MUC15 loss facilitates epithelial-mesenchymal transition and cancer stemness for prostate cancer metastasis through GSK3ß/ß-catenin signaling.

Patients with prostate cancer (PCa) have a high incidence of relapse and metastasis. Unfortunately, the molecular mechanisms underlying these processes have not been fully elucidated. In our study, we demonstrate that MUC15, a member of the mucin family, is a novel tumor suppressor in PCa that modulates epithelial-mesenchymal transition (EMT) and cancer stemness, contributing to PCa metastasis. First, MUC15 expression was found to be decreased in PCa tissues compared with para-carcinoma tissues. Moreover, we observed that MUC15 suppressed cell migration and invasion, both in vitro and in vivo, but had no effect on cell proliferation. Mechanistically, knockdown of MUC15 increased GSK3ß phosphorylation and promoted ß-catenin nuclear translocation. Therefore, the ß-catenin-specific inhibitors XAV939 and PRI-724 rescued EMT in MUC15-deficient cell lines. Taken together, these results indicate that MUC15 is downregulated in PCa tissues and serves as a potential target to prevent PCa metastasis, which can inhibit EMT and cancer stemness via the GSK3ß/ß-catenin signaling pathway.

Correction: Infiltrating mast cells increase prostate cancer chemotherapy and radiotherapy resistances via modulation of p38/p53/p21 and ATM signals.

[This corrects the article DOI: 10.18632/oncotarget.6372.].

The expression and function of RASAL2 in renal cell carcinoma angiogenesis.

Patients with renal cell carcinoma (RCC) often develop resistance to antivascular drugs and eventually succumb to disease. However, the underlying molecular mechanism remains poorly understood. In this study, we demonstrated that RASAL2, a RAS GTPase-activating protein, played a tumor-suppressive role in RCC by targeting tumor angiogenesis. Firstly, we showed that RASAL2 was frequently epigenetically silenced in RCC, and its loss was negatively correlated with overall survival of RCC patients. Furthermore, we discovered that RASAL2 could inhibit RCC angiogenesis in vitro and in vivo. Mechanistically, we identified that RASAL2 could activate GSK3ß by reducing Ser9 phosphorylation and subsequently decrease the expression of c-FOS and vascular endothelial growth factor A (VEGFA). Interruption of the p-GSK3ß/c-FOS pathway with the specific inhibitor or small interfering RNA could reverse the expression of VEGFA, which may provide a new insight to prevent RCC from resistance to antivascular therapy.

RASAL2 inhibits tumor angiogenesis via p-AKT/ETS1 signaling in bladder cancer.

Muscle-invasive or metastatic bladder cancer (BCa) is a life-threatening disease for patients, and tumor angiogenesis is believed to play a critical role in the progression of BCa. However, its underlying mechanism of tumor angiogenesis is still poorly understood. In this study, we discovered that RASAL2, a RAS GTPase activating protein, could inhibit BCa angiogenesis based on our shRNA/siRNA knockdown or ectopic cDNA expression experiments. Mechanistically, RASAL2 downregulation could enhance the phosphorylation of AKT and then subsequently upregulate the expression of ETS1 and VEGFA. Furthermore, there was a negative correlation between RASAL2 and VEGFA or CD31 expression in subcutaneous xenograft and human BCa specimens. Taken together, we provide a new insight into the molecular mechanism of BCa progression, in which RASAL2 can be a new therapeutic target.

Clinical-radiologic correlation of mixed epithelial and stromal tumor of the kidneys: Cases analysis.

BACKGROUND: Mixed epithelial and stromal tumor of the kidney (MESTK) is a rare tumor, with few malignant cases reported. Occurring mostly in middle-aged women, it is characterized by a biphasic pathological structure. METHODS: This study retrospectively reviewed the imaging findings and medical records of six MESTK cases of a single institution in a 10-year period. RESULTS: All of the patients were middle-aged women without hormone therapy history. The typical image was a renal tumor with varied cystic components. Half of the cases had sinus invagination, but only one had intratumor calcification. On imaging studies, four were Bosniak Category IV, one was Category III, and one presented as a solid tumor. The mean RENAL nephrometry score was 9.3. Five patients underwent partial nephrectomy, with no statistical renal functional deterioration after nephron-sparing surgery. There were no peri-operative complications. CONCLUSION: Surgery remains the treatment of choice for MESTK, and nephron-sparing surgery should be considered in feasible cases.

Infiltrating mast cells increase prostate cancer chemotherapy and radiotherapy resistances via modulation of p38/p53/p21 and ATM signals.

Early studies indicated that mast cells in prostate tumor microenvironment might influence prostate cancer (PCa) progression. Their impacts to PCa therapy, however, remained unclear. Here we found PCa could recruit more mast cells than normal prostate epithelial cells then alter PCa chemotherapy and radiotherapy sensitivity, leading to PCa more resistant to these therapies. Mechanism dissection revealed that infiltrated mast cells could increase p21 expression via modulation of p38/p53 signals, and interrupting p38-p53 signals via siRNAs of p53 or p21 could reverse mast cell-induced docetaxel chemotherapy resistance of PCa. Furthermore, recruited mast cells could also increase the phosphorylation of ATM at ser-1981 site, and inhibition of ATM activity could reverse mast cell-induced radiotherapy resistance. The in vivo mouse model with xenografted PCa C4-2 cells co-cultured with mast cells also confirmed that mast cells could increase PCa chemotherapy resistance via activating p38/p53/p21 signaling. Together, our results provide a new mechanism showing infiltrated mast cells could alter PCa chemotherapy and radiotherapy sensitivity via modulating the p38/p53/p21 signaling and phosphorylation of ATM. Targeting this newly identified signaling may help us better suppress PCa chemotherapy and radiotherapy resistance.

FGF2-mediated reciprocal tumor cell-endothelial cell interplay contributes to the growth of chemoresistant cells: a potential mechanism for superficial bladder cancer recurrence.

Patients with superficial bladder cancer can be definitively cured by one single transurethral resection (TUR) with additional intravesical chemotherapy; however, up to 75 % of cases display frequent and multiple recurrences. One of the major causes of recurrence is that chemotherapeutic drugs used in intravesical regimens may induce chemoresistance. However, the mechanisms by which these chemoresistant cells develop into recurrent tumors remain unclear. Recent clinical evidence revealed that the expression of pro-angiogenic factor FGF2 was associated with early local relapse in patients with superficial bladder cancer. In this study, we conducted a preliminary investigation of the mechanisms of chemoresistant cells mediated bladder cancer recurrence, focusing on FGF2-initiated tumor cell-endothelial cell interaction on chemoresistant cancer cell growth. We found that the expression of FGF2 was increased in chemoresistant bladder cell lines and in bladder tissues after intravesical chemotherapy. Although chemoresistant bladder cells grow slower than parental cells, chemoresistant bladder cancer cells had stronger ability than parental cells to stimulate endothelial cell migration, growth, and tube formation by producing FGF2. Inversely, endothelial cells significantly promoted chemoresistant bladder cancer growth in vitro and in vivo. Thus, targeting chemotherapy-induced FGF2 upregulation may provide a promising approach to manage the recurrence of superficial bladder cancer.

Silibinin induces apoptosis through inhibition of the mTOR-GLI1-BCL2 pathway in renal cell carcinoma.

The downstream transcriptional factor of the hedgehog (Hh) pathway, GLI family zinc finger 1 (GLI1), plays a crucial role in regulating tumor progression. In the present study, we demonstrated that silibinin, a natural flavonoid antioxidant isolated from extracts of the milk thistle herb, exerts its anticancer capabilities by restraining GLI1 function in renal cell carcinoma (RCC) cells in vitro and in vivo. In the present study, we confirmed that silibinin induced growth inhibition of RCC through caspase-dependent apoptosis and downregulation of GLI1 and BCL2, which could be partially reversed by GLI1 overexpression. Moreover, we determined that the decreased GLI1 expression by silibinin was mediated by the mammalian target of rapamycin (mTOR) pathway. The in vivo mouse xenograft study also showed that silibinin significantly reduced RCC tumor growth and specifically targeted the mTOR-GLI1-BCL2 signaling pathway. In conclusion, our findings demonstrated for the first time that silibinin induces apoptosis of RCC cells through inhibition of the mTOR-GLI1­BCL2 pathway. These findings also indicate that GLI1 is a novel regulator for the potential therapeutic application of silibinin against RCC.

Anti-androgen enzalutamide enhances prostate cancer neuroendocrine (NE) differentiation via altering the infiltrated mast cells → androgen receptor (AR) → miRNA32 signals.

The recently developed anti-androgen enzalutamide also known as (MDV3100) has the advantage to prolong by 4.8 months the survival of castration resistant prostate cancer (CRPC) patients. However, the mechanisms behind the potential side effects involving the induction of the prostate cancer (PCa) neuroendocrine (NE) differentiation remain unclear. Here we found PCa cells could recruit more mast cells than normal prostate epithelial cells, and enzalutamide (or casodex) treatment could further increase such recruitment that resulted in promoting the PCa NE differentiation. Mechanism dissection found infiltrated mast cells could function through positive feedback to enhance PCa to recruit more mast cells via modulation of the androgen receptor (AR) â†’ cytokines IL8 signals, and interruption by AR-siRNA or neutralizing anti-IL8 antibody could partially reverse the recruitment of mast cells. Importantly, targeting the PCa androgens/AR signals with AR-siRNA or enzalutamide (or casodex) also increased PCa NE differentiation via modulation of the miRNA32 expression, and adding miRNA32 inhibitor reversed the AR-siRNA- or enzalutamide-enhanced NE differentiation. Together, these results not only identified a new signal via infiltrated mast cells â†’ PCa AR â†’ miRNA32 to increase PCa NE differentiation, it also pointed out the potential unwanted side effects of enzalutamide (or casodex) to increase PCa NE differentiation. Targeting these newly identified signals, including AR, IL8, or miRNA32, may help us to better suppress PCa NE differentiation that is induced during ADT with anti-androgen enzalutamide (or casodex) treatment.

Kaempferol suppresses bladder cancer tumor growth by inhibiting cell proliferation and inducing apoptosis.

The effects of the flavonoid compound, kaempferol, which is an inhibitor of cancer cell proliferation and an inducer of cell apoptosis have been shown in various cancers, including lung, pancreatic, and ovarian, but its effect has never been studied in bladder cancer. Here, we investigated the effects of kaempferol on bladder cancer using multiple in vitro cell lines and in vivo mice studies. The MTT assay results on various bladder cancer cell lines showed that kaempferol enhanced bladder cancer cell cytotoxicity. In contrast, when analyzed by the flow cytometric analysis, DNA ladder experiment, and TUNEL assay, kaempferol significantly was shown to induce apoptosis and cell cycle arrest. These in vitro results were confirmed in in vivo mice studies using subcutaneous xenografted mouse models. Consistent with the in vitro results, we found that treating mice with kaempferol significant suppression in tumor growth compared to the control group mice. Tumor tissue staining results showed decreased expressions of the growth related markers, yet increased expressions in apoptosis markers in the kaempferol treated group mice tissues compared to the control group mice. In addition, our in vitro and in vivo data showed kaempferol can also inhibit bladder cancer invasion and metastasis. Further mechanism dissection studies showed that significant down-regulation of the c-Met/p38 signaling pathway is responsible for the kaempferol mediated cell proliferation inhibition. All these findings suggest kaempferol might be an effective and novel chemotherapeutic drug to apply for the future therapeutic agent to combat bladder cancer.

Curcumin promotes KLF5 proteasome degradation through downregulating YAP/TAZ in bladder cancer cells.

KLF5 (Krüppel-like factor 5) plays critical roles in normal and cancer cell proliferation through modulating cell cycle progression. In this study, we demonstrated that curcumin targeted KLF5 by promoting its proteasome degradation, but not by inhibiting its transcription in bladder cancer cells. We also demonstrated that lentivirus-based knockdown of KLF5 inhibited cancer cell growth, while over-expression of a Flag-tagged KLF5 could partially reverse the effects of curcumin on cell growth and cyclin D1 expression. Furthermore, we found that curcumin could down-regulate the expression of Hippo pathway effectors, YAP and TAZ, which have been reported to protect KLF5 protein from degradation. Indeed, knockdown of YAP by small interfering RNA caused the attenuation of KLF5 protein, but not KLF5 mRNA, which was reversed by co-incubation with proteasome inhibitor. A xenograft assay in nude mice finally proved the potent inhibitory effects of curcumin on tumor growth and the pro-proliferative YAP/TAZ/KLF5/cyclin D1 axis. Thus, our data indicates that curcumin promotes KLF5 proteasome-dependent degradation through targeting YAP/TAZ in bladder cancer cells and also suggests the therapeutic potential of curcumin in the treatment of bladder cancer.

Androgen receptor enhances kidney stone-CaOx crystal formation via modulation of oxalate biosynthesis & oxidative stress.

Males develop kidney stones far more frequently than females with a ratio of 2-3:1, suggesting that androgen receptor (AR) signaling might play a key role in the development of nephrolithiasis. Using the cre-loxP system to selectively knock out AR in glyoxylate-induced calcium oxalate (CaOx) crystal mouse models, we found that the mice lacking hepatic AR had less oxalate biosynthesis, which might lead to lower CaOx crystal formation, and that the mice lacking kidney proximal or distal epithelial AR also had lower CaOx crystal formation. We found that AR could directly up-regulate hepatic glycolate oxidase and kidney epithelial NADPH oxidase subunit p22-PHOX at the transcriptional level. This up-regulation might then increase oxalate biosynthesis and oxidative stress that resulted in induction of kidney tubular injury. Targeting AR with the AR degradation enhancer ASC-J9 led to suppression of CaOx crystal formation via modulation of oxalate biosynthesis and oxidative stress in both in vitro and in vivo studies. Taken together, these results established the roles of AR in CaOx crystal formation.

Novel green-light KTP laser en bloc enucleation for nonmuscle-invasive bladder cancer: technique and initial clinical experience.

BACKGROUND AND PURPOSE: The standard procedure for staging and treating nonmuscle-invasive bladder cancer (NMIBC) is still transurethral resection of bladder tumor (TURBT) via a wire loop. However, TURBT is associated with serious disadvantages that facilitate tumor recurrence. Recently, lasers have been explored as treatment tools for bladder tumors. Here, we report a novel tumor en bloc enucleation using a front-firing green-light potassium-titanyl-phosphate laser and its initial clinical application. PATIENTS AND METHODS: From March through June 2013, 45 patients with NMIBC received modified transurethral resection using a front-firing green-light laser. En bloc enucleation was performed on all tumors. Preoperative and intraoperative data were retrospectively collected. RESULTS: All patients successfully went through a session of treatment with front-firing green-light laser enucleation of the bladder tumor. Complications such as bladder hemorrhage, vesicle perforation, and obturator nerve reflex were not encountered during the treatment. The tumor diameter ranges from 0.3 to 3.0 cm with a mean value of 1.8 cm. Mean operative time and enucleation time were 21 (12-38) and 12 (4-23) minutes, respectively. Serum hemoglobin decreased 1.1 (0.1-2.4) mg/dL averagely. Mean catheter time was 2.0 (1.0-3.0) days, and mean postoperative hospital stay was 2.5 (1.5-4.0) days. The stages of bladder cancer included 27 Ta, 15 T1, and 3 T2a. No tumor recurrence was observed at the initial 6-month follow-up. CONCLUSIONS: The modified technique using a front-firing green-light laser to en bloc enucleate bladder tumors is effective and safe for treatment of NMIBC. Moreover, it may improve the accurate valuation of tumor stage and prediction of postoperative prognosis, although long-term outcomes and prospective clinical trials are needed.

Sinomenine reverses multidrug resistance in bladder cancer cells via P-glycoprotein-dependent and independent manners.

P-Glycoprotein-mediated multidrug resistance is a frequent event during chemotherapy and a key obstacle for bladder cancer therapy. Search for strategies to reverse multidrug resistance is a promising approach to improve the management of bladder cancer. In the present study, we reported a novel P-glycoprotein-mediated multidrug resistant cell model 253J/DOX, which was generated from human bladder cancer 253J cell line. Furthermore, we found that the multidrug resistant phenotype of 253J/DOX cells could be overcome by sinomenine, an alkaloid derived from the stem of Sinomenium acutum. Mechanistically, the chemosensitive effect by sinomenine was mediated by down-regulating P-glycoprotein expression, as well as triggering apoptotic pathways. The chemosensitive effect of sinomenine may make it a prime candidate agent to target bladder cancer.

[Expression and significance of FGFR2IIIc in chemoresistant bladder cancer cells].

OBJECTIVE: To investigate the role of fibroblast growth factor receptor-2 (FGFR2) splice variant FGFR2IIIc in the regulation of mesenchymal-epithelial transition (MET) in doxorubicin-resistant bladder cancer cells. METHODS: A doxorubicin-resistant human bladder cell line (253J/DOX) was generated from the bladder cancer cell line 253J by being continuously exposed to gradually increasing doses of doxorubicin. Chemosensitivity to doxorubicin was determined by MTT assay. The expressions of P-glycoprotein and FGFR2IIIc were evaluated by Western blotting and real-time RT-PCR, respectively. Changes in E-cadherin and vimentin were detected by Western blot analysis. Migration ability of 253J and 253J/DOX cells was analyzed by in vitro wound healing assay. RESULTS: The resistant cells, 253J/DOX, were more resistant to doxorubicin than the parent cells. Western blotting and RT-PCR analysis indicated the higher levels of P-glycoprotein and FGFR2IIIc in 253J/DOX cells (P<0.05). Additionally, compared with the 253J cells, 253J/DOX cells presented the upregulation of E-cadherin, the downregulation of vimentin and the inhibition of migration ability. CONCLUSION: FGFR2IIIc-induced MET in chemoresistant bladder cancer cells may play an important role in the formation of metastatic lesions.

Increased PrLZ-mediated androgen receptor transactivation promotes prostate cancer growth at castration-resistant stage.

Most advanced prostate cancers (PCa) will develop into the castration-resistant stage following androgen deprivation therapy, yet the molecular mechanisms remain unclear. In this study, we found PrLZ, a newly identified Prostate Leucine Zipper gene that is highly expressed in PCa could interact with the androgen receptor (AR) directly leading to enhance AR transactivation in the castration-resistant condition. PrLZ might enhance AR transactivation via a change of AR conformation that leads to promotion of AR nuclear translocation and suppression of AR degradation via modulating the proteasome pathway, which resulted in increased prostate-specific antigen expression and promoted PCa growth at the castration-resistant stage. Clinical PCa sample survey from same-patient paired specimens found increased PrLZ expression in castration-resistant PCa following the classical androgen deprivation therapy. Targeting the AR-PrLZ complex via ASC-J9® or PrLZ-siRNA resulted in suppression of PCa growth in various human PCa cells and in vivo mouse PCa models. Together, these data not only strengthen PrLZ roles in the transition from androgen dependence to androgen independence during the castration-resistant stage, but they may also provide a new potential therapy to battle PCa at the castration-resistant stage.

Twist confers chemoresistance to anthracyclines in bladder cancer through upregulating P-glycoprotein.

BACKGROUND: The membrane transporter P-glycoprotein (P-gp) was found to mediate chemoresistance, which is one of the obstacles to effective chemotherapy in several types of human cancer. The transcription factor Twist, which has been reported to participate in cancer invasion and metastasis, also plays a vital role in the progression of chemoresistance. However, the effect of Twist on P-gp-related chemoresistance remains dubious. METHODS AND RESULTS: We found that Twist can regulate the expression of P-gp and then confer resistance to anthracycline drugs in human bladder cancer cells. Firstly, Twist was found to be coexpressed with P-gp in human bladder cancer cells and tissues, which were associated with enhanced chemoresistance to anthracycline drugs. Secondly, knockdown of Twist by specific siRNA treatment significantly sensitized bladder cancer cells to anthracycline drugs via inhibiting P-gp expression. Bladder cancer cells that survived transient exposure to anthracycline drugs showed higher levels of P-gp expression and more nuclear localization of Twist than untreated cells. CONCLUSION: We report a novel mechanism of anthracycline chemoresistance in bladder cancer in which activated Twist mediates P-gp expression in addition to its antiapoptotic roles. Therapeutic strategies targeting Twist may improve the management of recurrent bladder cancer after chemotherapy.

Inhibitory effect of silibinin on EGFR signal-induced renal cell carcinoma progression via suppression of the EGFR/MMP-9 signaling pathway.

The activation of epidermal growth factor (EGF) through its receptor, EGFR, is one of the major mechanisms that mediate renal cell carcinoma (RCC) metastasis. Silibinin, a natural flavonoid antioxidant with pleiotropic anticancer capability, has shown anti-metastatic effects in a variety of cancers. However, the mechanism by which silibinin inhibits EGFR signal-induced migration and invasion of RCC cells is not clear. Here, we evaluated the potential roles of EGFR signaling cascade that affects RCC progression, and also investigated the inhibitory effect of silibinin on the EGFR signal-induced migration and invasion abilities of RCC cells. Our data indicated that silibinin inhibited migration and invasion of RCC cells in a dose-dependent manner via blocking the EGFR signal, especially in the EGFR highly expressing RCC cells. Silibinin inhibited phosphorylation of EGFR and its downstream molecules ERK1/2 but did not affect phosphorylation of other downstream molecules, STAT3 and Akt, in human RCC cell lines. Moreover, our data suggested that silibinin significantly reduced the MMP-9 expression and its activity that was promoted by the EGFR signal, and also suppressed MMP9-dependent migration and invasion abilities of RCC cells. Taken together, this study clearly demonstrated that silibinin inhibited EGFR induced migration and invasion of RCC cells via blockade of EGFR/MMP-9 signaling. Thus, we suggest that silibinin could be used as a potential effective drug for the inhibition of RCC metastasis.

Holmium laser enucleation of the prostate: a modified enucleation technique and initial results.

PURPOSE: Although holmium laser enucleation of the prostate has been proven to be an excellent technique for the treatment of benign prostatic hyperplasia, it has not been widely applied due to technical difficulties and longer operative time. We modified the current technique of enucleation and present our initial experience. MATERIALS AND METHODS: A total of 189 patients with benign prostatic hyperplasia underwent prostatectomy with our modified technique for holmium laser enucleation of the prostate. Intraoperative and postoperative data were prospectively collected. For followup International Prostate Symptom Score, quality of life, maximal flow rate and post-void residual urine were recorded. RESULTS: Mean±SD preoperative prostate volume was 78.1±24.3 cc and 60.9±39.2 gm tissue were enucleated. Mean operative and enucleation times were 54.7±21.1 and 36.5±16.3 minutes, respectively. Mean serum hemoglobin decrease was 0.98±0.72 gm/dl. Mean catheter time was 1.2±0.5 days and mean postoperative hospital stay was 4.9±3.4 days. Serious complications were not observed. Three patients complained of transient stress incontinence which resolved within 3 months. Significant improvement occurred in International Prostate Symptom Score, quality of life, maximal flow rate and post-void residual urine volume at 3 and 6-month followup compared with the preoperative baseline. CONCLUSIONS: The modified holmium laser enucleation of the prostate technique is effective and safe when treating benign prostatic hyperplasia.