Assessment of efficacy, safety and quality of life of 55 patients with metastatic renal cell carcinoma treated with temsirolimus: a single-center experience in Japan.

BACKGROUND: To evaluate experience of the use of temsirolimus for metastatic renal cell carcinoma (mRCC) in a single center in Japan. METHODS: This study included 55 consecutive patients with mRCC who received temsirolimus in a routine clinical setting, and retrospectively reviewed the comprehensive outcomes of these patients. RESULTS: Of the 55 patients, 20 had a Karnofsky performance status of ≤80, and 5, 41 and 9 were classified into favorable, intermediate and poor risk groups, respectively, according to the Memorial Sloan-Kettering Cancer Center model. Initially, 25 mg of temsirolimus was applied weekly; however, dose modification was required in 19 patients, resulting in a relative dose intensity of 90.5 % throughout this series. As the best responses to temsirolimus, 4, 44 and 7 were judged to have a partial response, stable disease and progressive disease, respectively. The median progression-free survival (PFS) and overall survival (OS) of these patients following the introduction of temsirolimus was 7.0 and 25.0 months, respectively. Of several factors examined, only the pretreatment C-reactive protein level was shown to be independently associated with both PFS and OS. The common adverse events related to temsirolimus corresponding to ≥grade 3 were anemia in 4, thrombocytopenia in 3, stomatitis in 3 and hyperglycemia in 3. Quality of life analysis using 36-Item Short Form showed that there were no significant differences in any scale scores between surveys performed before and 3 months after the introduction of temsirolimus. CONCLUSIONS: Temsirolimus was well tolerated and facilitated comparatively favorable cancer control in Japanese patients with mRCC.

Suppressed tumour growth and enhanced chemosensitivity by RNA interference targeting Aurora-A in the PC3 human prostate cancer model.

OBJECTIVES: To investigate the inhibitory effects of Aurora-A expression in prostate cancer cells on their growth and chemosensitivity. PATIENTS AND METHODS: Aurora-A expression in radical prostatectomy specimens obtained from 193 patients were evaluated by immunohistochemical staining. We then established PC3 cells in which the expression vector containing short-hairpin RNA (shRNA) targeting Aurora-A was introduced (PC3/sh-A). The growth and the sensitivity to docetaxel in PC3/sh-A were compared with those in PC3 transfected with control vector alone (PC3/C). RESULTS: Immunohistochemistry showed that there were various levels of Aurora-A expression in most prostate cancer tissues, and the expression levels of Aurora-A in prostate cancer were significantly related to Gleason score. Expression levels of both Aurora-A mRNA and protein in PC3/sh-A were approximately 20% of those in PC3/C. In vitro growth of PC3/sh-A was significantly worse than that of PC3/C, and the proportion of PC3/sh-A in the G2-M phase was significantly greater than that of PC3/C. The 50% inhibitory concentration of docetaxel in PC3/sh-A decreased by 67% compared with that in PC3/C. Tumour volume in nude mice injected with PC3/sh-A was significantly smaller than that with PC3/C. Furthermore, treatment of nude mice bearing PC3/sh-A tumour with docetaxel (10 mg/kg, once weekly for 4 weeks) achieved a synergistic cytotoxic effect, despite the lack of an enhanced antitumour effect of docetaxel on PC3/C tumours. CONCLUSIONS: The suppression of Aurora-A using shRNA could be a useful therapeutic strategy against androgen-independent prostate cancer, through growth inhibition as well as enhanced chemosensitivity.

Assessment of voiding function of orthotopic neobladders in elderly patients with long-term survival.

OBJECTIVE: To compare the voiding status in elderly patients (aged >or=80 years) with that in younger patients undergoing orthotopic neobladder substitution during long-term survival. PATIENTS AND METHODS: The voiding status was assessed in 111 patients (ileal neobladder in 62, ascending colonic neobladder in 14, sigmoid colonic neobladder in 21 and ileocolonic neobladder in 14) who lived for >5 years after radical cystectomy with an orthotopic neobladder, using a self-completed questionnaire and uroflowmetry. According to the age at the time of these assessments, patients were divided into two groups (group 1, <80 years, 94; group 2, >or=80 years, 17). The voiding status was compared between the groups. RESULTS: In all, 78 patients (92%) in group 1 and 16 (94%) in group 2 were capable of spontaneous voiding. In group 1 and 2, respectively, daytime continence was achieved by 67 (74%) and 12 (75%) patients, but night-time continence was achieved by 54 (60%) and six (38%), although the difference was not statistically significant. In groups 1 and 2, respectively, the median maximum flow rate was 13.3 and 11.7 mL/s and the median postvoid residual urine volume was 19 and 18 mL. The only statistically significant difference was for voiding posture, assessed in men. CONCLUSIONS: There was no significant difference in voiding status of patients with orthotopic neobladders except for voiding posture between patients aged <80 or carefully selected elderly patients aged >or=80 years during long-term survival. However, night-time continence might be clinically worse in the elderly than in the younger group.

Expression of urokinase-type plasminogen activator system in prostate cancer: correlation with clinicopathological outcomes in patients undergoing radical prostatectomy.

The objective of this study was to evaluate the expression levels of urokinase-type plasminogen activator (uPA) system in radical prostatectomy (RP) specimens in order to clarify the significance of the uPA system in prostate cancer. Expression levels of uPA, uPA receptor (uPAR), plasminogen activator inhibitor-1 (PAI-1), and PAI-2 in RP specimens obtained from 153 patients with clinically organ-confined prostate cancer who had not received any neoadjuvant therapies were evaluated by immunohistochemical staining. Various expression levels of uPA, uPAR, PAI-1, and PAI-2 were noted in the majority of prostate cancer specimens. Expression levels of uPA and uPAR were significantly associated with major prognostic indicators, including pathological stage, Gleason score, lymphatic invasion, surgical margin status and lymph node metastasis. However, PAI-1 expression was related to only pathological stage and surgical margin status, and there was no significant association between the expression level of PAI-2 and several parameters examined. Despite the lack of prognostic significance in PAI-2 expression, biochemical recurrence-free survival of patients with strong uPA, uPAR, and PAI-1 expression was significantly lower than that of those with weak uPA, uPAR, and PAI-1 expression, respectively. Furthermore, strong expression of uPA in addition to a Gleason score, positive surgical margin, and lymph node metastasis could be independent predictors for biochemical recurrence after RP. These findings suggest that the uPA system may be involved in the progression of prostate cancer, and that the expression level of uPA in prostate cancer tissue could be used as a useful predictor of biochemical recurrence in patients undergoing RP.

Bonding-Based Wafer-Level Vacuum Packaging Using Atomic Hydrogen Pre-Treated Cu Bonding Frames.

A novel surface activation technology for Cu-Cu bonding-based wafer-level vacuum packaging using hot-wire-generated atomic hydrogen treatment was developed. Vacuum sealing temperature at 300 °C was achieved by atomic hydrogen pre-treatment for Cu native oxide reduction, while 350 °C was needed by the conventional wet chemical oxide reduction procedure. A remote-type hot-wire tool was employed to minimize substrate overheating by thermal emission from the hot-wire. The maximum substrate temperature during the pre-treatment is lower than the temperature of Cu nano-grain re-crystallization, which enhances Cu atomic diffusion during the bonding process. Even after 24 h wafer storage in atmospheric conditions after atomic hydrogen irradiation, low-temperature vacuum sealing was achieved because surface hydrogen species grown by the atomic hydrogen treatment suppressed re-oxidation. Vacuum sealing yield, pressure in the sealed cavity and bonding shear strength by atomic hydrogen pre-treated Cu-Cu bonding are 90%, 5 kPa and 100 MPa, respectively, which are equivalent to conventional Cu-Cu bonding at higher temperature. Leak rate of the bonded device is less than 10-14 Pa m³ s-1 order, which is applicable for practical use. The developed technology can contribute to low-temperature hermetic packaging.

Prognostic analysis of Japanese men with metastatic germ cell tumors showing favorable response to bleomycin, etoposide and cisplatin as first-line chemotherapy.

The objective of this study was to evaluate the efficacy of first-line bleomycin, etoposide and cisplatin (BEP) chemotherapy in Japanese patients with metastatic germ cell tumors (GCTs). Between 1996 and 2006, 88 male patients with metastatic GCTs were treated with first-line BEP at our institution. Of these 88, 47 (16, seminoma; 31, nonseminoma), who did not receive high-dose chemotherapy following BEP because of the normalization of serum tumor markers, were included in this study. The primary site was the testis in 42 patients, retroperitoneum in 3, and mediastinum in 2. The full-dose regimen used for BEP consisted of cisplatin 20 mg/m2 on days 1 to 5, etoposide 100 mg/m2 on days 1 to 5, and bleomycin on days 2, 9 and 16. Therapeutic outcome was assessed according to several clinicopathological parameters. Following 2 to 4 cycles of BEP (median, 4 cycles), alpha-fetoprotein, beta-human chorionic gonadotropin and lactate dehydrogenase were normalized in all 47 patients. Eighteen patients (38.3%) achieved a complete response (CR) after BEP alone, while BEP resulted in a partial response and stable disease in the remaining 23 (48.9%) and 6 (12.8%), respectively. In addition, surgical resection of the residual tumors following BEP was performed in 15 patients, of whom 12 (80.0%) and 3 (20.0%) achieved pathological and surgical CR, respectively. At a median follow-up of 27 months, all patients were alive; however, disease recurrence occurred in 5 (seminoma, 1; nonseminoma, 4), and all these 5 were subsequently treated with high-dose chemotherapy as salvage therapy. In this series, 1-, 3- and 5-year recurrence-free survival rates were 95.0, 91.4 and 79.2%, respectively, and, there was no significant difference in recurrence-free survival between patients with seminoma and those with nonseminoma. These findings suggested that patients with metastatic GCTs, regardless of histological subtype (i.e., seminoma or nonseminoma), who showed favorable response to first-line BEP chemotherapy, could achieve an excellent prognostic outcome.

Stable hematopoietic recovery after peripheral blood stem cell transplantation in patients receiving high-dose chemotherapy for advanced germ cell tumors.

AIM: The objective of this study was to evaluate the utility of CD34-positive peripheral blood stem cell transplantation (PBSCT) in the hematopoietic recovery, in patients receiving high-dose chemotherapy (HDCT), for advanced germ cell tumor (GCT). MATERIALS AND METHODS: This study included 41 patients with advanced GCT, who were treated with HTCT combined with PBSCT. PBSCs were harvested after conditioning chemotherapy followed by the administration of granulocyte colony-stimulating factor. A retrospective analysis of a total of 86 PBSCTs was carried out focusing on the effects of several factors, including age (<35 years versus > or = 35 years), CD34-positive cell dose (<5.0 x10(6)/kg versus > or =5.0 x10(6)/kg), indication of HDCT (first-line versus salvage) and previous history of HDCT before PBSCT (with versus without), on hematopoietic recovery after PBSCT. RESULTS: The median number of CD34-positive PBSCs collected during a single apheresis and the median cumulative number of CD34-positive PBSCs from each patient were 8.3x10(6)/kg and 23.2x10(6)/kg, respectively. The median number of PBSCT performed in each patient was two and the median number of CD34-positive cells transplanted during a single course was 5.7x10(6)/kg. The median recovery times to white blood cells (WBC) greater than 500/microl, 1000/microl and 2000/microl were 8, 9 and 10 days, respectively, following PBSCT, while that to neutrophils greater than 500/microl and that to platelets greater than 50000/microl were 9 and 13 days, respectively, following PBSCT Only the recovery time to platelet count greater than 50000/microl was significantly affected by age; however, there were no significant differences in the recovery of WBC, neutrophils and platelets in relation to several parameters examined. CONCLUSION: These findings suggest that CD34-positive PBSCT may facilitate stable hematopoietic recovery after HDCT in patients with advanced GCT, and that HDCT, if combined with PBSCT, could be performed with comparative safety in such patients, irrespective of their individual characteristics.

[Pathological prognostic factors of renal pelvic and ureteral cancer].

OBJECTIVE: To determine the pathologic risk factors after nephroureterectomy in patients with urothelial carcinoma of the renal pelvis and ureter. PATIENTS AND METHODS: We investigated the clinicopathological features of 131 patients (94 males and 37 females) with urothelial carcinoma of the renal pelvis and ureter who underwent nephroureterectomy at our department and related facilities from August, 1994 to August, 1997. The mean age of the patients was 68 years, ranging from 24 to 86 years. RESULTS: The 1-, 3- and 5-year cause-specific survival rates (Kaplan-Meier's method) for all of the patients were 91.8%, 76.7%, and 67.8%, respectively. The significant prognostic factors for survival rates by univariate analysis using the log rank test were tumor stage, infiltration pattern, lymphatic invasion, vessel invasion and lymph node metastasis. On the other hand, multivariate analysis using Cox proportional hazards regression model showed the most influential prognostic factors to be vessel invasion and tumor stage. CONCLUSIONS: From these results, in urothelial carcinoma of the renal pelvis and ureter underwent nephroureterectomy, we suggested that vessel invasion and tumor stage were the independent prognostic factors.

[Is second course intravesical Bacillus Calmette-Guerin therapy for recurrent carcinoma in situ of the bladder useful?].

We evaluated the usefulness of second course intravesical bacillus Calmette-Guerin (BCG) therapy for carcinoma in situ (CIS) of the bladder that failed to respond to the initial BCG therapy. Between January 1995 and December 2000, 185 patients with CIS of the bladder underwent an initial 6- or 8-week course of intravesical BCG instillation with an average follow-up period of 40.9 months (range: 3.8 to 94.8 months). Of the 185 patients, 160 (86.5%) completely responded to an initial course of BCG therapy. During follow up, 49 (30.6%) of the complete responders had recurrent transitional cell carcinoma. Overall, 9 (36.0%) of the 25 patients who did not respond completely to the initial 6- or 8-week course of BCG therapy and 22 (44.9%) of the 49 who had recurrent tumor after initial complete response, a total of 31 patients received the second course intravesical BCG therapy. Of the 9 incomplete responders, 8 (88.9%) achieved a complete response after the second course BCG therapy. With an average follow-up period of 39.6 months (range: 2.8 to 62.2 months), 2 (22.2%) of the 8 had recurrence. On the other hand, 17 (77.3%) of the 22 with recurrent tumor after the initial complete response developed recurrence with an average follow-up period of 14.1 months (range: 2.8 to 55.2 months). Seven (31.8%) of the 17 patients had disease progression to muscle invasion. Subsequently, cystectomy was done in 10 (58.8%) and radiation in 1 (5.9%). Our results suggest that a selected group of incomplete responders with initial BCG therapy may benefit from continued second course BCG. However, in patients who had recurrence after initial BCG success, the benefits of second course BCG therapy are limited. Careful surveillance and aggressive therapy on optimal timing are mandatory.

Hsp27 Inhibition with OGX-427 Sensitizes Non-Small Cell Lung Cancer Cells to Erlotinib and Chemotherapy.

Non-small cell lung cancer (NSCLC) is the most frequent cause of death from cancer worldwide. Despite the availability of active chemotherapy regimens and EGFR tyrosine kinase inhibitors, all advanced patients develop recurrent disease after first-line therapy. Although Hsp27 is a stress-induced chaperone that promotes acquired resistance in several cancers, its relationship to treatment resistance in NSCLC has not been defined. Understanding adaptive responses of acquired resistance will help guide new strategies to control NSCLC. Hsp27 levels were evaluated in an HCC827 erlotinib-resistant-derived cell line (HCC-827Resistant), and sensitivity to erlotinib was examined in Hsp27-overexpressing A549 cells. The role of Hsp27 in both erlotinib and cytotoxic treatment resistance was evaluated in HCC-827 and A549 NSCLC cells using the Hsp27 antisense drug OGX-427. The effect of OGX-427 in combination with erlotinib was also assessed in mice bearing A549 xenografts. Hsp27 is induced by erlotinib and protects NSCLC cells from treatment-induced apoptosis, whereas OGX-427 sensitizes NSCLC cells to erlotinib. Interestingly, increased resistance to erlotinib was observed when Hsp27 was increased either in HCC827 erlotinib-resistant or overexpressing A549 cells. Combining OGX-427 with erlotinib significantly enhanced antitumor effects in vitro and delayed A549 xenograft growth in vivo. OGX-427 also significantly enhanced the activity of cytotoxic drugs used for NSCLC. These data indicate that treatment-induced Hsp27 contributes to the development of resistance, and provides preclinical proof-of-principle that inhibition of stress adaptive pathways mediated by Hsp27 enhances the activity of erlotinib and chemotherapeutics.

Clusterin facilitates stress-induced lipidation of LC3 and autophagosome biogenesis to enhance cancer cell survival.

We define stress-induced adaptive survival pathways linking autophagy with the molecular chaperone clusterin (CLU) that function to promote anticancer treatment resistance. During treatment stress, CLU co-localizes with LC3 via an LIR-binding sequence within autophagosome membranes, functioning to facilitate LC3-Atg3 heterocomplex stability and LC3 lipidation, and thereby enhance autophagosome biogenesis and autophagy activation. Stress-induced autophagy is attenuated with CLU silencing in CLU(-/-) mice and human prostate cancer cells. CLU-enhanced cell survival occurs via autophagy-dependent pathways, and is reduced following autophagy inhibition. Combining CLU inhibition with anticancer treatments attenuates autophagy activation, increases apoptosis and reduces prostate cancer growth. This study defines a novel adaptor protein function for CLU under stress conditions, and highlights how co-targeting CLU and autophagy can amplify proteotoxic stress to delay cancer progression.

Sequential use of mammalian target of rapamycin inhibitors in patients with metastatic renal cell carcinoma following failure of tyrosine kinase inhibitors.

The aim of the study is to evaluate the clinical experience of the sequential use of mammalian target of rapamycin inhibitors (mTORIs) for metastatic renal cell carcinoma (mRCC) refractory to tyrosine kinase inhibitors (TKIs). This study retrospectively investigated the clinical outcomes in a total of 83 consecutive Japanese patients with mRCC who were treated with either everolimus or temsirolimus following the failure of sorafenib and/or sunitinib. Of the 83 patients, 15, 61, and 7 were classified into favorable-, intermediate-, and poor-risk groups, respectively, according to the Memorial Sloan-Kettering Cancer Center model, and 47 and 36 patients were administered mTORIs as second- and third-line therapy, respectively. As the best responses to mTORIs, 6, 53, and 24 were judged to have a partial response, stable disease, and progressive disease, respectively. The median progression-free survival (PFS) and overall survival (OS) of these patients following the introduction of mTORIs were 5.8 and 20.4 months, respectively. Of the several factors examined, liver metastasis and pretreatment C-reactive protein (CRP) level were shown to be independently associated with PFS, while only pretreatment CRP level had an independent impact on OS. Adverse events related to mTORIs corresponding to ≥grade 3 were observed in 26 patients, including anemia in 7, pneumonitis in 7, neutropenia in 4, and stomatitis in 3. Despite the low response rate, mTORIs are well tolerated and could provide comparatively favorable prognostic outcomes in Japanese patients with mRCC after the failure of TKIs.

Significance of random bladder biopsies in non-muscle invasive bladder cancer.

BACKGROUND/AIMS: To evaluate retrospectively the clinical outcome of random bladder biopsies in patients with non-muscle invasive bladder cancer (NMIBC) undergoing transurethral resection (TUR). PATIENTS AND METHOD: This study included 234 consecutive patients with NMIBC who underwent random biopsies from normal-appearing urothelium of the bladder, including the anterior wall, posterior wall, right wall, left wall, dome, trigone and/or prostatic urethra, during TUR. RESULT: Thirty-seven patients (15.8%) were diagnosed by random biopsies as having urothelial cancer. Among several factors available prior to TUR, preoperative urinary cytology appeared to be independently related to the detection of urothelial cancer in random biopsies on multivariate analysis. Urinary cytology prior to TUR gave 50.0% sensitivity, 91.7% specificity, 56.8% positive predictive value and 89.3% negative predictive value for predicting the findings of the random biopsies. CONCLUSION: Biopsies of normal-appearing urothelium resulted in the additional detection of urothelial cancer in a definite proportion of NMIBC patients, and it remains difficult to find a reliable alternative to random biopsies. Collectively, these findings suggest that it would be beneficial to perform random biopsies as part of the routine management of NMIBC.

ß4-integrin-mediated cytotoxic activity of AexU in human prostate cancer PC3 cells.

The present study aimed to characterize the cytotoxic activity of AexU, an effector-mediating type three secretion system (TTSS) of gram-negative bacteria, in human prostate cancer cells, focusing on the association with ß4-integrin expression. The cytotoxic effects of AexU either alone or in combination with chemotherapeutic agents were evaluated using several human prostate cancer cell lines. Human prostate cancer PC3 cells, in which an expression vector containing siRNA targeting ß4-integrin had been introduced, were established (PC3/sh-In), and the cytotoxic effects of AexU on the PC3/sh-In cells were compared with the PC3 cells that were transfected with a control vector (PC3/C). The expression levels of ß4-integrin in the PC3 cells were markedly higher compared with those in the LNCaP or DU145 cells, and the cytotoxic effects of AexU in the PC3 cells were more pronounced compared with those in the LNCaP or DU145 cells. The sensitivity of the PC3 cells to docetaxel and cisplatin was significantly enhanced following treatment with AexU, resulting in a decrease in the IC50 of the two agents by ~90%. The cytotoxic effect of AexU in the PC3/C cells was more marked compared with that in the PC3/sh-In cells, and the phosphorylation of Akt in the PC3/C cells appeared to be significantly more inhibited by the treatment with AexU compared with the PC3/sh-In cells. In conclusion, treatment with AexU may be a useful therapeutic option for prostate cancer when ß4-integrin is overexpressed. The treatment appears to exert its effects through growth inhibition and by enhancing the sensitivity of the cancer cells to chemotherapeutic agents.

Blocked autophagy using lysosomotropic agents sensitizes resistant prostate tumor cells to the novel Akt inhibitor AZD5363.

PURPOSE: Prostate cancer development is often associated with deletion or silencing of tumor suppressor phosphatase and tensin homolog (PTEN), a negative regulator of the phosphoinositide 3 kinase (PI3K)-Akt pathway, leading to resistance to various therapies in both the preclinical and clinical setting. Therefore, the PI3K-Akt pathway plays a central role in various cellular processes promoting survival signaling that can contribute to the malignant phenotype, and, consequently, is an attractive pharmacologic target. However, as single agents, the efficacy of AKT inhibitors may be limited by resistance mechanisms that result in minimal cell death in tumor cells. EXPERIMENTAL DESIGN: We investigated the effects of the Akt inhibitor AZD5363 on cell proliferation, cell cycle, apoptosis, and Akt downstream pathway proteins. Survival mechanisms induced by AZD5363 were investigated. We then examined the impacts of inhibition of autophagy in combination with AZD5363 on cell proliferation and apoptosis. Furthermore, the anticancer activity of combination treatment of the lysosomotropic inhibitor of autophagy (chloroquine) with the Akt inhibitor AZD5363 was evaluated in PC-3 prostate cancer xenografts. RESULTS: Here, we show that the Akt inhibitor AZD5363 affected the Akt downstream pathway by reducing p-mTOR, p-P70S6K, and p-S6K. While AZD5363 monotherapy induced G(2) growth arrest and autophagy, it failed to induce significant apoptosis in PC-3 and DU145 prostate cancer cell lines. Blocking autophagy using pharmacologic inhibitors (3-methyladenine, chloroquine, and bafilomycin A) or genetic inhibitors (siRNA targeting Atg3 and Atg7) enhanced cell death induced by Akt inhibitor AZD5363 in these tumor prostate cell lines. Importantly, the combination of AZD5363 with chloroquine significantly reduced tumor volume by 84.9% compared with the control group and by 77.5% compared with either drug alone in PC3 xenografts. CONCLUSION: Taken together, these data show that the Akt inhibitor AZD5363 synergizes with the lysosomotropic inhibitor of autophagy chloroquine to induce apoptosis and delay tumor progression in prostate cancer models that are resistant to monotherapy AZD5363, providing a new therapeutic approach potentially translatable to patients.

Expression profile of E-cadherin and N-cadherin in non-muscle-invasive bladder cancer as a novel predictor of intravesical recurrence following transurethral resection.

The objective of this study was to investigate the impact of the expression profile of E-cadherin and N-cadherin in newly diagnosed non-muscle-invasive bladder cancer (NMIBC) on the probability of intravesical recurrence in patients undergoing transurethral resection (TUR). This study included 115 consecutive patients diagnosed as having NMIBC following TUR. Expression levels of E-cadherin and N-cadherin in TUR specimens from these patients were measured by immunohistochemical staining. In this series, intravesical recurrence occurred in 35 of 115 patients (30.4%). Immunohistochemical study showed that positive expression of E-cadherin and N-cadherin were noted in 62 (53.9%) and 48 (41.7%) specimens, respectively. Intravesical recurrence was detected in only 7 of 62 patients (11.3%) with positive E-cadherin expression, while 33 of 48 patients (68.8%) with positive N-cadherin expression developed intravesical recurrence. When patients were divided into 4 groups according to the positivities of E-cadherin and N-cadherin expression, intravesical recurrence was detected in 27 of 30 patients (90.0%) with negative E-cadherin as well as positive N-cadherin expression, and the intravesical recurrence-free survival of this group was significantly poorer than those of the remaining 3 groups. Furthermore, negative E-cadherin as well as positive N-cadherin expression was identified as the most powerful independent predictor for intravesical recurrence following TUR on multivariate analysis. These findings suggest that the loss of E-cadherin and gain of N-cadherin expression in on NMIBC appeared to be significantly associated with postoperative recurrence; therefore, the switch from E-cadherin to N-cadherin expression might be involved in the mechanism underlying intravesical recurrence of on NMIBC.

Clusterin mediates TGF-ß-induced epithelial-mesenchymal transition and metastasis via Twist1 in prostate cancer cells.

TGF-ß promotes epithelial-mesenchymal transition (EMT) and induces clusterin (CLU) expression, linking these genes to cancer metastasis. CLU is a pleiotropic molecular chaperone that confers survival and proliferative advantage to cancer cells. However, the molecular mechanisms by which TGF-ß regulates CLU expression and CLU affects metastasis remain unknown. In this study, we report that the transcription factor Twist1 mediates TGF-ß-induced CLU expression. By binding to E-boxes in the distal promoter region of CLU gene, Twist1 regulated basal and TGF-ß-induced CLU transcription. In addition, CLU reduction reduced TGF-ß induction of the mesenchymal markers, N-cadherin and fibronectin, thereby inhibiting the migratory and invasive properties induced by TGF-ß. Targeted inhibition of CLU also suppressed metastasis in an in vivo model. Taken together, our findings indicate that CLU is an important mediator of TGF-ß-induced EMT, and suggest that CLU suppression may represent a promising therapeutic option for suppressing prostate cancer metastatic progression.

Cotargeting stress-activated Hsp27 and autophagy as a combinatorial strategy to amplify endoplasmic reticular stress in prostate cancer.

Hsp27 is a stress-activated multifunctional chaperone that inhibits treatment-induced apoptosis and causes treatment resistance in prostate and other cancers. We previously showed that targeted suppression of Hsp27 sensitizes cancer cells to hormone and chemotherapy. However, mechanisms by which Hsp27 confers cell treatment resistance are incompletely defined. Here, we report that Hsp27 protects human prostate cancer cells against proteotoxic stress induced by proteasome inhibition, and that Hsp27 silencing using siRNA or antisense (OGX-427) induced both apoptosis and autophagy through mechanisms involving reduced proteasome activity and induction of endoplasmic reticulum (ER) stress. We found that autophagy activation protected against ER stress-induced cell death, whereas inhibition of autophagy activation following Hsp27 silencing using either pharmacologic inhibitors or atg3 silencing enhanced cell death. Importantly, cotargeting Hsp27 and autophagy by combining OGX-427 with the autophagy inhibitor, chloroquine, significantly delayed PC-3 prostate tumor growth in vivo. These findings identify autophagy as a cytoprotective, stress-induced adaptive pathway, activated following disruption of protein homeostasis and ER stress induced by Hsp27 silencing. Combinatorial cotargeting cytoprotective Hsp27 and autophagy illustrates potential benefits of blocking activation of adaptive pathways to improve treatment outcomes in cancer.

Growth inhibition and enhanced chemosensitivity induced by down-regulation of Aurora-A in human renal cell carcinoma Caki-2 cells using short hairpin RNA.

The objective of this study was to investigate the inhibitory effects of Aurora-A expression on the growth and chemosensitivity of Caki-2 cells in human renal cell carcinoma (RCC). Caki-2 cells were established, in which an expression vector containing short hairpin RNA (shRNA) targeting Aurora-A was introduced (Caki-2/sh-A). The growth and sensitivity of chemotherapeutic agents in Caki-2/sh-A cells were compared to those in Caki-2 cells transfected with control vector alone (Caki-2/C). The expression levels of both Aurora-A mRNA and protein in Caki-2/sh-A cells were less than 10% of those in Caki-2/C cells. The in vitro growth of Caki-2/sh-A cells was significantly inferior to that of Caki-2/C cells, and the proportion of Caki-2/sh-A cells in the G2-M phase was significantly greater compared to that of Caki-2/C cells. In addition, the expression level of Bax in Caki-2/sh-A cells was significantly higher as compared to that in Caki-2/C cells, while phosphorylated Akt in Caki-2/sh-A cells was markedly down-regulated compared to that in Caki-2/C cells. Among several chemotherapeutic agents examined, the most significant difference between Caki-2/sh-A and Caki-2/C cells was observed in the sensitivity to docetaxel. Thus, the IC(50) value of docetaxel in Caki-2/sh-A cells was decreased by approximately 90% compared to that in Caki-2/C cells. Treatment of Caki-2/sh-A cells, but not Caki-2/C ones, with 5 nM docetaxel resulted in the induction of apoptotic cell death accompanying the induction of p53. The findings suggest that the suppression of Aurora-A expression using shRNA is a useful therapeutic strategy against RCC through growth inhibition as well as enhanced chemosensitivity.

Clusterin is a critical downstream mediator of stress-induced YB-1 transactivation in prostate cancer.

Clusterin is a stress-activated, cytoprotective chaperone that confers broad-spectrum treatment resistance in cancer. However, the molecular mechanisms mediating CLU transcription following anticancer treatment stress remain incompletely defined. We report that Y-box binding protein-1 (YB-1) directly binds to CLU promoter regions to transcriptionally regulate clusterin expression. In response to endoplasmic reticulum stress inducers, including paclitaxel, YB-1 is translocated to the nucleus to transactivate clusterin. Furthermore, higher levels of activated YB-1 and clusterin are seen in taxane-resistant, compared with parental, prostate cancer cells. Knockdown of either YB-1 or clusterin sensitized prostate cancer cells to paclitaxel, whereas their overexpression increased resistance to taxane. Clusterin overexpression rescued cells from increased paclitaxel-induced apoptosis following YB-1 knockdown; in contrast, however, YB-1 overexpression did not rescue cells from increased paclitaxel-induced apoptosis following clusterin knockdown. Collectively, these data indicate that YB-1 transactivation of clusterin in response to stress is a critical mediator of paclitaxel resistance in prostate cancer.