Narrow-band imaging assisted cystoscopy in the follow-up of patients with transitional cell carcinoma of the bladder: a randomized study in comparison with white light cystoscopy.

PURPOSE: To evaluate the diagnostic accuracy of a second look narrow-band imaging (NBI) cystoscopy in the follow-up of patients with NMIBC as compared to a second white light cystoscopy (WLI). PATIENTS AND METHODS: From August 2013 to October 2014, 600 patients with history of non-muscle invasive bladder cancer (NMIBC), who presented for follow-up cystoscopy at an academic outpatient clinic, were randomized to flexible WLI-cystoscopy plus second look NBI-cystoscopy (n = 300) or flexible WLI-cystoscopy plus second look WLI-cystoscopy (n = 300) in the same session. We analysed the detection rate of bladder tumours in second look cystoscopy as primary endpoint. In addition, we evaluated recurrence rates before study enrolment and after transurethral resection (TUR-BT) in each group. RESULTS: In 600 patients with a history of NMIBC, 78 out of 300 patients (26%) with WLI-NBI-cystoscopy and 70 out of 300 patients (23%) with WLI-WLI-cystoscopy were diagnosed with cancer recurrence (p = 0.507). Overall, WLI-NBI detected 404 and WLI-WLI 234 lesions, respectively. The second look cystoscopy detected 57 additional cancer lesions: 45 tumours in 18 patients with WLI-NBI and 12 tumours in 9 patients with WLI-WLI (p = 0.035). After initial examination without tumour detection an improvement was determined by the second cystoscopy in 3 patients (75 vs. 78 pat.) with WLI-NBI and in only one patient (69 vs. 70 pat.) with WLI-WLI (p = 0.137). Second look cystoscopy did not influence the detection of carcinoma in situ in both groups (p = 0.120). After TUR-BT the median recurrence-free survival was 4 months in 57 recurring patients (73%) in the group with WLI-NBI- and 6 months in 56 patients (80%) with WLI-WLI-cystoscopy (p = 0.373), respectively. CONCLUSION: Our study showed no differences in per-patient tumour detection between WLI and NBI. Although NBI has significant benefits for detecting individual lesions overlooked by WLI-cystoscopy, this did not positively affect recurrence-free survival after transurethral resection.

Diagnosis and Empirical Treatment of Urinary Tract Infections in Urologic Outpatients.

INTRODUCTION: Due to a continuing increase of bacterial resistance in common uropathogens, we wanted to revisit our standards for the diagnosis and treatment of lower urinary tract infections, in the setting of urological outpatient care in a conurbation in Germany. PATIENTS AND METHODS: All subjects presenting with significant bacteriuria at our urology clinics in Mülheim, Germany, in 2011 were included. Comorbidity, bacterial species, urinary tract symptoms, and empirically prescribed antibiotics were taken from the patients' records. RESULTS: In 2011, a total of 1,324 patients were included (793 female, 531 male). Of the 771 patients with symptomatic bacteriuria, 647 received antibiotic treatment, as well as 116 of 409 patients with asymptomatic bacteriuria. Escherichia coli was identified in 60% of the included patients. In 427 E. coli infections, bacterial resistance was found in 14% of 316 cases treated with quinolone, in 21% of 53 cases treated with co-trimoxazole, and in only 3% of 58 cases treated with nitrofurantoin. CONCLUSIONS: We found a high use of fluoroquinolones for empirical first-line antibiotics in the treatment of lower urinary tract infections. In our regional setting, antibiotic stewardship needs to be promoted, along national and international guidelines, to avoid unnecessary prescription of fluoroquinolones for empirical treatment.

Haematuria work-up in general care-A German observational study.

BACKGROUND: Haematuria is a common finding in the population and the diagnostic workflow of this symptom represents a large proportion of "work-load" in the urological outpatient clinic. AIMS: The intention of this study was to verify if the intensive diagnostic procedures of haematuria patients is justified by detection of a significant proportion of genito-urinary tract cancers. MATERIALS AND METHODS: In a retrospective design 1049 consecutive patients, who presented themselves with macro- or microhaematuria in the outpatient clinic PURR in the time from 2011 to 2012, were included in the study and the diagnostic procedures including ultrasound, intravenous urography, computed tomography of the abdomen and urethrocystoscopy as well as therapeutic consequences with its results were analysed. RESULTS: The study group comprised 570 women (54.3%) and 479 men (45.7%) with a median age of 58 years and macrohaematuria occurred in 89 patients. Diagnostics revealed seven patients with renal cell cancer, six patients with urothelial cell cancer of the renal pelvis, four patients with urothelial cell cancer of the ureter, 65 patients with urothelial cell cancer of the lower urinary tract and 17 patients with prostate cancer. Age, male gender and macrohaematuria were associated with a higher risk of cancer. CONCLUSIONS: The high incidence of urinary tract cancer in the data presented here support the rationale for diagnostic work-up of patients with micro- or macrohaematuria. Prospective randomised trials are necessary to identify index patients for second work-up after a primarily negative investigation as well as the role of molecular markers, which possibly enable to omit invasive work-up.

Mistletoe Plant Extract in Patients with Nonmuscle Invasive Bladder Cancer: Results of a Phase Ib/IIa Single Group Dose Escalation Study.

PURPOSE: We determined the maximum tolerated dose, safety and effectiveness of intravesical instillation of mistletoe extract after transurethral resection of nonmuscle invasive bladder cancer. MATERIALS AND METHODS: In this single group dose escalation study patients with nonmuscle invasive bladder cancer were treated with weekly instillations of mistletoe extract for 6 weeks. Four weeks before instillation therapy all patients underwent transurethral resection of bladder tumors. During this procedure a marker tumor was left. At 12 weeks after the start of instillation therapy transurethral resection of the marker tumor or biopsy of the former marker tumor location was done so that patients were tumor free when entering followup until week 48. During the followup clinical assessment laboratory tests for safety and cystoscopy were done every 12 weeks. RESULTS: A total of 36 patients were treated with increasing doses of mistletoe extract. We found no dose limiting toxicity up to a dose of 675 mg of plant extract. Besides local reactions we saw hints that pyrexia may develop. All adverse events were well manageable. At 12 weeks a marker tumor remission rate of 55.6% (95% CI 38.1 to 72.1) was achieved. At 1 year a recurrence rate of 26.3% (95% CI 9.1 to 51.2) was observed. CONCLUSIONS: In this study intravesical instillation of mistletoe extract as treatment in patients with nonmuscle invasive bladder cancer was shown to be safe and well tolerated. Promising data on efficacy were observed and will be further investigated in a phase III study.

Open-label, 9-month extension study investigating the uro-selective alpha-blocker silodosin in men with LUTS associated with BPH.

OBJECTIVES: To evaluate the long-term safety (primary objective) and efficacy/impact on quality of life (QoL, secondary objectives) of silodosin 8 mg once daily in men with LUTS/BPH. PATIENTS AND METHODS: Men who completed the 12-week double-blind study with silodosin 8 mg, tamsulosin 0.4 mg, or placebo were offered to continue with the 9-month open-label study during which all patients received silodosin 8 mg once daily. Safety was assessed by analysing vital signs, electrocardiograms, laboratory tests, and adverse events. Efficacy was evaluated with the International Prostate Symptom Score (IPSS), IPSS voiding and storage sub-scores, IPSS-QoL, and maximum urinary flow rate (Q max). RESULTS: A total of 500 patients (mean age 66 years) entered the 9-month open-label study. Treatment-emergent adverse events (TEAE) were experienced by 33.4% patients. Ejaculation dysfunction was the most common TEAE (9.0%) but led to study discontinuations in only 1.6% of patients. Dizziness without orthostatic hypotension occurred in 0.8%. A marked reduction in total IPSS (-2.7 ± 3.8) was documented at the first visit of this extension phase in patients having de novo silodosin compared with lesser improvement in patients previously treated with silodosin (-0.82 ± 4.2) or tamsulosin (-0.83 ± 3.8). Improvements were maintained throughout the open-label phase. QoL also improved, with the greatest improvement in de novo silodosin patients. No relevant changes in Q max occurred. CONCLUSIONS: Long-term treatment with silodosin was safe and efficacious. Abnormal ejaculation was the most common TEAE, but led to treatment discontinuation in only 1.6% of patients. Orthostatic hypotension was not seen, and only a few patients experienced dizziness.

Consistent and significant improvement of nighttime voiding frequency (nocturia) with silodosin in men with LUTS suggestive of BPH: pooled analysis of three randomized, placebo-controlled, double-blind phase III studies.

OBJECTIVES: Nocturia is prevalent and bothersome in men with lower urinary tract symptoms suggestive of BPH (LUTS/BPH). α-Adrenoceptor antagonists without subtype selectivity have inconsistently shown significant effects on nocturia in these patients. We explored the effects of the α1A-adrenoceptor subtype-selective antagonist silodosin on nocturia by analyzing three placebo-controlled registration studies. METHODS: Responses to question 7 of the IPSS questionnaire were analyzed for the entire study population and patients with ≥ 2 voids/night at baseline. Improvement/worsening rates for nocturia were calculated for once-daily silodosin 8 mg and placebo. Silodosin effects on the mean number of nocturnal voids were compared with placebo, and the number of patients in whom nocturia was reduced to <2 times was calculated. RESULTS: In total, 1,479 men were treated with silodosin or placebo; 1,266 men (85 %) had ≥ 2 voids/night at baseline. Compared to placebo, more men treated with silodosin reported about nocturia improvement (53.4 vs. 42.8 %, p < 0.0001) and fewer patients about worsening (9.0 vs. 14.3 %, p < 0.0001). Silodosin significantly reduced nocturia within each study and pooled cohort compared to placebo (p < 0.001). In men with ≥ 2 nocturnal voids at baseline, 61 and 49 % of patients with silodosin and placebo had reductions of ≥ 1 voids/night, respectively (p = 0.0003), and significantly more patients with silodosin had <2 nocturia episodes at study end compared to placebo (29.3 vs. 19.0 %; p = 0.0002). CONCLUSIONS: Although a weak impact on nocturia is already known from α-adrenoceptor antagonists without subtype selectivity, the individual placebo-controlled studies and the pooled data analysis showed that the α1A-adrenoceptor subtype-selective antagonist silodosin consistently and significantly improves nocturia in men with LUTS/BPH.

Pudendal nerve block in HDR-brachytherapy patients: do we really need general or regional anesthesia?

PURPOSE: In male patients, the pudendal block was applied only in rare cases as a therapy of neuralgia of the pudendal nerve. We compared pudendal nerve block (NPB) and combined spinal-epidural anesthesia (CSE) in order to perform a pain-free high-dose-rate (HDR) brachytherapy in a former pilot study in 2010. Regarding this background, in the present study, we only performed the bilateral perineal infiltration of the pudendal nerve. METHODS: In 25 patients (71.8 ± 4.18 years) suffering from a high-risk prostate carcinoma, we performed the HDR-brachytherapy with the NPB. The perioperative compatibility, the subjective feeling (German school marks principle 1-6), subjective pain (VAS 1-10) and the early postoperative course (mobility, complications) were examined. RESULTS: All patients preferred the NPB. There was no change of anesthesia form necessary. The expense time of NPB was 10.68 ± 2.34 min. The hollow needles (mean 24, range 13-27) for the HDR-brachytherapy remained on average 79.92 ± 12.41 min. During and postoperative, pain feeling was between 1.4 ± 1.08 and 1.08 ± 1.00. A transurethral 22 French Foley catheter was left in place for 6 h. All patients felt the bladder catheter as annoying, but they considered postoperative mobility as more important as complete lack of pain. The subjective feeling was described as 2.28 ± 0.74. Any side effects or complications did not appear. CONCLUSIONS: Bilateral NPB is a safe and effective analgesic option in HDR-brachytherapy and can replace CSE. It offers the advantage of almost no impaired mobility of the patient and can be performed by the urologist himself. Using transrectal ultrasound guidance, the method can be learned quickly.

Central venous port implantations via the cephalic vein applying an intravasal electrographic control of the catheter tip position: a single-center experience of 316 cases.

PURPOSE: The aim of this study was the analysis of the feasibility and complication rate of central venous port implantation with the surgical cut-down technique applying an intravasal electrographic control of the catheter tip position performed by urologists. PATIENTS AND METHODS: In the time from December 1999 to November 2010, implantation of 324 subcutaneously implanted venous port systems (NuPort-System) has been performed in 316 patients, 221 men (mean age 59.5 years, range 11-87 years) and 95 women (mean age 57.6 years, range 7-85 years). Two hundred and fifty-nine (79.9%) port systems were placed under electrographic control of the catheter tip position. Duration of procedure, long-term device function, and complications such as infections, occlusions, dislocations, and thrombosis were all retrospectively measured and recorded until removal of the device, patient's death or the last known recorded documentation. RESULTS: In total, 359 devices have been used in 348 surgical procedures, 324 implantations (90.25%), and 35 explantations (9.75%). Port systems were implanted using the cephalic vein in 275 patients (84.9%), and in 49 (15.1%), the subclavian vein was used for insertion of the catheter. Mean surgical implantation time was 38.8 min (15-85 min). The median follow-up was 490.6 days (range 2-2,568); 159,764 catheter days (mean, 234 days, range 2-2,604) were documented. Of 35 explanted devices, the explantation was necessary due to complications in 28 cases (8.6%) with infection n = 6 (1.9%, 0.037 per 1,000 catheter days), occlusion n = 8 (2.5%, 0.050 per 1,000 catheter days), dislocation n = 7 (2.2%, 0.044 per 1,000 catheter days), deep vein thrombosis of the upper extremity n = 6 (1.9%, 0.037 per 1,000 catheter days), and clotting n = 1 (0.3%, 0.006 per 1,000 catheter days). Premature catheter removal (<30d post-operatively) was required in 6 patients (1.9%, 0.037 per 1,000 catheter days) due to complications: 3 catheter dislocations/malfunctions (0.9%, 0.019 per 1,000 catheter days), one port related infection, one pocket port infection, and one deep vein thrombosis of the upper extremity (0.3%, 0.006 per 1,000 catheter days). CONCLUSIONS: The intra-atrial ECG techniques to judge correct tip positioning for central venous port implantations are simple and economical. The exact position can be determined intraoperatively. It can justify a delayed postoperative chest X-ray to confirm CVC line tip placement. Nevertheless, the procedure and handling of the device later on has to be performed with care in order to avoid infections and technical problems.

Regulatory BCL2 promoter polymorphism (-938C>A) is associated with adverse outcome in patients with prostate carcinoma.

Molecular markers predictive of prostate cancer prognosis are urgently needed. Overexpression of the antiapoptotic protein, Bcl-2, has repeatedly been shown to be associated with adverse outcome in this malignancy. We hypothesized that a regulatory BCL2 -938C>A promoter polymorphism, which significantly affects promoter activity and Bcl-2 expression in different malignancies, may influence survival. Reporter assays and electrophoretic mobility shift assays reveled that the -938C>A BCL2 promoter polymorphism significantly affects promoter activity and transcription factor binding in prostate cancer cells. Significantly higher BCL2 mRNA expression was observed in primary prostate carcinomas derived from patients with the AA, compared to CC, genotype. Survival analysis showed that the -938AA genotype was an independent, unfavorable prognostic factor for relapse-free survival in a primary cohort of 142 patients and in an independent replication cohort of 148 patients, with hazard ratios (HR) of 4.4 (95% CI, 1.3-15.1; p = 0.018) and 4.6 (95% CI, 1.5-14.2; p = 0.009). Furthermore, the -938AA genotype was independently associated with worse overall survival in the replication series, with a HR of 10.9 (95% CI, 1.2-99.3; p = 0.034). We conclude that the BCL2 -938C>A polymorphism is an independent predictor of relapse-free and overall survival in patients with prostate cancer. The BCL2 -938C>A polymorphism should be evaluated prospectively and may also have promise in assisting optimal patient choice for treatment with BCL2-targeted drugs already in evaluation for prostate cancer treatment.

Serum and tissue syndecan-1 levels in renal cell carcinoma.

BACKGROUND: The proteoglycan syndecan-1 is involved in cell proliferation, adhesion and angiogenesis. It was shown to be involved in cancer progression in different tumor entities. So far, the role of syndecan-1 in renal cell carcinoma (RCC), one of the most common diseases in urologic oncology, was little described. Purpose of the present study was to obtain serum concentrations and tissue expression levels of syndecan-1 in a cohort of patients diagnosed with RCC. METHODS: Clinical and follow-up data were obtained from 413 RCC patients. SDC1 levels were determined in serum samples of 100 patients by enzyme-linked immunosorbent assay and tissue SDC1 expression was measured by immunohistochemistry (IHC) in 343 cases. Results were correlated with clinicopathological and follow-up data. RESULTS: Five and ten years overall and cancer specific survival were 67% and 56% [overall survival (OS)] and 79% and 76% [cancer-specific survival (CSS)]. In female patients and locally advanced disease (≥T3), tissue SDC1 expression was decreased (female 85.6% vs. male 71.1% low tissue SDC1 expression, P=0.0153 and ≤T2 70.0% vs. ≥T3 87.2% low tissue SDC1 expression, P=0.0055) compared to male patients and organ confined disease. Locally advanced tumor stage, presence of lymph node or distant metastases, high Fuhrman grading and clear cell carcinoma as histopathological subtype were independent prognostic factors for reduced CSS and OS. There was no impact of serum SDC1 (sSDC1) serum concentration or SDC1 tissue protein expression on OS, CSS or recurrence free survival (RFS) in uni- or multivariable analysis. CONCLUSIONS: sSDC1 concentration or SDC1 tissue protein expression levels had no influence on patients' prognosis in the present cohort of patients diagnosed with RCC.

The Investigation of Hematuria.

BACKGROUND: Hematuria can be either grossly visible (macrohematuria) or only detectable under a microscope (microhematuria). Microhematuria is often asymptomatic and has a prevalence of 4-5% in routine clinical practice. It may be due to an underlying disease of the kidneys or the urogenital tract. In this article, we provide an overview of the causes of hematuria and of the recommendations of current guidelines for its diagnostic evaluation. A risk-adapted diagnostic strategy for the evaluation of asymptomatic microhematuria (aMH) is presented. METHODS: This review is based on pertinent publications retrieved by a selective search in PubMed, as well as on guidelines from Germany and abroad. RESULTS: Hematuria has many causes, and a broad urological and nephrological differential diagnosis must be considered. In the absence of high-quality scientific evidence, the recommendations of current guidelines for the diagnostic evaluation of hematuria are not uniform; this is particularly so for aMH. Microhematuria is said to be present when urine microscopy reveals three or more erythrocytes per highpower field. The basic diagnostic evaluation consists of a thorough history and physical examination, measurement of inflammatory parameters and renal function tests, and ultrasonography of the kidneys and bladder. Patients with non-glomerular aMH who have risk factors such as smoking, advanced age, and male sex are more likely to have relevant underlying conditions and should therefore undergo augmented, risk-adapted diagnostic evaluation with urethrocystoscopy, urine cytology, and, when indicated, CT urography. Patients with isolated glomerular hematuria are at elevated risk for renal disease and should undergo follow-up checks at six-month intervals. CONCLUSION: Although hematuria is common, there is no uniform, internationally accepted, evidence-based algorithm for its diagnostic evaluation. All potential causes of hematuria must be considered, and all individual risk factors taken into account, so that an underlying disease requiring treatment can be identified or ruled out.

A double-blind, randomized, placebo-controlled clinical trial evaluating the safety and efficacy of autologous muscle derived cells in female subjects with stress urinary incontinence.

PURPOSE: The purpose of the study was to assess safety and efficacy of autologous muscle derived cells for urinary sphincter repair (AMDC-USR) in female subjects with predominant stress urinary incontinence. METHODS: A randomized, double-blind, multicenter trial examined intra-sphincteric injection of 150 × 106 AMDC-USR versus placebo in female subjects with stress or stress predominant, mixed urinary incontinence. AMDC-USR products were generated from vastus lateralis needle biopsies. Subjects were randomized 2:1 to receive AMDC-USR or placebo and 1:1 to receive 1 or 2 treatments (6 months after the first). Primary outcome was composite of ≥ 50% reduction in stress incontinence episode frequency (IEF), 24-h or in-office pad weight tests at 12 months. Other outcome data included validated subject-recorded questionnaires. Subjects randomized to placebo could elect to receive open-label AMDC-USR treatment after 12 months. Subject follow-up was up to 2 years. RESULTS: AMDC-USR was safe and well-tolerated with no product-related serious adverse events or discontinuations due to adverse events. Interim analysis revealed an unexpectedly high placebo response rate (90%) using the composite primary outcome which prevented assessment of treatment effect as designed and thus enrollment was halted at 61% of planned subjects. Post hoc analyses suggested that more stringent endpoints lowered placebo response rates and revealed a possible treatment effect. CONCLUSIONS: Although the primary efficacy finding was inconclusive, these results inform future trial design of AMDC-USR to identify clinically meaningful efficacy endpoints based on IEF reduction, understanding of placebo response rate, and refinement of subject selection criteria to more appropriately align with AMDC-USR's proposed mechanism of action.

The GNAS1 T393C polymorphism predicts survival in patients with clear cell renal cell carcinoma.

PURPOSE: G proteins mediate signaling from cell surface receptors to specific intracellular proteins. In vitro cancer cell line studies revealed a link between the Galphas protein and proapoptotic processes. We have recently shown that TT genotypes of the GNAS1 T393C polymorphism display increased transcription of Galphas and a more favorable clinical course in bladder and colorectal cancer patients compared both with TC or CC genotypes. EXPERIMENTAL DESIGN: In the present study, 150 patients with clear cell renal cell carcinoma surgically treated by nephrectomy with curative intent were retrospectively genotyped to elucidate a potential association between T393C genotypes and clinical outcome. RESULTS: The C-allele frequency in the renal cell carcinoma patient group was 0.51, which is not significantly different from that of a healthy blood donor group. Kaplan-Meier curves for tumor progression, development of metastasis, and tumor-related death showed a significant association of the T393C polymorphism with outcome (5-year cancer-specific survival rates: TT, 91%; TC, 81%; CC, 69%; P = 0.015). Multivariate Cox proportional analysis of a 10-year follow-up confirmed the T393C polymorphism as an independent prognostic factor in clear cell renal cell carcinoma. Homozygous CC patients were at highest risk for progression (hazard ratio, 2.48; P = 0.009) or tumor-related death (hazard ratio, 3.15; P = 0.018) compared with T-allele carriers. CONCLUSION: Our results show that besides tumor stage, lymph node status, and tumor grade, the GNAS1 T393C status is a novel independent host factor for disease progression in patients with clear cell renal cell carcinoma and provides further evidence for the T393C polymorphism as a general prognostic tumor marker.

Impact of BCL2 polymorphisms on survival in transitional cell carcinoma of the bladder.

PURPOSE: To evaluate the impact of three BCL2 single-nucleotide polymorphisms, i.e., c.-938C>A (rs2279115), c.21G>A (rs1801018), and c.*2203A>G (rs4987853) on survival in patients with transitional cell carcinoma of the bladder. METHODS: We analyzed 179 patients who underwent surgical treatment for bladder cancer at the Clinic of Urology, University Hospital Essen, Germany. Genomic DNA was extracted and genotyped for the polymorphisms. For all polymorphisms, linkage analysis was performed. Kaplan-Meier and Cox regression analyses were used to determine the putative impact of the three polymorphisms on outcome. RESULTS: c.-938C>A and c.21G>A, but not c.*2203A>G, are in strong linkage disequilibrium (D' 0.96). We found a significant association between c.-938C>A and relapse-free survival (p = 0.024) with an allele dose effect. In the same way, c.21G>A had a significant impact on both relapse-free survival (p = 0.009) and progression-free survival (p = 0.012), as well as a pronounced allele dose effect. Regression analysis proved c.21G>A and c.-938C>A, to be an independent risk factor in univariate and multivariable analyses. CONCLUSIONS: In our cohort, both c.-938C>A and c.21G>A showed a significant impact on outcome with TCC of the bladder. Due to the linkage disequilibrium of both SNPs, maybe, only one of them could mediate this effect. In multivariable analysis, however, both proved to be independently associated with overall survival. Contrary to other findings which found the c.-938C>A mainly influencing outcome, our data may suggest that the main effect on TCC could be due to the c.21G>A polymorphism.

Characterization of the splice variant Gbeta3v of the human G-protein Gbeta3 subunit.

A polymorphism (C825T) in the gene of the G-protein Gbeta3 (GNB3) has been the subject of numerous studies which have shown that the 825T-allele is associated with several cardiovascular and metabolic disorders. Furthermore, the T-allele is associated with the occurrence of the splice variant Gbeta3s which has been implicated in the pathogenesis of these disorders. Here, we characterise a novel splice variant of GNB3, termed Gbeta3v, which is generated by alternative splicing of parts from intron 9 as a novel exon 10a. Gbeta proteins belong to the superfamily of propeller proteins composed of seven regular WD-domains. In Gbeta3v, four of these WD-domains are retained but the protein has a novel C terminus. Gbeta3v forms dimers with Ggamma3 and Ggamma12 but these Gbetagamma complexes do not stimulate phospholipase C-beta2. Thus, a physiological role of Gbeta3v remains to be established. Gbeta3v transcripts are detectable in a wide variety of cells and tissues including fibroblasts, B lymphoblasts, retinoblastoma cells, retina, brain, umbilical cord and colon. However, there is no association with an allele of the GNB3 C825T polymorphism, which suggests that Gbeta3v does not contribute to the complex phenotype observed in association with the 825T-allele.

The T393C polymorphism of the G alpha s gene (GNAS1) is a novel prognostic marker in bladder cancer.

The G protein G(alpha)s pathway is linked to proapoptotic signaling in cancer cell lines. To assess the role of the GNAS1 locus encoding G(alpha)s as a genetic factor for disease progression of transitional cell carcinoma (TCC) of the bladder, we genotyped the synonymous T393C polymorphism in 254 patients with TCC (minor allele frequency: 0.43) to examine a potential association between genotypes and disease progression. Using Kaplan-Meier estimates to calculate 5-year probabilities of follow-up, we could show that progression-free survival, metastasis-free survival, and cancer-specific survival was significantly increased in TT genotypes (56%, 84%, 82%) compared with CC genotypes (35%, 53%, 58%). In multivariate Cox proportional hazard analysis, the T393C polymorphism was an independent prognostic factor for clinical outcome. Homozygous CC patients were at highest risk for progression [odds ratio (OR), 1.94; P = 0.020], metastasis (OR, 3.49; P = 0.005), and tumor-related death (OR, 2.49; P = 0.031) compared with TT genotypes. Heterozygous patients had an intermediate risk compatible with a gene-dose effect. Real-time PCR analysis of urothelial tumor tissue as well as adipose and heart tissue revealed that G(alpha)s mRNA expression was highest in TT genotypes, indicating a proapoptotic effect in these genotypes. In conclusion, the GNAS1 T393C status associated with differential G(alpha)s mRNA expression is a novel independent prognostic marker for clinical outcome supporting a functional role of G(alpha)s in bladder cancer progression.

Quantification of G protein Gaalphas subunit splice variants in different human tissues and cells using pyrosequencing.

The G protein Galphas is derived from four alternatively spliced transcripts, two long variants (Galphas(L)+CAG and Galphas(L)-CAG), which include an extra 45-bp segment, and two short variants (Galphas(S)+CAG and Galphas(S)-CAG). The long and short forms differ in each case by splicing in or out of a serine residue encoded at the 3' end of the variable exon 3. The relative expression of all four variants in human tissues is poorly investigated due to experimental limitations. We therefore established a method for reliable relative mRNA quantification of these splice variants based on the Pyrosequencing technology, and determined Galphas transcript ratios in various human tissues and cells. Galphas(S)/Galphas ratio was highest in blood mononuclear cells (0.84 +/- 0.02, n = 16) and lowest in the brain (0.51 +/- 0.14, n = 3). The different ranges resulted from differences in Galphas(S)+CAG ratios, which ranged from a total Galphas ratio of 0.32 +/- 0.07 (n = 12) in heart tissue to 0.57 +/- 0.03 (n = 16) in blood mononuclear cells (p < 0.0001), whereas the Galphas(S)-CAG ratio was rather constant and ranged from 0.22 +/- 0.04 (n = 7) in retinoblastoma cells to 0.27 +/- 0.04 in lymphocytes (p = 0.19). The Galphas(L)+CAG ratio ranged from 0.02 +/- 0.02 in heart tissue to 0.05 +/- 0.01 in retinoblastoma cells, with a varying proportion of Galphas(L)-CAG, which ranged from 0.14 +/- 0.02 in blood mononuclear cells to 0.41 +/- 0.08 in heart tissue. Stimulation of immortalized B lymphoblasts with isoproterenol resulted in significant changes of splice variant ratios. Our data indicate that changes of long and short ratios of Galphas in different tissues affected Galphas(L)-CAG and Gas(S)+CAG rather than Galphas(L)+CAG and Galphas(S-)CAG. Furthermore, stimulation of cells seemed to affect splice variant ratios. These results are, therefore, suggestive of different biological functions of these variants.

Pharmacokinetic, Pharmacodynamic, and Activity Evaluation of TMX-101 in a Multicenter Phase 1 Study in Patients With Papillary Non-Muscle-Invasive Bladder Cancer.

INTRODUCTION/BACKGROUND: Non-muscle-invasive bladder cancer (NMIBC) has a strong tendency to recur despite adjuvant instillations. TMX-101 is a new liquid form of imiquimod for intravesical instillation and has activity in vitro against urothelial carcinoma. The purpose was to analyze the activity of TMX-101 in low-grade NMIBC. Furthermore, pharmacokinetic and pharmacodynamic characteristics and adverse events were evaluated. PATIENTS AND METHODS: A multicenter, prospective phase 1 trial in 7 patients with low-grade NMIBC was conducted. All patients underwent a marker lesion transurethral resection of the bladder tumor and 6 weekly instillations with TMX-101 0.2% or 0.4%. Cystoscopy 2 to 4 weeks after the last instillation evaluated the effect of TMX-101. RESULTS: The effective biologic dose (EBD = complete response [CR] in > 2 patients) could not be defined because none of the patients experienced CR. Maximum plasma concentration was 75.1 ng/mL in the 0.4% dose group. No drug accumulation was observed. In the pharmacodynamic analysis, urinary interleukin 1 receptor agonist (IL-1ra) represents the most sensitive and uniform response after TMX-101 instillation. A total of 87.0% reported at least 1 adverse event. All events were of grade 2 severity or less (Common Terminology Criteria of Adverse Events version 4.02). No clinically significant changes in laboratory parameters or vital signs were observed during or after treatment. CONCLUSION: Toll-like receptor 7 (TLR-7) agonists are effective in urothelial carcinoma in preclinical research. The EBD in this phase 1 study could not be determined because no patient experienced CR. IL-1ra could be valuable as a urinary biomarker in future developments. The safety of TMX-101 has been reconfirmed. New doses, other schedules, and NMIBC subgroups should be tested to define the EBD. A pilot study in carcinoma-in-situ patients is currently ongoing and results are expected shortly.

The prognostic value of cadherin switch in bladder cancer.

Loss of E-cadherin expression and gain of N-cadherin expression ('cadherin switch') is shown to be characteristic in epithelial to mesenchymal transition (EMT), a mechanism associated with cancer progression. Furthermore, the prognostic role of P-cadherin in different cancers is controversial. The aim of this study was to evaluate the prognostic significance of 'cadherin switch' on the gene expression level in bladder cancer. Frozen tissue samples of 181 bladder cancer patients and 7 control individuals were analyzed by quantitative real-time PCR. Kaplan-Meier log-rank test and Cox univariate and multivariate analysis were performed to assess the prognostic relevance of gene expression of E-, N- and P-cadherin. Cox univariate analysis revealed that the decrease of E-cadherin and the gain of N-cadherin gene expression are risk factors for cancer-related death (P=0.087, P=0.005, respectively). Fourteen percent (13/92) of muscle-invasive bladder cancers were N-cadherin- negative. These patients had a significantly poorer prognosis than those with N-cadherin-positive muscle-invasive tumors (P=0.024). P-cadherin gene expression proved to be a significant independent prognostic factor for both cancer-specific and recurrence-free survival (P=0.011, P=0.036). The characteristic 'cadherin switch' between low- and high-stage tumors that we observed and the prognostic significance of E-, N- and P-cadherin suggests the importance of these markers in bladder cancer progression. The poor patient prognosis in N-cadherin-negative muscle-invasive tumors indicates an alternative, N-cadherin-independent way in bladder cancer progression.