Dah-Shyong Yu

Estrogen inhibits renal cell carcinoma cell progression through estrogen receptor-β activation.

Renal cell carcinoma (RCC) originates in the lining of the proximal convoluted tubule and accounts for approximately 3% of adult malignancies. The RCC incidence rate increases annually and is twofold higher in males than in females. Female hormones such as estrogen may play important roles during RCC carcinogenesis and result in significantly different incidence rates between males and females. In this study, we found that estrogen receptor β (ERβ) was more highly expressed in RCC cell lines (A498, RCC-1, 786-O, ACHN, and Caki-1) than in breast cancer cell lines (MCF-7 and HBL-100); however, no androgen receptor (AR) or estrogen receptor α (ERα) could be detected by western blot. In addition, proliferation of RCC cell lines was significantly decreased after estrogen (17-β-estradiol, E2) treatment. Since ERβ had been documented to be a potential tumor suppressor gene, we hypothesized that estrogen activates ERβ tumor suppressive function, which leads to different RCC incidence rates between males and females. We found that estrogen treatment inhibited cell proliferation, migration, invasion, and increased apoptosis of 786-O (high endogenous ERβ), and ERβ siRNA-induced silencing attenuated the estrogen-induced effects. Otherwise, ectopic ERβ expression in A498 (low endogenous ERβ) increased estrogen sensitivity and thus inhibited cell proliferation, migration, invasion, and increased apoptosis. Analysis of the molecular mechanisms revealed that estrogen-activated ERβ not only remarkably reduced growth hormone downstream signaling activation of the AKT, ERK, and JAK signaling pathways but also increased apoptotic cascade activation. In conclusion, this study found that estrogen-activated ERβ acts as a tumor suppressor. It may explain the different RCC incidence rates between males and females. Furthermore, it implies that ERβ may be a useful prognostic marker for RCC progression and a novel developmental direction for RCC treatment improvement.

Taxol synergizes with antioxidants in inhibiting hormal refractory prostate cancer cell growth

Taxanes are chemotherapeutic agents commonly used to treat various carcinomas. Dietary antioxidants, such as vitamin E, green tea extracts, and isoflavones have been used against prostate cancer, and exhibit anticancer effects both in vitro and in vivo. We evaluated the combined effect of taxol (paclitaxel) with pyrrolidine dithiocarbamate, vitamin E, epigallocatechin gallate, and genistein in killing hormone-refractory prostate cancer cells. Those agents were tested on the hormone-refractory prostate cancer cell line PC-3, and the viability of the cells was determined using MTT {3 (4, 5-dimethylthiazo-2-yl)-2, 5-diphenyl tetrazolium} assay after drug treatment. PC-3 cells were sensitive to these drugs with 50% inhibitory concentrations of 0.1, 23, 220, 1122, and 260 microM, for taxol, pyrrolidine dithiocarbamate, epigallocatechin gallate, genistein, and vitamin E, respectively. Genistein, pyrrolidine dithiocarbamate, and epigallocatechin gallate showed synergistic cytotoxicity to PC-3 cells when combined with 0.01 microM taxol. Only high concentration of vitamin E showed a synergistic effect with this dose of taxol. Further study revealed that 3 combinations could induce sub-G1 phase of cell cycle, induce apoptosis, and increase caspase activity and decrease Bcl-2 expression simultaneously. In conclusion, in addition to vitamin E, incorporation of these antioxidants with taxan-based cytotoxic therapies offers encouraging strategies for combating hormone-refractory prostate cancers.

Sunitinib can enhance BCG mediated cytotoxicity to transitional cell carcinoma through apoptosis pathway.

Bacillus Calmette-Guerin (BCG)-refractory generated a high risk to patients with bladder cancer during treatment. Tyrosine kinase receptor (TKR) and TKR-mediated signal transduction pathways play an important role in tumor initiation, maintenance, angiogenesis, and vascular proliferation. Theoretically, it is helpful in adjuvant treatment for transitional cell carcinoma (TCC). Hence, we proposed that sunitinib, a endothelial growth factor receptor (VEGFR) inhibitor, may have a synergistic effect with BCG in enhancing its cytotoxicity to bladder cancer. The level of VEGF in various TCC cell lines was quantified by real time PCR. High grade TCC-T24 cell line with high level of VEGF expression was selected as representative tumor cells for further study. The single drug and combined inhibitory effects of BCG and sunitinib in T24 cells were determined by MTT method. The drug mediated cell apoptosis in T24 cells was characterized by flow cytometry with PI and annexin V stain. Bcl-2 apoptotic pathway induction by BCG and sunitinib treatment was evaluated by Western blotting method. Inhibitory ability of sunitinib in BCG induced cell migration was verified by cell migration assay. The results shown that expression level of VEGF mRNA in high grade T24 cells was higher than low grade J82, TSGH 8301, and TCC 9202 cell lines. Both BCG and sunitinib treatment presented cytotoxic effect to T24 cells in a dose-dependent manner. Combination of BCG and sunitinib revealed superior cytotoxicity effect than single agent when cells were pretreated with low dosage BCG before sunitinib treatment. By Annexin V analysis it was observed that cell death associated with increased early and late apoptosis process individually. Furthermore, the bcl-2 expression was significant reduced in T24 cells in metachronous BCG and sunitinib combination treatment than single agent. Tumor cell migration activity was also markedly inhibited with BCG and sunitinib combination treatment. In conclusion, these results suggested that during BCG and sunitinib combination treatment both reagents interacted with each other and caused TCC cells apoptosis in addition to direct cytotoxicity. This combination therapeutic model may have the potential for future clinical application to bladder cancer treatment.

Tyrosine kinase receptor inhibitor-targeted combined chemotherapy for metastatic bladder cancer

Overexpression of hypoxia‐inducible factor‐1 alpha is noted during the invasive and metastatic process of transitional cell carcinoma. It will upregulate vascular endothelial growth factor (VEGF) and drive proliferation, invasiveness, metastasis, and antiapoptotic ability of cancer cells. We proposed that tyrosine kinase receptor inhibitor, sunitinib malate—(Sutent; Pfizer Inc., Taiwan), combined with chemotherapeutic drug may present synergistic cytotoxic enhancement to transitional cell carcinoma cells with subsequent inhibition of their cellular behaviors, including proliferation, invasiveness, and metastatic activity. The contents of VEGF‐A in mouse bladder tumor cells (MBT‐2) and culture medium were detected by quantification‐polymerase chain reaction and Western blot individually. The inhibitory concentrations of various chemotherapeutic drugs, sunitinib, and their combination treatment in MBT‐2 were determined by 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2H‐tetrazolium bromide (MTT) assay. Microchamber transmembrane migration assay was applied in evaluation of the inhibitory effects of different dosages of sunitinib and combination treatment on tumor cells. The cell cycle and apoptosis were analyzed after combination therapy by flow cytometry. Variation in apoptotic pathway was elucidated by Western blot using specific antibodies with cleaved PARP and caspase‐3. Metastatic animal model mimicked by tail vein injection of MBT‐2 cells was used to evaluate the treatment efficiency in tumor weight and survival rate. The mRNA and protein level of VEGF‐A in MBT‐2 cells increased by 70% at 48 hours interval under hypoxia stress condition. In MTT assay, MBT‐2 cells had shown the highest sensitivity to epirubicin. Sunitinib combined with epirubicin had shown a synergistic cytotoxic effect to MBT‐2 cells. Sunitinib and its combination with epirubicin showed significant inhibition on MBT‐2 cells migration in microchambers. G2/M phase arrest and increased subG1 in cell cycle was seen in the epirubicin and sunitinib combination treatment group. The activation of apoptosis pathway was confirmed by increased cleaved caspase‐3 and cleaved PARP in MBT‐2 cells. In tail vein tumor inoculation C3H mice model, epirubicin alone and sunitinib combination therapy decreased tumor growth in lungs with marginal effect. Sunitinib and epirubicin combination had shown a synergistic cytotoxic effect and inhibited cell migration ability in MBT‐2 cells. The combination can induce cell cycle arrest at G2/M phase and increase subG1 cells. Metastatic animal study also showed that sunitinib combined with epirubicin has a marginal effect on inhibition of tumor growth of lungs. The tyrosine kinase receptor inhibitor‐targeted combined chemotherapy regimen may provide as a new treatment modality for advanced bladder cancer in the future.

NGAL Can Alternately Mediate Sunitinib Resistance in Renal Cell Carcinoma

PURPOSEnSerum NGAL is highly expressed in patients with advanced renal cancer treated with sunitinib. We investigated the role of NGAL in sunitinib resistance in renal cell carcinoma to identify potential tactics to overcome it.nnnMATERIALS AND METHODSnNGAL expression was correlated with sunitinib sensitivity. Vascular endothelial growth factor related upstream Ras, Erk1/2 and STAT1 phosphorylation activity in Caki-1 and NGAL transfected Caki-1 cells after sunitinib treatment was analyzed using Western blot. NGAL and vascular endothelial growth factor-A interaction with sunitinib therapeutic efficacy was monitored in renal cell carcinoma tumor xenografted mice by tumor growth inhibition, serum NGAL and vascular endothelial growth factor-a levels, and microscopic examination of tumor microvascular density.nnnRESULTSnSunitinib cytotoxicity in various renal cell carcinoma cell lines was reversibly related to NGAL expression. Sunitinib showed the lowest 50% inhibitory concentration (5.53 μM) in Caki-1 cells, which had the lowest NGAL expression of these renal cell carcinoma cell lines. After sunitinib treatment adding NGAL inhibited Ras and Erk1/2 phosphorylation but activated STAT1α phosphorylation in Caki-1 cells and Caki-1 cells transfected with NGAL. In a xenograft mouse model sunitinib significantly inhibited tumor growth in Caki-1 mice. NGAL transfected Caki-1 mice had higher serum NGAL and lower vascular endothelial growth factor-A than Caki-1 mice. Microvascular density was decreased in Caki-1 mice with sunitinib treatment.nnnCONCLUSIONSnNGAL in tumor cells may show crosstalk with vascular endothelial growth factor-a and alternative activation in stimulating tumor growth during sunitinib treatment. It may become a therapeutic target to reverse sunitinib resistance in renal cell carcinoma.

Bacille Calmette-Guerin can induce cellular apoptosis of urothelial cancer directly through toll-like receptor 7 activation.

Immunotherapy using bacille Calmette‐Guerin (BCG) instillation is the mainstay treatment modality for superficial urothelial cancer (UC) through toll‐like receptor (TLR) activation of cognitive immune response. We investigated the roles of TLR7 in the activation of apoptosis in UC cells after BCG treatment. The inu2009vitro cytotoxicity effect of BCG on UC cells was measured by a modified 3‐(4,5‐dimethylthiazo‐2‐yl)‐2,5‐diphenyl tetrazolium assay. Expressions of TLR7 mRNA and protein in native UC cells prior to and after BCG treatment were analyzed using real‐time quantitative polymerase chain reaction and western blot methods. Phagocytotic processes after BCG treatment in UC cells were observed microscopically using a specific immunostain, subsequent cellular apoptosis‐related signals induced by TLR7 were analyzed by western blot. Low‐grade UC cells, TSGH8301, showed significant cellular death (4.23‐fold higher than the high‐grade UC cells T24 and J82) when treated with BCG and the BCG cytotoxicity was displayed in a dose–time‐dependent manner. TSGH8301 cells had the highest content of TLR7 mRNA, 7.2‐ and 4.5‐fold higher than that of T24 and J82 cells, respectively. TLR7 protein expression was also significantly increased in TSGH8301 cells. Phagocytosis‐related markers, including beclin 1, ATG2, and LC3, were increased when TSGH8301 cells were treated by BCG. Interleukin‐1 receptor‐associated kinases 2 and 4 were also increased markedly in TSGH8301 cells. On the contrary, cellular apoptosis of TSGH8301 cells decreased by 34% when TLR7 activation was suppressed by the TLR antagonist IRS661 after BCG treatment. Our findings suggest that well differentiated TCC cells have higher expression of TLR7 and BCG can drive cellular death of TCC cells directly via TLR7 activation and related apoptotic pathway.

Combined Y-shaped common channel transureteroureterostomy with Boari flap to treat bilateral long-segment ureteral strictures

BackgroundUreteral stricture is a complication of several etiologies including idiopathic retroperitoneal fibrosis, infection, radiotherapy, instrumentation, and surgical procedures. A variety of techniques have been reported for management. The transureteroureterostomy and bladder flap have been the standard procedures for repairing distal ureteral defects of unilateral ureter. Bilateral ureteral stricture is an uncommon condition that challenges usual reconstructive procedures. It is a difficult task to reconstruct the complex situation of bilateral ureteral strictures.Case presentationA 54-year-old female underwent concurrent chemoradiotherapy for stage IVB squamous cell carcinoma of cervix. Subsequently, she had stricture of bilateral distal ureters with bilateral hydroureteronephrosis which was found by computed tomography. The renal function deteriorated during the follow-up period. She had periodic change of double-J stents and percutaneous nephrostomy. However, the renal function still deteriorated. We performed a combined Y-shaped common channel transureteroureterostomy with Boari flap to reconstruct bilateral long-segment ureteral strictures. The patient recovered uneventfully.ConclusionReconstruction of bilateral ureteral strictures is a difficult treatment. We developed a modified technique for the complex situation of bilateral ureteral strictures. To our knowledge, this has not been previously reported in the scientific literature and it is a feasible procedure to treat bilateral long-segment ureteral strictures.

Combining prostrate-specific antigen and Gleason score increases the diagnostic power of endorectal coil magnetic resonance imaging in prostate cancer pathological stage.

Background: The proper use of endorectal coil MRI (eMRI) images provide detailed information for the real extent of locally prostate cancer invasion and involvement of pelvic lymph nodes. This study evaluated the accuracy of endorectal coil magnetic resonance imaging (eMRI) results, combining the preoperative prostate‐specific antigen (PSA), and the biopsy Gleason score to improve the diagnostic accuracy of prostate cancer (PCa) with organ‐confined disease (OCD) or extracapsular extension (ECE)/seminal vesicle invasion (SVI). Methods: Between 2001 and 2007, 94 PCa patients received eMRI testing during presurgical evaluation and underwent radical prostatectomy. As a part of routine patient workup, serum PSA level and Gleason score after pathology examination were recorded. The eMRI images were used to help assess patient PCa staging status regarding OCD or ECE/SVI. These stage assessments as evaluated through the use of MRI were compared with the final specimen pathological stage after the patients underwent radical prostatectomy. Results: Of the total 94 patients in our study, 65 had stage pT2, 12 had stage pT3a, and 17 had stage pT3b PCa. In patients with clinical stage T2 PCa, the Gleason score significantly improved the discriminative ability of eMRI to successfully predict PCa at the OCD stage. Otherwise, in cases of clinical stage T3 PCa, accurate determination of PSA levels significantly improved eMRI predictive ability to assess ECE or SVI staging. Conclusion: In clinical stage T2 PCa patients, integrating the biopsy Gleason score improved the discriminative ability to assess OCD PCa staging. Additionally, combining the preoperative PSA levels of clinical T3 prostate cancer cases with Gleason scores significantly improved the sensitivity and accuracy of eMRI diagnosis to distinguish ECE from SVI.

Fistula Involving Urinary Bladder: Experience with the Management of 23 Cases

PURPOSE: Fistula involving urinary bladder is a common disease caused by pelvic malignancies, radiotherapy, previous operations, primary intestinal diseases and trauma. We have reviewed our experience in diagnosis and management of 23 cases of fistulae involving urinary bladder. nMATERIALS AND METHODS: From 1995 to 2000, 23 patients with fistulae involving urinary bladder were diagnosed and treated at eh Tri-Service General Hospital. Each patient’s record were reviewed retrospectively regarding symptoms, causes, primary disease processes, diagnostic studies, managements, complications, mortality and follow-up. nRESULTS: The most common fistula involving urinary bladder was vesicointestinal fistula (57%), followed by vesicovaginal (26%) and vesicocutaneous (17%) fistulae. Fecaluria (46%) was the most common symptom of vesicointestinal fistula, while the major cause of vesicointestinal fistula was malignancy (54%). Urinary leakage from the vagina (100%) was a major symptom found in the patients of vesicovaginal fistula, which was caused by malignancy (33%), previous operation (33%) and radiotherapy (33%). Previous operation (75%) was found to be a major cause for vesicocutaneous fistula. Most of the fistulae involving urinary bladder may be diagnosed by cystoscopy (83%). The treatment and outcome were individualized, dependent on patient’s general condition and progression of the disease. nCONCLUSIONS: The diagnosis of fistula involving urinary bladder can be established by combining the results of cystoscopy, colposcopy lower G-I series and cystography. Two-stage operation is preferable to single-stage operation in the case of vesicointestinal fistula, while a single-stage operation is recommended for the vesicovaginal and vesicocutaneous fistulae.

Granulomatous Prostatitis after Intravesical BCG Therapy for Bladder Cancer-Report of a Case and Review of the Literature

Intravesical bacillus Calmette-Guerin (BCG) instillation is a well estalished ther-apy for superficial bladder cancer. Urinary tract complications induced by the BCG have been reported by various investigators. The granulomatous prostatits is an un-common complication with reported incidence of 0.9%. We report a patient, underwent 100mg Taipei-NIPM BCG intravesical instillation wddkly for consecutive 6 doses, devel-oped granulomatous prostatitis one year later. This case initially presented symptoms of prostatism and granulomatous prostatitis was diagnosed incidentally after the TURP. Bacillus Calmette-Guerin related granulomas of the prostate may be differentiated histo-logically from nonspecific granulomatous prosatitis, mycotic prostatitis and postoperative prostatitis. No specific treatment is required since this kind of entity usually is self-limited.