequently proportional to their degree of aggressiveness. The objective of the present study was to determine the correlation between the expression of three antigens associated with proliferation and the stage and ploidy of bladder cancers. The reactivity of antibodies against the nuclear antigen Ki-67 and membrane antigens T43 and the epidermal growth factor receptor (EGFR) was studied by flow cytometry on a series of 35 clinical samples of superficial and infiltrating bladder cancer and as well as on 5 specimens of normal urothelial cells. A preferential expression of the T43 antigen was observed on invasive cancer. EGFR was less frequently expressed; however, seven of the nine positive samples were from invasive cancers. Ki67 on the other hand did not show any selective expression according to tumor stage. When comparing Ki-67 and T43 expression on individual tumor samples, a negative correlation was found in that no tumor strongly expressed both markers simultaneously. These results suggest that markers of proliferation and activation do not all measure the same tumor parameters. Together, they may provide prognostic information that may be useful in the follow-up of patients with bladder cancer.

Purpose: We have previously demonstrated that p53 overexpression is predictive of disease progression and survival in Ta, Tis, and T1 tumors. Instillation of Bacillus Calmette-Guérin (BCG) is now accepted to be the most efficient adjuvant therapy for superficial bladder carcinoma. The aim of this study was to determine if p53 status, assessed before and after intravesical BCG therapy, can predict clinical outcome in a high-risk population of patients with superficial bladder carcinoma., Materials and Methods: We examined 196 tissue specimens from 98 patients, obtained immediately before and after intravesical BCG therapy. The pretherapy population was composed of 22 Ta, 57 Tis, and 19 T1 tumors. After BCG, 66 specimens were TO and 32 had residual tumors. Nuclear p53 overexpression was analyzed in relation to time to disease progression and disease-specific survival., Results: The median follow-up duration was 44 months. The detection of nuclear p53 overexpression before BCG therapy did not predict response to BCG therapy. Pre-BCG p53 protein overexpression, response to BCG therapy, and pre-BCG stage were all independent markers of disease progression. In patients with residual disease after BCG therapy (nonresponders), multivariate analysis confirmed that posttherapy p53 overexpression was the only independent marker of disease progression., Conclusion: In this high-risk population of patients with superficial bladder tumors, patients who have p53 nuclear overexpression in the tumor and stage T1 disease before BCG therapy are at high risk of disease progression. Furthermore, in the group of patients with residual disease after BCG therapy, p53 status is a better predictor of disease progression than post-BCG stage.

The p21WAF1/CIP1 gene is regulated by p53 and encodes a cyclin-dependent kinase (Cdk)-inhibitor involved in senescence and cell quiescence. The role of p21 as a negative regulator of cell proliferation suggests that it may function as a tumor suppressor gene. However, only a few mutations of the p21WAF1/CIP1 gene have been reported to date. In order to assess potential p21WAF1/CIP1 gene alterations in human bladder cancer, we have examined this gene and its encoded product in a well-characterized cohort of 27 primary bladder tumors. Mobility shifts by single-strand conformation polymorphism in the p21WAF1/CIP1 gene were identified in 2 cases. Sequencing analyses revealed that one of these cases had point mutations in the 3' untranslated region, while the other case had a frame shift mutation at positions 322 (C to A) and a deletion of 8 nucleotides (323-->331; CCG-->ACG, codon 81 Arg-->Thr) that produced a stop signal at codon 83 (Gly--Stop). This tumor had a p21-negative phenotype by immunohistochemistry, but did not lose any allele. We further characterized these cases by the study of TP53 mutations using single-strand conformation polymorphism (PCR-SSCP) and sequencing, as well as immunohistochemical assays. Seven mobility shifts were identified and seven cases showed p53 nuclear accumulation. The two cases displaying mutated p21WAF1/CIP1 had wild-type TP53. It is concluded that p21WAF1/CIP1 gene aberrations are infrequent in bladder carcinoma but may be occasionally identified in primary bladder tumors.

Background: Two genes, p16 (also known as CDKN2, INK4A, or MTS1) and p15 (also described as INK4B or MTS2), are found in tandem at chromosome 9p21. These genes are designated as candidate tumor suppressor genes because they encode proteins that function as negative cell cycle regulators. (The encoded polypeptides inactivate specific cyclin-protein kinase complexes that are required for progression through the cell cycle.) Molecular genetic studies have revealed that deletion of the p16 and p15 genes occurs frequently in cancer cell lines and in certain malignant neoplasms., Purpose: We evaluated the frequency of p16 and p15 gene alterations in a well-characterized cohort of human transitional cell bladder cancers, and we explored potential associations between alterations in these genes and tumor stage and/or grade., Methods: Tumor tissue and normal tissue from 110 patients with transitional cell carcinoma of the urinary bladder were examined. The status of the p16 and p15 genes in these tissues was determined by Southern blotting and hybridization with gene-specific probes, by coupled polymerase chain reaction and single-strand conformation polymorphism analysis (PCR-SSCP), and by sequencing DNA fragments produced during PCR. Associations between alterations in the genes and tumor stage and/or grade were evaluated using the two-tailed Fisher's exact test., Results: Homozygous deletion (both alleles lost) of the p16 and the p15 genes was observed in 11 and nine bladder tumors, respectively. Eight of the 11 tumors exhibiting complete loss of the p16 gene also displayed homozygous deletion of the p15 gene. Exclusive loss of either gene was detected in only three tumors. Hemizygous deletion (one allele lost, also referred to as loss of heterozygosity [LOH] of the p16 and/or p15 genes was observed in eight tumors. Rearrangement of the two genes was indicated in three additional tumors. No point mutations were identified in either gene. The overall frequency of alteration in this cohort of bladder tumors was approximately 18% for each gene (in 20 [18.3%, 95% confidence interval (CI) = 11.1%-25.6%] of 109 informative tumors for the p16 gene and in 18 [18%, 95% CI = 10.5%-25.5%] of 100 informative tumors for the p15 gene). A statistically significant association between p16 gene alteration and bladder tumors of low stage (P < .01) and grade (P < .01) was observed; a significant association between p15 gene alteration and tumors of low stage (P < .01) was also detected., Conclusions: Alteration of the p16 and p15 genes, especially coincident homozygous deletion, appears to be a common event in bladder cancer.

The 9p21 region of human chromosome 9 is a hot spot for chromosomal aberrations in both cultured cell lines and primary tumors. This region contains a gene, P16 (also called MTS1, CDKN2 and p16INK4), that encodes a presumptive negative cell cycle regulator called p16. P16 is deleted or mutated at high frequency in a variety of tumor cell lines including melanoma and bladder carcinoma lines. As such, it is likely to be a tumor suppressor gene. Here we show that P16 is mutated in primary bladder carcinomas (3 of 33) and melanomas (5 of 34). These findings support studies that show P16 mutations are not solely a product of growth in tissue culture but rather are involved in formation of tumors in viva. Some bladder primary tumors and some bladder and melanoma tumor cell lines contain mutations in both P16 and P53 at frequencies that suggest that p53 and p16 function in different pathways, each of which is important in suppressing malignant transformation.

Chromosome 9 allelic losses have been reported as a frequent and early event occurring in bladder cancer. It has been postulated that a candidate tumor suppressor gene may reside on this chromosome, alterations of which may lead to the development of a subset of superficial bladder tumors. More recently, the involvement of two different regions harboring suppressor loci, one on each of both chromosome 9 arms, has been proposed. We undertook the present study with the objectives of better defining the deleted regions of chromosome 9 in bladder tumors, as well as evaluating the frequency of microsatellite alterations affecting certain loci on this chromosome in urothelial neoplasia. Seventy-three primary bladder tumors were analyzed using a set of highly polymorphic markers, and results were correlated with pathological parameters associated with poor clinical outcome. We observed that, overall, 77% of the tumors studied showed either loss of heterozygosity for one or more chromosome 9 markers and/or microsatellite abnormalities at chromosome 9 loci. Detailed analyses showed that two regions, one on 9p at the interferon cluster, and the other on 9q associated with the q34.1-2 bands, had the highest frequencies of allelic losses. Furthermore, Ta lesions appeared to present mainly with 9q abnormalities, while T1 tumors displayed a mixture of aberrant 9p and 9q genotypes. These observations indicate that loss of heterozygosity of 9p may be associated with the development of superficial tumors with a more aggressive biological behavior or, alternatively, they may be related to early disease progression. In addition, microsatellite alterations were documented in over 40% of amplified cases. Taken together, these data suggest that two different tumor suppressor gene loci on chromosome 9 are involved as tumorigenic events in bladder cancer and that chromosome 9 microsatellite alterations are frequent events occurring in urothelial neoplasia.