Search

Purpose: An investigation of various convolutional neural network (CNN)-based deep learning algorithms was conducted to select the appropriate artificial intelligence (AI) model for calculating the diagnostic performance of bladder tumor classification on cystoscopy images, with the performance of the selected model to be compared against that of medical students and urologists. Methods: A total of 3,731 cystoscopic images that contained 2,191 tumor images were obtained from 543 bladder tumor cases and 219 normal cases were evaluated. A total of 17 CNN models were trained for tumor classification with various hyperparameters. The diagnostic performance of the selected AI model was compared with the results obtained from urologists and medical students by using the receiver operating characteristic (ROC) curve graph and metrics. Results: EfficientNetB0 was selected as the appropriate AI model. In the test results, EfficientNetB0 achieved a balanced accuracy of 81%, sensitivity of 88%, specificity of 74%, and an area under the curve (AUC) of 92%. In contrast, human-derived diagnostic statistics for the test data showed an average balanced accuracy of 75%, sensitivity of 94%, and specificity of 55%. Specifically, urologists had an average balanced accuracy of 91%, sensitivity of 95%, and specificity of 88%, while medical students had an average balanced accuracy of 69%, sensitivity of 94%, and specificity of 44%. Conclusions: Among the various AI models, we suggest that EfficientNetB0 is an appropriate AI classification model for determining the presence of bladder tumors in cystoscopic images. EfficientNetB0 showed the highest performance among several models and showed high accuracy and specificity compared to medical students. This AI technology will be helpful for less experienced urologists or nonurologists in making diagnoses. Image-based deep learning classifies bladder cancer using cystoscopy images and shows promise for generalized applications in biomedical image analysis and clinical decision making.

Purpose: This study investigated the efficacy of intravesical gemcitabine as an alternative to bacillus Calmette-Guérin (BCG) therapy., Materials and Methods: Data were retrospectively collected across seven institutions from February 1999 to May 2023. Inclusion criteria included patients with intermediate- or high-risk non-muscle invasive bladder cancer (NMIBC) who underwent transurethral resection of bladder tumors (TURBT) and received at least four sessions of intravesical gemcitabine or BCG induction therapy. Patient characteristics, complete remission (CR), occurrence, and progression rates were compared., Results: In total, 149 patients were included in this study (gemcitabine, 63; BCG, 86). No differences were apparent between the two groups in baseline characteristics, except for the follow-up period (gemcitabine, 9.2±5.9 months vs. BCG, 43.9±41.4 months, p<0.001). There were no consistent significant differences observed between the two groups in the 3-month (gemcitabine, 98.4% vs. BCG, 95.3%; p=0.848), 6-month (94.9% vs. 90.0%, respectively; p=0.793) and 1-year CR rates (84.2% vs. 83.3%, respectively; p=0.950). Also, there was no significant statistical difference in progression-free survival between the two groups (p=0.953). The occurrence rates of adverse events were similar between the groups (22.2% vs. 22.1%; p=0.989); however, the rate of Clavien-Dindo grade 2 or higher was significantly higher in the BCG group (1.6% vs. 16.3%, respectively; p<0.001)., Conclusions: Intravesical gemcitabine demonstrated efficacy comparable to BCG therapy for the first year in patients with intermediate- and high-risk NMIBC. However, long-term follow-up studies are warranted., Competing Interests: The authors have nothing to disclose., (© The Korean Urological Association.)

Purpose: We investigated the efficacy and optimal dosage of recombinant Bacillus Calmette-Guérin-dltA (rBCG-dltA) in a high-throughput 3D bio-printed bladder cancer-on-a-chip (BCOC) and orthotopic bladder cancer mouse model., Materials and Methods: We fabricated high-throughput BCOC with microfluidic systems, enabling efficient drug screening. The efficacy of rBCG-dltA was evaluated using BCOC by the cell viability assay, monocyte migration assay, and measuring cytokine levels. The anti-tumor effect was compared using the orthotopic bladder cancer mouse model., Results: The cell proliferation rates of T24 and 253J bladder cancer cell lines (mean±standard error) were measured at three days after treatment. In T24 cell line, there was significantly decreased T24 cells compared to control at rBCG 1 multiplicity of infection (MOI) and 10 MOI (30 MOI: 63.1±6.4, 10 MOI: 47.4±5.2, 1 MOI: 50.5±7.5, control: 100.0±14.5, p<0.05). In 253J cell line, a statistically significant decrease in 253J cell count compared to control and mock BCG 30 MOI (30 MOI: 11.2±1.3, 10 MOI: 22.5±2.3, 1 MOI: 39.4±4.7, Mock: 54.9±10.8, control: 100.0±5.6, p<0.05). The migration rates of THP-1 cells showed increased patterns after rBCG-dltA treatment in BCOC. The concentration of tumor necrosis factor-α and interleukin-6 after rBCG-dltA 30 MOI treatment was higher than control in T24 and 253J cell line., Conclusions: In conclusion, rBCG-dltA has the potential to have better anti-tumor activity and immunomodulatory effects than BCG. Furthermore, high-throughput BCOCs have potential to reflect the bladder cancer microenvironment., Competing Interests: The authors have nothing to disclose., (© The Korean Urological Association.)

This study assessed the trends in methotrexate, vinblastine, doxorubicin, and cisplatin (MVAC) and gemcitabine-cisplatin (GC) regimens in Korean patients with metastatic urothelial carcinoma (UC) and compared the side effects and overall survival (OS) rates of the two regimens using nationwide population-based data. The data of patients diagnosed with UC between 2004 and 2016 were collected using the National Health Insurance Service database. The overall treatment trends were assessed according to the chemotherapy regimens. The MVAC and GC groups were matched by propensity scores. Cox proportional hazard analysis and Kaplan-Meier analysis were performed to assess survival. Of 3108 patients with UC, 2,880 patients were treated with GC and 228 (7.3%) were treated with MVAC. The transfusion rate and volume were similar in both the groups, but the granulocyte colony-stimulating factor (G-CSF) usage rate and number were higher in the MVAC group than in the GC group. Both groups had similar OS. Multivariate analysis revealed that the chemotherapy regimen was not a significant factor for OS. Subgroup analysis revealed that a period of ≥ 3 months from diagnosis to systemic therapy enhanced the prognostic effects of the GC regimen. The GC regimen was widely used as the first-line chemotherapy in more than 90% of our study population with metastatic UC. The MVAC regimen showed similar OS to the GC regimen but needed greater use of G-CSF. The GC regimen could be a suitable treatment option for metastatic UC after ≥ 3 months from diagnosis., (© 2023. The Author(s).)

Purpose: This study aimed to evaluate the trend of adjuvant chemotherapy (AC) and neoadjuvant chemotherapy (NAC) in patients who underwent radical nephroureterectomy with bladder cuff excision (NUx) for upper tract urothelial carcinoma (UTUC) to compare the perioperative outcomes and overall survival (OS) between AC and NAC using nationwide population-based data., Materials and Methods: We collected data on patients diagnosed with UTUC and treated with NUx between 2004 and 2016 using the National Health Insurance Service database, and evaluated the overall treatment trends. The AC and NAC groups were propensity score-matched. Cox proportional hazard and Kaplan-Meier analyses were used to assess survival., Results: Of the 8,705 enrolled patients, 6,627 underwent NUx only, 94 underwent NAC, and 1,984 underwent AC. The rate of NUx without perioperative chemotherapy increased from 70.8 to 78.2% (R 2  = 0.632; p < 0.001). The rates of dialysis (p = 0.398), TUR-BT (p = 1.000), and radiotherapy (p = 0.497) after NUx were similar. In the Kaplan-Meier curve, the NAC and AC groups showed no significant difference (p = 0.480). In multivariate analysis, treatment with AC or NAC was not associated with OS (hazard ratio 0.83, 95% confidence interval 0.49-1.40, p = 0.477)., Conclusion: The use of NUx without perioperative chemotherapy has tended to increase in South Korea. Dialysis, TUR-BT, and radiotherapy rates after NUx were similar between the NAC and AC groups. There was no significant difference in OS between the NAC and AC groups. Proper perioperative chemotherapy according to patient and tumor conditions should be determined by obtaining more evidence of UTUC., (© 2022. The Author(s).)

Purpose: Radical cystectomy is the standard of care for muscle-invasive bladder cancer. However, the 5-year survival rate is only about 50%. Therefore, additional treatments are needed. We compared the perioperative outcomes, overall survival, and treatment trends in patients with bladder cancer who underwent radical cystectomy and either neoadjuvant or adjuvant chemotherapy using nationwide population-based data., Materials and Methods: We collected the data of patients diagnosed with bladder cancer treated with radical cystectomy between 2004 and 2016 using the National Health Insurance Service database. We evaluated overall treatment trends. The neoadjuvant chemotherapy and adjuvant chemotherapy groups were matched by propensity score. Cox proportional hazard analysis and Kaplan-Meier analysis were used to assess survival., Results: Of 6134 patients, 1379 underwent adjuvant chemotherapy and 389 underwent neoadjuvant chemotherapy. The utilization rate of neoadjuvant chemotherapy increased from 6.4 to 12.2% from 2004 to 2016 (p = 0.018). The administration rate and number of granulocyte colony-stimulating factor cycles were lower in the neoadjuvant chemotherapy group than in the adjuvant chemotherapy group (p < 0.001 and p = 0.027, respectively). After propensity score matching, the neoadjuvant chemotherapy group had significantly better overall survival than the adjuvant chemotherapy group (p = 0.004). In multivariate analysis, neoadjuvant chemotherapy was associated with better overall survival (hazard ratio 0.77, 95% confidence interval 0.65-0.92, p = 0.003)., Conclusions: Neoadjuvant chemotherapy was associated with lower granulocyte colony-stimulating factor administration and better overall survival than adjuvant chemotherapy. Neoadjuvant chemotherapy should be considered for patients with bladder cancer who undergo radical cystectomy., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

In the past, there was no second-line chemotherapeutic agent suitable for use when urothelial carcinoma (UC) progressed to platinum-resistant UC. However, recently, several new treatment options, such as immune checkpoint inhibitors or targeted therapy have shifted the treatment paradigm regarding second-line therapeutic modalities. A novel class of therapeutic agents includes an antibody-drug conjugate (ADC). ADCs consist of three characteristics: a monoclonal antibody, linker, and payload. The specificity of the monoclonal antibody facilitates the delivery of a linked cytotoxic drug directly into the target tumor cell. Although various ADCs have been developed and approved for use in treating several solid tumors, almost all ADCs for the treatment of UC are still in the testing phase. Here, we review the key points about ADCs and summarize the novel ADCs that are approved or are involved in ongoing studies in UC., Competing Interests: The authors have nothing to disclose., (© The Korean Urological Association.)

Purpose: Intravesical BCG (bacille Calmette-Guérin) instillation in patients with non-muscle-invasive bladder cancer decreases the risk for tumor recurrence and progression. After one BCG product was discontinued, a chronic global BCG shortage occurred. We focused on identifying a reduced dose of BCG that could maintain efficacy and reduce adverse effects., Materials and Methods: We conducted a comprehensive literature search of PubMed, Embase, the Cochrane Library, CINAHL, Web of Science, and Scopus to identify randomized controlled trials through April 2021. The odds ratios (ORs) and 95% confidence intervals (CIs) for the low and standard doses in nine studies were compared. A low dose was defined as a low volume of BCG compared with the standard BCG dose (Armand Frappier, 120 mg; Connaught, 81 mg; Danish 1331, 120 mg; modified Danish 1331, 120 mg; Tokyo 172, 80 mg)., Results: The low-dose group experienced aggravated recurrence (OR, 1.45; 95% CI, 1.09-1.94; p=0.01) but similar progression (OR, 1.11; 95% CI, 0.76-1.62; p=0.59), similar cancer-specific survival (OR, 1.02; 95% CI, 0.60-1.75; p=0.93), similar overall survival (OR, 1.09; 95% CI, 0.76-1.56; p=0.65), favorable adverse effects (OR, 0.41; 95% CI, 0.28-0.62; p<0.0001), and favorable withdrawal (OR, 0.42; 95% CI, 0.25-0.71; p=0.001)., Conclusions: Low-dose BCG had more unfavorable outcomes than did standard-dose BCG in terms of recurrence. Tumor progression, cancer-specific survival, and overall survival were similar between the doses. Low-dose BCG improved adverse effects and withdrawal. In the setting of BCG shortage, low-dose BCG may have strong potential as an alternative., Competing Interests: The authors have nothing to disclose., (© The Korean Urological Association.)

Purpose: The recombinant Bacillus Calmette-Guérin (BCG) containing the streptococcal inhibitor of the complement gene (rBCG- sic ) may be more resistant to antimicrobial peptides and improve internalization; therefore, it can enhance the immunotherapeutic effect of the BCG. Here we determined the optimal dose of rBCG- sic and compared its effectiveness with that of BCG., Materials and Methods: We fabricated a high-throughput 3D-bioprinted bladder cancer-on-a-chip (BCOC) and used it to evaluate the effectiveness of the rBCG- sic in terms of cell viability, cell migration, and cytokine concentrations. Using an orthotopic mouse model, we evaluated its anticancer effect and toxicity via bioluminescence imaging., Results: T24 cell viability was decreased after treatment with rBCG- sic 30 multiplicities of infection (MOI) versus the same dosage of mock BCG (42.8%±6.4% vs. 75.7%±6.6%, p<0.05). THP-1 cell migration was positively correlated with rBCG- sic concentration (2.42-fold at 30MOI, p<0.01). The interleukin-6 concentration of rBCG- sic 30MOI was significantly higher than that of mock BCG 30MOI (11.2±1.3 pg/mL vs. 6.7±0.6 pg/mL, p<0.05). In the orthotopic bladder cancer mouse model, lower tumor volume was observed in the rBCG- sic 30MOI group than in the BCG 30MOI group after 10 days of treatment (p<0.05)., Conclusions: We concluded that rBCG- sic is a useful tool for overcoming BCG unresponsiveness in non-muscle invasive bladder cancer. Additionally, high-throughput BCOC with a microfluidic system can successfully reflect the bladder cancer microenvironment., Competing Interests: The authors have nothing to disclose., (© The Korean Urological Association.)

Immunotherapy of bladder cancer is known to have favorable effects, although it is difficult to determine which patients will show a good response because of the different tumor microenvironments (TME). Here, we developed a bladder cancer-on-a-chip (BCOC) to mimic the TME using three-dimensional (3D) bioprinting and microfluidic technology. We fabricated a T24 and a 5637-cell line-based BCOC that also incorporated MRC-5, HUVEC, and THP-1 cells. We evaluated the effects of TME and assessed the immunologic reactions in response to different concentrations of Bacillus Calmette-Guérin (BCG) via live/dead assay and THP-1 monocytic migration, and concentrations of growth factors and cytokines. The results show that cell viability was maintained at 15% filling density in circle-shaped cell constructs at 20 μL/min microfluidic flow rate. A 3D co-culture increased the proliferation of BCOCs. We found that the appropriate time to evaluate the viability of BCOC, concentration of cytokines, and migration of monocytes was 6 h, 24 h, and three days after BGC treatment. Lastly, the immunotherapeutic effects of BCOC increased according to BCG dosage. To predict effects of immunotherapeutic agent in bladder cancer, we constructed a 3D bioprinted BCOC model. The BCOC was validated with BCG, which has been proven to be effective in the immunotherapy of bladder cancer.

Rapamycin is well-recognized in the clinical therapeutic intervention for patients with cancer by specifically targeting mammalian target of rapamycin (mTOR) kinase. Rapamycin regulates general autophagy to clear damaged cells. Previously, we identified increased expression of messenger RNA levels of NBR1 (the neighbor of BRCA1 gene; autophagy cargo receptor) in human urothelial cancer (URCa) cells, which were not exhibited in response to rapamycin treatment for cell growth inhibition. Autophagy plays an important role in cellular physiology and offers protection against chemotherapeutic agents as an adaptive response required for maintaining cellular energy. Here, we hypothesized that loss of NBR1 sensitizes human URCa cells to growth inhibition induced by rapamycin treatment, leading to interruption of protective autophagic activation. Also, the potential role of mitochondria in regulating autophagy was tested to clarify the mechanism by which rapamycin induces apoptosis in NBR1-knockdown URCa cells. NBR1-knockdown URCa cells exhibited enhanced sensitivity to rapamycin associated with the suppression of autophagosomal elongation and mitochondrial defects. Loss of NBR1 expression altered the cellular responses to rapamycin treatment, resulting in impaired ATP homeostasis and an increase in reactive oxygen species (ROS). Although rapamycin treatment-induced autophagy by adenosine monophosphate-activated protein kinase (AMPK) phosphorylation in NBR1-knockdown cells, it did not process the conjugated form of LC3B-II after activation by unc-51 like autophagy-activating kinase 1 (ULK1). NBR1-knockdown URCa cells exhibited rather profound mitochondrial dysfunctions in response to rapamycin treatment as evidenced by Δψm collapse, ATP depletion, ROS accumulation, and apoptosis activation. Therefore, our findings provide a rationale for rapamycin treatment of NBR1-knockdown human urothelial cancer through the regulation of autophagy and mitochondrial dysfunction by regulating the AMPK/mTOR signaling pathway, indicating that NBR1 can be a potential therapeutic target of human urothelial cancer., (© 2019 Wiley Periodicals, Inc.)

Rapamycin, a mammalian target of rapamycin (mTOR) inhibitor, has significant potential for application in the treatment of urothelial carcinoma (URCa) of the bladder. Previous studies have shown that regulation of the AMP-activated serine/threonine protein kinase (AMPK)-mTOR signaling pathway enhances apoptosis by inducing autophagy or mitophagy in bladder cancer. Alteration of liver kinase B1 (LKB1)-AMPK signaling leads to mitochondrial dysfunction and the accumulation of autophagy-related proteins as a result of mitophagy, resulting in enhanced cell sensitivity to drug treatments. Therefore, we hypothesized that LKB1 deficiency in URCa cells could lead to increased sensitivity to rapamycin by inducing mitochondrial defect-mediated mitophagy. To test this, we established stable LKBI-knockdown URCa cells and analyzed the effects of rapamycin on their growth. Rapamycin enhanced growth inhibition and apoptosis in stable LKB1-knockdown URCa cells and in a xenograft mouse model. In spite of the stable downregulation of LKB1 expression, rapamycin induced AMPK activation in URCa cells, causing loss of the mitochondrial membrane potential, ATP depletion, and ROS accumulation, indicating an alteration of mitochondrial biogenesis. Our findings suggest that the absence of LKB1 can be targeted to induce dysregulated mitochondrial biogenesis by rapamycin treatment in the design of novel therapeutic strategies for bladder cancer., (© 2018 Wiley Periodicals, Inc.)

Purpose: Although Mycobacterium bovis Bacillus Calmette-Guérin (BCG) is the most widely used bladder cancer immunotherapy, innate immune responses involving antimicrobial peptides (AMPs) cause BCG failure and unwanted side effects. Here, we generated genetically modified BCG strains with improved immunotherapeutic effects by adding genes that confer evasion of AMPs., Materials and Methods: We constructed recombinant BCG (rBCG) strains expressing Streptococcal inhibitor of complement (Sic), which confers resistance to human α-defensin-1 and cathelicidin, and d-alanyl carrier protein ligase (dltA), which confers resistance to cationic AMPs. Sic and dltA were separately cloned into the pMV306 plasmid and introduced into BCG via electroporation. Then, the efficacy of the rBCGs was tested in a growth inhibition assay using two bladder cancer cell lines (5637, T24)., Results: We confirmed the presence of cDNA segments corresponding to the Sic and dltA genes in total mRNA of the rBCG strains containing Sic (rBCG-Sic) and dltA (rBCG-dltA), and these rBCGs showed higher survival against AMPs. The growth inhibitory effects of rBCGs on bladder cancer cells were significantly enhanced compared to those of the parent BCG, and THP-1 migration also increased. After 8 h of infection, the levels of internalization were higher in rBCG-infected bladder cancer cells than in BCG-infected cells, and cells infected with rBCGs showed increased release of antitumor cytokines, such as IL-6/12, TNF-α, and INF-γ, resulting in inhibition of bacterial killing and immune modulation via antimicrobial peptides., Conclusions: rBCG-Sic and rBCG-dltA can effectively evade BCG-stimulated AMPs, and may be significantly improved immunotherapeutic tools to treat bladder cancer., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Microarray analysis was used to investigate the lack of identified mammalian target of rapamycin (mTOR) pathway downstream genes to overcome cross-talk at non-muscle invasive high-grade (HG)-urothelial carcinoma (UC) of the bladder, gene expression patterns, gene ontology, and gene clustering by triple (p70S6K, S6K, and eIF4E) small interfering RNAs (siRNAs) or rapamycin in 5637 and T24 cell lines. We selected mTOR pathway downstream genes that were suppressed by siRNAs more than 2-fold, or were up-regulated or down-regulated by rapamycin more than 2-fold. We validated mTOR downstream genes with immunohistochemistry using a tissue microarray (TMA) of 125 non-muscle invasive HG-UC patients and knockout study to evaluate the synergistic effect with rapamycin. The microarray analysis selected mTOR pathway downstream genes consisting of 4 rapamycin up-regulated genes (FABP4, H19, ANXA10, and UPK3A) and 4 rapamycin down-regulated genes (FOXD3, ATP7A, plexin D1, and ADAMTS5). In the TMA, FABP4, and ATP7A were more expressed at T1 and FOXD3 was at Ta. ANXA10 and ADAMTS5 were more expressed in tumors ≤ 3 cm in diameter. In a multivariate Cox regression model, ANXA10 was a significant predictor of recurrence and ATP7A was a significant predictor of progression in non-muscle invasive HG-UC of the bladder. In an ATP7A knock-out model, rapamycin treatment synergistically inhibited cell viability, wound healing, and invasion ability compared to rapamycin only. Activity of the ANXA10 and ATP7A mTOR pathway downstream genes might predict recurrence and progression in non-muscle invasive HG-UC of the bladder. ATP7A knockout overcomes rapamycin cross-talk., Competing Interests: The authors have no potential conflicts of interest to disclose., (© 2017 The Korean Academy of Medical Sciences.)

Bladder cancer is a significant clinical and economic problem. Despite intravesical chemotherapy and immunotherapy, up to 80% of patients with non-muscle-invasive bladder cancer develop recurrent tumors, of which 20% to 30% evolve into more aggressive, potentially lethal tumors. Recently, bladder cancer cells are considered to be mediators of resistance to current therapies and therefore represent strong candidates as biologic targets. No effective chemotherapy has yet been developed for advanced bladder cancer. It is desirable that a drug can be delivered directly and specifically to bladder cancer cells. Stem cells have selective migration ability toward cancer cells, and therapeutic genes can be easily transduced into stem cells. In suicide gene therapy for cancer, stem cells carry a gene encoding a carboxylesterase (CE) enzyme that transforms an inert CPT-11 prodrug into a toxic SN-38 product, a topoisomerase 1 inhibitor. In immunodeficient mice, systemically transplanted HB1.F3.CE stem cells migrated toward the tumor implanted by the TCCSUP bladder cancer cell line, and, in combination with CPT-11, the volume of tumors was significantly reduced. These findings may contribute to the development of a new selective chemotherapeutic strategy against bladder cancer. Mol Cancer Ther; 15(6); 1201-7. ©2016 AACR., (©2016 American Association for Cancer Research.)

Objective: To confirm the role of cathelicidin (LL-37) in the internalization of bacille Calmette-Guérin (BCG) into bladder cancer cells., Methods: Enzyme-linked immunosorbent assay and reverse transcription polymerase chain reaction analysis evaluated the changes in protein and messenger ribonucleic acid (RNA) expression with BCG incubation after LL-37 pretreatment in 5637 and T24 human bladder cancer cells. The internalization rate was evaluated by a double immunofluorescence assay, and confocal microscopy confirmed the function of LL-37 in BCG internalization. We also investigated the difference in internalization rates and cell viability between LL-37, anti-LL-37 antibody, and LL-37 plus anti-LL-37 antibody., Results: The levels of LL-37 increased after BCG exposure in bladder cancer cells in dose- and time-dependent manners. Increasing LL-37 levels using recombinant LL-37 protein further dose dependently decreased BCG internalization in both cell lines. The internalization rates of BCG after LL-37 instillation were lower compared with the controls, and the internalization rate of BCG after anti-LL-37 antibody instillation was significantly higher compared with the controls in both cell lines (P <.05). Viability of LL-37 plus BCG group was higher compared with the BCG-alone group. The anti-LL-37 antibody plus BCG group had decreased cell viability compared with the BCG-alone group in both cell lines., Conclusion: Bladder cancer cells produce cathelicidin when infected with BCG and upregulate cathelicidin to defend against BCG by inhibiting its internalization. Blocking the action of cathelicidin may increase the internalization and effectiveness of BCG in reducing bladder cancer cell proliferation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

We investigated how the dual inhibition of the molecular mechanism of the mammalian target of the rapamycin (mTOR) downstreams, P70S6 kinase (P70S6K) and eukaryotic initiation factor 4E (eIF4E), can lead to a suppression of the proliferation and progression of urothelial carcinoma (UC) in an orthotopic mouse non-muscle invasive bladder tumor (NMIBT) model. A KU-7-luc cell intravesically instilled orthotopic mouse NMIBC model was monitored using bioluminescence imaging (BLI) in vivo by interfering with different molecular components using rapamycin and siRNA technology. We then analyzed the effects on molecular activation status, cell growth, proliferation, and progression. A high concentration of rapamycin (10 µM) blocked both P70S6K and elF4E phosphorylation and inhibited cell proliferation in the KU-7-luc cells. It also reduced cell viability and proliferation more than the transfection of siRNA against p70S6K or elF4E. The groups with dual p70S6K and elF4E siRNA, and rapamycin reduced tumor volume and lamina propria invasion more than the groups with p70S6K or elF4E siRNA instillation, although all groups reduced photon density compared to the control. These findings suggest that both the mTOR pathway downstream of eIF4E and p70S6K can be successfully inhibited by high dose rapamycin only, and p70S6K and Elf4E dual inhibition is essential to control bladder tumor growth and progression.

Purpose: This study aimed to confirm the utility of the European Organization for Research and Treatment of Cancer (EORTC) and the Spanish Urological Club for Oncological Treatment (CUETO) scoring systems and to determine which model is preferred as a prognostic model in Korean patients with non-muscle-invasive bladder cancer., Materials and Methods: Between 1985 and 2011, 531 patients who were treated by transurethral resection of bladder cancer were retrospectively analyzed by use of the EORTC and CUETO models. Statistically, we performed Kaplan-Meier survival analysis; calculated Harrell's concordance index, receiver operating characteristic (ROC) curve, and cutoff values; and performed univariate and multivariate Cox proportional hazards regression analyses., Results: For risk of recurrence, with the use of the EORTC model, all groups had statistically significant differences except between the group with a score of 0 and the group with a score of 1-4. With the use of the CUETO model, all groups differed significantly. For risk of progression, with the use of the EORTC model, significant differences were observed between all groups except between the group with a score of 2-6 and the group with a score of 7-13. With the use of the CUETO model, a significant difference was observed between the group with a score of 0 and the other groups. The concordance index of the EORTC and CUETO models was 0.759 and 0.836 for recurrence and 0.704 and 0.745 for progression, respectively. The area under the ROC curve for the EORTC and CUETO models was 0.832 and 0.894 for recurrence and 0.722 and 0.724 for progression, respectively., Conclusions: Both scoring systems, especially the CUETO model, showed value in predicting recurrence and progression in Korean patients, which will help in individualizing treatment and follow-up schedules.

We investigated whether inhibiting phosphorylated p70S6K (p-p70S6K) suppresses the proliferation and growth of noninvasive low-grade urothelial carcinoma (LG-URCa) in vitroand whether p-p70S6K can serve as a predictive biomarker for the recurrence of noninvasive LG-URCa of the bladder in patients. We constructed a tissue microarray (TMA) for 95 LG-URCa and 35 benign urothelium samples and performed immunohistochemical staining for p-p70S6K and p-4E-BP1. A Cox regression model was used to investigate the predictive factors for recurrence of LG-URCa. We investigated the dose-dependent antiproliferative effect of rapamycin, its antiproliferative effect and the growth-inhibition effect of p70S6K siRNA transfection in RT4 and 253J cell lines. The pT1 staged group (P < 0.05; hazard ratio (HR), 2.415) and the high p-p70S6K staining group (P < 0.05; HR, 2.249) were independent factors for predicting recurrence. Rapamycin inhibited RT4 and 253J cell proliferation in a dose-dependent manner (r = -0.850, P< 0.001 in RT4 cells; r = -0.835, P< 0.001 in 253J cells). RT4 and 253J cell proliferation and growth were inhibited by the transfection of p70S6K siRNA and rapamycin, respectively (P < 0.05). Transfection of p70S6K siRNA resulted in inhibitory effects on cell proliferation and growth that were similar to those of rapamycin. Our results suggest that inhibiting p70S6K phosphorylation is important to prevent recurrence and that p70S6K phosphorylation can be used as a molecular biomarker to predict recurrence of certain LG-URCa of the bladder.

We established an orthotopic non-muscle invasive bladder cancer (NMIBC) mouse model expressing the mammalian target of the rapamycin (mTOR) signaling pathway. After intravesical instillation of KU-7-lucs (day 0), animals were subsequently monitored by bioluminescence imaging (BLI) on days 4, 7, 14, and 21, and performed histopathological examination. We also validated the orthotopic mouse model expressing the mTOR signaling pathway immunohistochemically. In vitro BLI photon density was correlated with KU-7-luc cell number (r (2) = 0.97, P < 0.01) and in vivo BLI photon densities increased steadily with time after intravesical instillation. The tumor take rate was 84.2%, formed initially on day 4 and remained NMIBC up to day 21. T1 photon densities were significantly higher than Ta (P < 0.01), and histological tumor volume was positively correlated with BLI photon density (r (2) = 0.87, P < 0.01). The mTOR signaling pathway-related proteins were expressed in the bladder, and were correlated with the western blot results. Our results suggest successful establishment of an orthotopic mouse NMIBC model expressing the mTOR signaling pathway using KU-7-luc cells. This model is expected to be helpful to evaluate preclinical testing of intravesical therapy based on the mTOR signaling pathway against NMIBC.

Objective: To investigate whether secretion of human β-defensin 2 (HBD-2) is induced by bacillus Calmette-Guérin (BCG) and to determine whether HBD-2 affects BCG internalisation in bladder cancer cells., Materials and Methods: Reverse transcription-polymerase chain reaction analysis was used to determine whether HBD-2 mRNA increases after incubation with BCG. HBD-2 proteins in 5637 and T24 human bladder cancer cell lines were assayed by enzyme-linked immunosorbent assay. The internalisation rate was evaluated by double immunofluorescence assay and confocal microscopy to test the optimal dose of HBD-2 for BCG internalisation. We also investigated the difference in internalisation rates and cell viability between recombinant HBD-2 protein, anti-HBD-2 antibody, and HBD-2 plus anti-HBD-2 antibody pretreatments., Results: BCG induced HBD-2 mRNA expression and HBD-2 production dose and time-dependently in bladder cancer cells and affected BCG internalisation. Pretreatment with recombinant HBD-2 protein lowered internalisation of BCG dose-dependently. Moreover, anti-HBD-2 antibody prevented the effect of HBD-2 on BCG internalisation in bladder cancer cells. The internalisation rate of BCG pretreated with anti-HBD-2 antibody was higher than that in the control in 5637 (P < 0.01) and T24 cells (P < 0.05). The BCG internalisation rate in cells pretreated with anti-HBD-2 antibody plus recombinant HBD-2 protein was higher than that in the control in 5637 (P < 0.01) and T24 cells (P < 0.05). Mycobacterium bovis BCG decreased bladder cancer cell viability, and anti-HBD-2 antibody prevented the inhibitory role of HBD-2 on the anti-proliferative effects of M. bovis BCG in bladder cancer cells, Conclusion: Bladder cancer cells produce HBD-2 when they are infected by BCG to defend themselves against BCG internalisation, which plays an important role during the initiation and propagation of the immunotherapeutic response in bladder cancer cells., (© 2013 The Authors. BJU International © 2013 BJU International.)

The aim of this study was to determine the impact that age and comorbidity status have on both overall and bladder cancer-specific survival of bladder cancer patients. We obtained medical information pertaining to a population of 528 patients with newly diagnosed bladder cancer from Chung-Ang University Hospital cancer registry. The Adult Comorbidity Evaluation-27 (ACE-27) test, which has been previously validated in adult cancer patients, was used to assess comorbidity. We evaluated differences in the demographic and clinical characteristics of included patients, as well as differences in the treatments they received after categorizing them by age. The median age at the time of bladder cancer diagnosis of the entire cohort was 63 years, and the median follow-up time was 97 months. Of the 528 patients who were included in our study, 303 had at least one comorbid condition and 249 died during the follow-up period. When patients were stratified by age, we found that older patients had a higher proportion of severe comorbidities (P < 0.01) than younger patients, and that a lower proportion of them underwent radical cystectomy for invasive bladder cancer (IBC) (P < 0.01). By multivariate analysis, we found that older age was predictive of lower overall survival (OS) and bladder cancer-specific survival (BCSS) rates among patients with superficial bladder cancer (SBC) and of lower OS rates among patients with IBC. We also found that moderate-severe comorbidity status and treatment through a bladder-conserving approach were predictive of lower OS and cancer-specific survival rates among patients with IBC. The disparity between overall deaths and bladder cancer deaths was shown in SBC and increased along with age and higher comorbidity. Age and comorbidity were found to be independent predictive factors of OS and BCSS among bladder cancer patients, and explained the disparity that we observed between overall bladder cancer-specific mortality rates.

Introduction: We investigated the clinical significance of preoperatively-unsuspected ureteral involvement of cancer detected on intraoperative frozen section analysis of ureteral margins during radical cystectomy., Patients and Methods: We performed a retrospective study of 115 patients who received radical cystectomy for locally-advanced but node-negative transitional cell carcinoma of the bladder between 1995 and 2002 by reviewing their records., Results: Of 115 total patients, 5 (4.3%) demonstrated carcinoma in situ at the ureteral margin with 2 of 5 having a positive margin on final pathologic analysis. Meanwhile, only 3 of 115 total patients (2.6%) later showed upper urinary tract recurrence at a median interval of 30 months after cystectomy. And all 3 patients had intramural or juxtavesical ureter involvement of disease (p = 0.006), while not demonstrating carcinoma in situ in ureteral margins resected during cystectomy. On multivariate analysis, only the stage and grade of bladder cancer along with pathologic feature of vascular invasion were observed to be independent prognostic predictors of disease-specific survival., Conclusions: Cancer involvement of the distal ureteral margin detected through intraoperative frozen section analysis may not be a significant factor regarding upper tract recurrence and survival of patients with locally-advanced bladder cancer after radical cystectomy. Upper tract recurrence may be more prone to occur in patients with cancer involvement at the intramural or juxtavesical ureter.