Focal therapy for primary tumor and metastases in de novo or recurrent oligometastatic prostate cancer: current standing and future perspectives.

PURPOSE: Focal therapy (FT) is gaining increasing acceptance in the management of localized prostate cancer particularly due to its favorable safety. Preliminary evidence suggests advantageous utilization of local treatment in the field of oligometastatic prostate cancer (OMPC). Since data on the utilization of FT in OMPC are scarce, we sought to summarize available evidence. METHODS: For this narrative comprehensive review, we employed PubMed®, Web of Science™, Embase®, Scopus®, and clinicaltrial.gov databases and Google web search engine to seek peer-reviewed articles, published abstracts from international congresses, and ongoing trials in the English language using the terms "prostate cancer", "oligometastatic", "hormone-sensitive", "focal therapy", "focal treatment", "cryotherapy", "ablation", "cancer" as well as "metastasis-directed therapy. We focused on relevant publications on FT utilized in OMPC targeting the primary or metastatic sites as well as completed and ongoing clinical trials. RESULTS: Growing evidence points to distinct differences in the biologic behavior and molecular signaling processes of OMPC as compared to polymetastatic disease (PMPC). No established biomarkers are available to accurately identify OMPC yet, while several candidates are currently under investigation. The evolution of molecular imaging is set to aid in selecting patients benefitting most from local management. Differences between OMPC and PMPC should be considered when designing the optimal therapeutic strategy. While efficacy data for FT in comparison to standard care in OMPC are scarce, longer progression-free survival and time to castration resistance have been demonstrated for bone metastatic prostate cancer with the primary tumor treated by cryosurgery followed by androgen deprivation therapy (ADT) compared to ADT alone. CONCLUSION: Ongoing research efforts are eagerly awaited to better characterize OMPC and establish customized strategies for patients with this condition.

Cucumis sativus (Cucurbitaceae) seed oil prevents benzo(a)pyrene-induced prostate cancer in vitro and in vivo.

Despite enormous progress in modern medicine, prostate cancer (PCa) remains a major public health problem due to its high incidence and mortality. Although studies have shown in vitro antitumor effects of cucurbitacins from Cucumis sativus, the in vivo anticancer effect of the seed oil as a whole, has yet to be demonstrated. The present study evaluated the in vitro anticancer mechanisms of C. sativus (CS) seed oil and its possible chemopreventive potential on benzo(a)pyrene (BaP)-induced PCa in Wistar rat. In vitro cell growth, clone formation, cell death mechanism, cell adhesion and migration as well as expression of integrins ß-1 and ß-4 were assessed. In vivo PCa was induced in 56 male rats versus 8 normal control rats, randomized in normal (NOR) and negative (BaP) control groups which, received distilled water; the positive control group (Caso) was treated with casodex (13.5 mg/kg BW). One group received the total seed extract at the dose of 500 mg/kg BW; while the remaining three groups were treated with CS seed oil at 42.5, 85, and 170 mg/kg BW. The endpoints were: morphologically (prostate tumor weight and volume), biochemically (total protein, prostate specific antigen (PSA), oxidative stress markers such as MDA, GSH, catalase, and SOD) and histologically. As results, CS seed oil significantly and concentration-dependently reduced the DU145 prostate cancer cell growth and clone formation (optimum = 100 µg/mL). It slightly increased the number of apoptotic cells and inhibited the migration and invasion of DU145 cells, while it decreased their adhesion to immobilized collagen and fibrinogen. The expression of integrin ß-1 and ß-4 was increased in presence of 100 µg/mL CS oil. In vivo, the BaP significantly elevated the incidence of PC tumors (75%), the total protein and PSA levels, pro-inflammatory cytokines (TNF-α, IL-1, and IL-6) and MDA levels compared to NOR. CS seeds oil significantly counteracted the effect of BaP by decreasing significantly the PC incidence (12.5%), and increasing the level of antioxidant (SOD, GSH, and catalase) and anti-inflammatory cytokine IL-10 in serum. While in BaP group PCa adenocarninoma was the most representative neoplasm, rats treated with 85 and 170 mg/kg prevented it in the light of the casodex. It is conclude that CS may provide tumor suppressive effects in vitro and in vivo which makes it an interesting candidate to support the current treatment protocol.

Curcumin 3.0-Therapeutic and Diagnostic Potential in Cancer and Beyond.

Curcumin is one of the most interesting plant-derived polyphenols with a high potential for therapeutic, and even diagnostic, application in various diseases [...].

Allyl-, Butyl- and Phenylethyl-Isothiocyanate Modulate Akt-mTOR and Cyclin-CDK Signaling in Gemcitabine- and Cisplatin-Resistant Bladder Cancer Cell Lines.

Combined cisplatin-gemcitabine treatment causes rapid resistance development in patients with advanced urothelial carcinoma. The present study investigated the potential of the natural isothiocyanates (ITCs) allyl-isothiocyanate (AITC), butyl-isothiocyanate (BITC), and phenylethyl-isothiocyanate (PEITC) to suppress growth and proliferation of gemcitabine- and cisplatin-resistant bladder cancer cells lines. Sensitive and gemcitabine- and cisplatin-resistant RT112, T24, and TCCSUP cells were treated with the ITCs, and tumor cell growth, proliferation, and clone formation were evaluated. Apoptosis induction and cell cycle progression were investigated as well. The molecular mode of action was investigated by evaluating cell cycle-regulating proteins (cyclin-dependent kinases (CDKs) and cyclins A and B) and the mechanistic target of the rapamycin (mTOR)-AKT signaling pathway. The ITCs significantly inhibited growth, proliferation and clone formation of all tumor cell lines (sensitive and resistant). Cells were arrested in the G2/M phase, independent of the type of resistance. Alterations of both the CDK-cyclin axis and the Akt-mTOR signaling pathway were observed in AITC-treated T24 cells with minor effects on apoptosis induction. In contrast, AITC de-activated Akt-mTOR signaling and induced apoptosis in RT112 cells, with only minor effects on CDK expression. It is concluded that AITC, BITC, and PEITC exert tumor-suppressive properties on cisplatin- and gemcitabine-resistant bladder cancer cells, whereby the molecular action may differ among the cell lines. The integration of these ITCs into the gemcitabine-/cisplatin-based treatment regimen might optimize bladder cancer therapy.

Sulforaphane Impact on Reactive Oxygen Species (ROS) in Bladder Carcinoma.

Sulforaphane (SFN) is a natural glucosinolate found in cruciferous vegetables that acts as a chemopreventive agent, but its mechanism of action is not clear. Due to antioxidative mechanisms being thought central in preventing cancer progression, SFN could play a role in oxidative processes. Since redox imbalance with increased levels of reactive oxygen species (ROS) is involved in the initiation and progression of bladder cancer, this mechanism might be involved when chemoresistance occurs. This review summarizes current understanding regarding the influence of SFN on ROS and ROS-related pathways and appraises a possible role of SFN in bladder cancer treatment.

Growth, Proliferation and Metastasis of Prostate Cancer Cells Is Blocked by Low-Dose Curcumin in Combination with Light Irradiation.

Although anti-cancer properties of the natural compound curcumin have been reported, low absorption and rapid metabolisation limit clinical use. The present study investigated whether irradiation with visible light may enhance the inhibitory effects of low-dosed curcumin on prostate cancer cell growth, proliferation, and metastasis in vitro. DU145 and PC3 cells were incubated with low-dosed curcumin (0.1-0.4 µg/mL) and subsequently irradiated with 1.65 J/cm2 visible light for 5 min. Controls remained untreated and/or non-irradiated. Cell growth, proliferation, apoptosis, adhesion, and chemotaxis were evaluated, as was cell cycle regulating protein expression (CDK, Cyclins), and integrins of the α- and ß-family. Curcumin or light alone did not cause any significant effects on tumor growth, proliferation, or metastasis. However, curcumin combined with light irradiation significantly suppressed tumor growth, adhesion, and migration. Phosphorylation of CDK1 decreased and expression of the counter-receptors cyclin A and B was diminished. Integrin α and ß subtypes were also reduced, compared to controls. Irradiation distinctly enhances the anti-tumor potential of curcumin in vitro and may hold promise in treating prostate cancer.

Amygdalin Modulates Prostate Cancer Cell Adhesion and Migration In Vitro.

The natural compound, amygdalin, is notably popular with prostate cancer patients as an alternative or complementary treatment option. However, knowledge about its mode of action is sparse. We investigated amygdalin's impact on prostate cancer adhesion and motile behavior. DU-145 and PC3 cancer cells were exposed to amygdalin. Adhesion to human vascular endothelium or immobilized collagen was then explored. The influence of amygdalin on chemotaxis and migration was also investigated, as well as amygdalin induced alteration to surface and total cellular α and ß integrin expression. Integrin knockdown was performed to evaluate the integrin influence on chemotaxis and adhesion. Amygdalin significantly reduced chemotactic activity, migration, and adhesion of DU-145 but not of PC3 cells. Amygdalin elevated integrin α2 in both cell lines. Integrin α6 was reduced by amygdalin only in DU-145 cells, whereas ß1 increased only in PC3 cells. Functional blocking revealed a negative association of α2 with PC3 and DU-145 chemotaxis. The ß1 increase correlated with enhanced chemotaxis, the diminished α6 expression with reduced chemotaxis. Amygdalin acted on prostate cancer cells in vitro. It induced downregulation of α6 integrin in DU-145 but not in PC3 cells, suggesting that exposing certain prostate cancer cells to amygdalin might inhibit metastatic spread promoted by this particular integrin.

Ethanol-extracted Cameroonian propolis: Antiproliferative effects and potential mechanism of action in prostate cancer.

The present study was conducted to evaluate in vitro and in vivo antiproliferative potential of the Cameroonian propolis and to elucidate its underlying mechanism. In vitro, ethanol-extracted propolis (EEP) was tested on cell growth, cell proliferation, cell cycle, cell death mechanism and cell migration. The cell cycle- and apoptosis-regulating proteins were assessed by Western blotting. In vivo the testosterone-induced benign prostatic hyperplasia (BPH) in Wistar rat was used to evaluate the antiproliferative potential of EEP. EEP reduced DU145 and PC3 cell survival with an IC50 of 70 and 22 µg/ml respectively. It increased the number of late apoptotic cells, the amount of cells in G0/G1 phase in DU145 and PC3 cells at 50 µg/ml. Cell cycle proteins (cdk1, pcdk1 and their related cyclins A and B) were down-regulated in both DU145 and PC3 cells, while cdk2 and pcdk2 were down-regulated only in PC3 cells. The pro-apoptotic Bax protein was up-regulated, while the anti-apoptotic Akt and pAKT, and Bcl-2 proteins were down-regulated. It increased prostate cell adhesion and chemotaxis. EEP reduced prostate weight, volume and epithelial thickness in rats. We demonstrated for the first time that Cameroonian propolis is endowed with in vitro and in vivo antiproliferative properties in the prostate.

Bladder Cancer Metastasis Induced by Chronic Everolimus Application Can Be Counteracted by Sulforaphane In Vitro.

Chronic treatment with the mTOR inhibitor, everolimus, fails long-term in preventing tumor growth and dissemination in cancer patients. Thus, patients experiencing treatment resistance seek complementary measures, hoping to improve therapeutic efficacy. This study investigated metastatic characteristics of bladder carcinoma cells exposed to everolimus combined with the isothiocyanate sulforaphane (SFN), which has been shown to exert cancer inhibiting properties. RT112, UMUC3, or TCCSUP bladder carcinoma cells were exposed short- (24 h) or long-term (8 weeks) to everolimus (0.5 nM) or SFN (2.5 µM), alone or in combination. Adhesion and chemotaxis along with profiling details of CD44 receptor variants (v) and integrin α and ß subtypes were evaluated. The functional impact of CD44 and integrins was explored by blocking studies and siRNA knock-down. Long-term exposure to everolimus enhanced chemotactic activity, whereas long-term exposure to SFN or the SFN-everolimus combination diminished chemotaxis. CD44v4 and v7 increased on RT112 cells following exposure to SFN or SFN-everolimus. Up-regulation of the integrins α6, αV, and ß1 and down-regulation of ß4 that was present with everolimus alone could be prevented by combining SFN and everolimus. Down-regulation of αV, ß1, and ß4 reduced chemotactic activity, whereas knock-down of CD44 correlated with enhanced chemotaxis. SFN could, therefore, inhibit resistance-related tumor dissemination during everolimus-based bladder cancer treatment.

Sulforaphane Reduces Prostate Cancer Cell Growth and Proliferation In Vitro by Modulating the Cdk-Cyclin Axis and Expression of the CD44 Variants 4, 5, and 7.

Prostate cancer patients whose tumors develop resistance to conventional treatment often turn to natural, plant-derived products, one of which is sulforaphane (SFN). This study was designed to determine whether anti-tumor properties of SFN, identified in other tumor entities, are also evident in cultivated DU145 and PC3 prostate cancer cells. The cells were incubated with SFN (1-20 µM) and tumor cell growth and proliferative activity were evaluated. Having found a considerable anti-growth, anti-proliferative, and anti-clonogenic influence of SFN on both prostate cancer cell lines, further investigation into possible mechanisms of action were performed by evaluating the cell cycle phases and cell-cycle-regulating proteins. SFN induced a cell cycle arrest at the S- and G2/M-phase in both DU145 and PC3 cells. Elevation of histone H3 and H4 acetylation was also evident in both cell lines following SFN exposure. However, alterations occurring in the Cdk-cyclin axis, modification of the p19 and p27 proteins and changes in CD44v4, v5, and v7 expression because of SFN exposure differed in the two cell lines. SFN, therefore, does exert anti-tumor properties on these two prostate cancer cell lines by histone acetylation and altering the intracellular signaling cascade, but not through the same molecular mechanisms.

Chronic Sulforaphane Administration Inhibits Resistance to the mTOR-Inhibitor Everolimus in Bladder Cancer Cells.

Progressive bladder cancer growth is associated with abnormal activation of the mammalian target of the rapamycin (mTOR) pathway, but treatment with an mTOR inhibitor has not been as effective as expected. Rather, resistance develops under chronic drug use, prompting many patients to lower their relapse risk by turning to natural, plant-derived products. The present study was designed to evaluate whether the natural compound, sulforaphane (SFN), combined with the mTOR inhibitor everolimus, could block the growth and proliferation of bladder cancer cells in the short- and long-term. The bladder cancer cell lines RT112, UMUC3, and TCCSUP were exposed short- (24 h) or long-term (8 weeks) to everolimus (0.5 nM) or SFN (2.5 µM) alone or in combination. Cell growth, proliferation, apoptosis, cell cycle progression, and cell cycle regulating proteins were evaluated. siRNA blockade was used to investigate the functional impact of the proteins. Short-term application of SFN and/or everolimus resulted in significant tumor growth suppression, with additive inhibition on clonogenic tumor growth. Long-term everolimus treatment resulted in resistance development characterized by continued growth, and was associated with elevated Akt-mTOR signaling and cyclin-dependent kinase (CDK)1 phosphorylation and down-regulation of p19 and p27. In contrast, SFN alone or SFN+everolimus reduced cell growth and proliferation. Akt and Rictor signaling remained low, and p19 and p27 expressions were high under combined drug treatment. Long-term exposure to SFN+everolimus also induced acetylation of the H3 and H4 histones. Phosphorylation of CDK1 was diminished, whereby down-regulation of CDK1 and its binding partner, Cyclin B, inhibited tumor growth. In conclusion, the addition of SFN to the long-term everolimus application inhibits resistance development in bladder cancer cells in vitro. Therefore, sulforaphane may hold potential for treating bladder carcinoma in patients with resistance to an mTOR inhibitor.

Curcumin-A Viable Agent for Better Bladder Cancer Treatment.

Although the therapeutic armamentarium for bladder cancer has considerably widened in the last few years, severe side effects and the development of resistance hamper long-term treatment success. Thus, patients turn to natural plant products as alternative or complementary therapeutic options. One of these is curcumin, the principal component of Curcuma longa that has shown chemopreventive effects in experimental cancer models. Clinical and preclinical studies point to its role as a chemosensitizer, and it has been shown to protect organs from toxicity induced by chemotherapy. These properties indicate that curcumin could hold promise as a candidate for additive cancer treatment. This review evaluates the relevance of curcumin as an integral part of therapy for bladder cancer.

Growth and Proliferation of Renal Cell Carcinoma Cells Is Blocked by Low Curcumin Concentrations Combined with Visible Light Irradiation.

The anti-cancer properties of curcumin in vitro have been documented. However, its clinical use is limited due to rapid metabolization. Since irradiation of curcumin has been found to increase its anti-cancer effect on several tumor types, this investigation was designed to determine whether irradiation with visible light may enhance the anti-tumor effects of low-dosed curcumin on renal cell carcinoma (RCC) cell growth and proliferation. A498, Caki1, and KTCTL-26 cells were incubated with curcumin (0.1⁻0.4 µg/mL) and irradiated with 1.65 J/cm² visible light for 5 min. Controls were exposed to curcumin or light alone or remained untreated. Curcumin plus light, but not curcumin or light exposure alone altered growth, proliferation, and apoptosis of all three RCC tumor cell lines. Cells were arrested in the G0/G1 phase of the cell cycle. Phosphorylated (p) CDK1 and pCDK2, along with their counter-receptors Cyclin B and A decreased, whereas p27 increased. Akt-mTOR-signaling was suppressed, the pro-apoptotic protein Bcl-2 became elevated, and the anti-apoptotic protein Bax diminished. H3 acetylation was elevated when cells were treated with curcumin plus light, pointing to an epigenetic mechanism. The present findings substantiate the potential of combining low curcumin concentrations and light as a new therapeutic concept to increase the efficacy of curcumin in RCC.

Knockdown of BAG3 sensitizes bladder cancer cells to treatment with the BH3 mimetic ABT-737.

PURPOSE: BAG3 is overexpressed in several malignancies and mediates a non-canonical, selective form of (macro)autophagy. By stabilizing pro-survival Bcl-2 proteins in complex with HSP70, BAG3 can also exert an apoptosis-antagonizing function. ABT-737 is a high affinity Bcl-2 inhibitor that fails to target Mcl-1. This failure may confer resistance in various cancers. METHODS: Urothelial cancer cells were treated with the BH3 mimetics ABT-737 and (-)-gossypol, a pan-Bcl-2 inhibitor which inhibits also Mcl-1. To clarify the importance of the core autophagy regulator ATG5 and BAG3 in ABT-737 treatment, cell lines carrying a stable lentiviral knockdown of ATG5 and BAG3 were created. The synergistic effect of ABT-737 and pharmaceutical inhibition of BAG3 with the HSF1 inhibitor KRIBB11 or sorafenib was also evaluated. Total cell death and apoptosis were quantified by FACS analysis of propidium iodide, annexin. Target protein analysis was conducted by Western blotting. RESULTS: Knockdown of BAG3 significantly downregulated Mcl-1 protein levels and sensitized urothelial cancer cells to apoptotic cell death induced by ABT-737, while inhibition of bulk autophagy through depletion of ATG5 had no discernible effect on cell death. Similar to knockdown of BAG3, pharmacological targeting of the BAG3/Mcl-1 pathway with KRIBB11 was capable to sensitize both cell lines to treatment with ABT-737. CONCLUSION: Our results show that BAG3, but not bulk autophagy has a major role in the response of bladder cancer cells to BH3 mimetics. They also suggest that BAG3 is a suitable target for combined therapies aimed at synergistically inducing apoptosis in bladder cancer.

Molecular analysis of sunitinib resistant renal cell carcinoma cells after sequential treatment with RAD001 (everolimus) or sorafenib.

Sequential application of target drugs is standard procedure after renal cell carcinoma (RCC) patients develop resistance. To optimize the sequence, antitumour effects of the mTOR inhibitor RAD001 or the tyrosine kinase inhibitor (TKI) sorafenib on RCC cells with acquired resistance to the TKI sunitinib was evaluated. RCC cells were exposed to 1 µM sunitinib for 24 hrs (as control) and for 8 weeks (to induce resistance) and then switched to RAD001 (5 nM) or sorafenib (5 µM) for a further 8 weeks. Tumour cell growth, cell cycle progression, cell cycle regulating proteins and intracellular signalling were then investigated. Short-term application of sunitinib (24 hrs) induced cell growth blockade with accumulation in the G2/M phase. RCC cells became resistant to sunitinib after 8 weeks, demonstrated by accelerated cell growth along with enhanced cdk1, cdk2, loss of p27, activation of Akt, Rictor and Raptor. Switching to sorafenib only slightly reduced growth of the sunitinib resistant RCC cells and molecular analysis indicated distinct cross-resistance. In contrast, full response was achieved when the cancer cells were treated with RAD001. p19 and p27 strongly increased, phosphorylated Akt, Rictor and Raptor decreased and the tumour cells accumulated in G0/G1. It is concluded that an mTOR-inhibitor for second-line therapy could be the strategy of choice after first-line sunitinib failure.

Intensified antineoplastic effect by combining an HDAC-inhibitor, an mTOR-inhibitor and low dosed interferon alpha in prostate cancer cells.

A significant proportion of men diagnosed with prostate cancer (PCa) eventually develop metastatic disease, which progresses to castration resistance, despite initial response to androgen deprivation. As anticancer therapy has become increasingly effective, acquired drug resistance has emerged, limiting efficacy. Combination treatment, utilizing different drug classes, exemplifies a possible strategy to foil resistance development. The effects of the triple application of the histone deacetylase (HDAC) inhibitor valproic acid (VPA), the mammalian target of rapamycin inhibitor everolimus and low dosed interferon alpha (IFNα) on PCa cell growth and dissemination capacity were investigated. For that purpose, the human PCa cell lines, PC-3, DU-145 and LNCaP were treated with the combined regimen or separate single agents. Cell growth was investigated by the MTT dye reduction assay. Flow cytometry served to analyse cell cycle progression. Adhesion to vascular endothelium or immobilized collagen, fibronectin and laminin was quantified. Migration and invasion characteristics were determined by the modified Boyden chamber assay. Integrin α and ß subtypes were investigated by flow cytometry, western blotting and RT-PCR. Integrin related signalling, Epidermal Growth Factor Receptor (EGFr), Akt, p70S6kinase and extracellular signal-regulated kinases (ERK)1/2 activation were also assessed. The triple application of VPA, everolimus and low dosed IFNα blocked tumour cell growth and dissemination significantly better than any agent alone. Antitumour effects were associated with pronounced alteration in the cell cycle machinery, intracellular signalling and integrin expression profile. Combining VPA, everolimus and low dosed IFNα might be a promising option to counteract resistance development and improve outcome in PCa patients.

Latent heparanase facilitates VLA-4-mediated melanoma cell binding and emerges as a relevant target of heparin in the interference with metastatic progression.

Heparanase is an endo-ß-glucuronidase that enzymatically cleaves heparan sulfates (HS) and heparan sulfate proteoglycan (HSPG) structures. Heparanase expression levels by tumors were correlated with cell invasion, angiogenic activity, and poor prognosis. Heparanase can also possess pro-tumorigenic effects independent of its enzymatic activity. Using human melanoma MV3 cells, we demonstrate that latent heparanase activates in a tightly temporary-regulated manner the binding function of the integrin very late antigen-4 (VLA-4), an important component in the metastatic spread of melanoma cells. shRNA-mediated knockdown of syndecan-4 (SDC-4) indicated that this proteoglycan is the key element to convey heparanase binding via focal adhesion complex formation, detected by vinculin staining, to an upregulated VLA-4 binding function. This inside-out signaling pathway of VLA-4 involved activated FAK and Akt, but apparently not PKCα/δ. VLA-4, however, appears representative of other integrins which together impact the heparanase/integrin activation axis in tumorigenicity. Biosensor measurements provided an insight as to how heparin can interfere with this activation process. While low-molecular-weight heparin (LMWH) cannot replace heparanase bound to SDC-4, LMWH can compete with SDC-4 binding of heparanase. Since blockade of heparanase by LMWH has functional consequences for reduced VLA-4 binding, latent heparanase appears as a novel, so far unnoticed target of heparin, underlying its antimetastatic activity.

Chemoresistance is associated with increased cytoprotective autophagy and diminished apoptosis in bladder cancer cells treated with the BH3 mimetic (-)-Gossypol (AT-101).

BACKGROUND: Acquired resistance to standard chemotherapy causes treatment failure in patients with metastatic bladder cancer. Overexpression of pro-survival Bcl-2 family proteins has been associated with a poor chemotherapeutic response, suggesting that Bcl-2-targeted therapy may be a feasible strategy in patients with these tumors. The small-molecule pan-Bcl-2 inhibitor (-)-gossypol (AT-101) is known to induce apoptotic cell death, but can also induce autophagy through release of the pro-autophagic BH3 only protein Beclin-1 from Bcl-2. The potential therapeutic effects of (-)-gossypol in chemoresistant bladder cancer and the role of autophagy in this context are hitherto unknown. METHODS: Cisplatin (5637(r)CDDP(1000), RT4(r)CDDP(1000)) and gemcitabine (5637(r)GEMCI(20), RT4(r)GEMCI(20)) chemoresistant sub-lines of the chemo-sensitive bladder cancer cell lines 5637 and RT4 were established for the investigation of acquired resistance mechanisms. Cell lines carrying a stable lentiviral knockdown of the core autophagy regulator ATG5 were created from chemosensitive 5637 and chemoresistant 5637(r)GEMCI(20) and 5637(r)CDDP(1000) cell lines. Cell death and autophagy were quantified by FACS analysis of propidium iodide, Annexin and Lysotracker staining, as well as LC3 translocation. RESULTS: Here we demonstrate that (-)-gossypol induces an apoptotic type of cell death in 5637 and RT4 cells which is partially inhibited by the pan-caspase inhibitor z-VAD. Cisplatin- and gemcitabine-resistant bladder cancer cells exhibit enhanced basal and drug-induced autophagosome formation and lysosomal activity which is accompanied by an attenuated apoptotic cell death after treatment with both (-)-gossypol and ABT-737, a Bcl-2 inhibitor which spares Mcl-1, in comparison to parental cells. Knockdown of ATG5 and inhibition of autophagy by 3-MA had no discernible effect on apoptotic cell death induced by (-)-gossypol and ABT-737 in parental 5637 cells, but evoked a significant increase in early apoptosis and overall cell death in BH3 mimetic-treated 5637(r)GEMCI(20) and 5637(r)CDDP(1000) cells. CONCLUSIONS: Our findings show for the first time that (-)-gossypol concomitantly triggers apoptosis and a cytoprotective type of autophagy in bladder cancer and support the notion that enhanced autophagy may underlie the chemoresistant phenotype of these tumors. Simultaneous targeting of Bcl-2 proteins and the autophagy pathway may be an efficient new strategy to overcome their "autophagy addiction" and acquired resistance to current therapy.

CCL2 promotes integrin-mediated adhesion of prostate cancer cells in vitro.

PURPOSE: Chemokines undergo alterations during neoplasia. However, knowledge about their functional significance in prostate cancer (PCa) progression is still sparse. Since chemokine (C-C motif) ligand 2 (CCL2) is significantly up-regulated in patients with PCa, aim of the current study was to assess whether CCL2 contributes to invasive behavior of prostate cancer cells in vitro. METHODS: The human PCa cell line PC3 was stimulated with CCL2. Cell growth was investigated by MTT dye reduction assay. Cell adhesion was analyzed by measuring attachment to a human endothelial cell (HUVEC) monolayer and immobilized collagen. Cell migration was assessed by a chemotactic assay. Integrin expression on the cell surface was evaluated by Western blot. Blocking studies were performed with anti-integrin α3, anti-integrin α6 and anti-integrin ß4 monoclonal antibodies. RESULTS: PC3 cell growth 72 h after CCL2 exposure was significantly increased, compared to controls. Activation of tumor cells by CCL2 significantly enhanced tumor cell adhesion to HUVEC and immobilized collagen. CCL2, added for 4 or 24 h, elevated α6 and ß4 (4 > 24 h) integrin expression. α3 was enhanced after 4 h, but reduced after 24 h. Blocking either α3, α6 or ß4 led to significant suppression of tumor cell binding to immobilized collagen. CONCLUSIONS: CCL2 stimulates PCa cell adhesion and induces alterations in α3-, α6- and ß4-integrin expression on the cell surface. Blocking these integrins leads to a significant reduction in cell adhesion.

Globalization in Urology: A Bibliographical Analysis of Cross-Continent Publication between 2002 and 2012.

INTRODUCTION: Asian scientists have now increasingly begun to contribute to globalization; yet it is not clear whether publishing in the field of urology is paralleled by elevated cross-continental scientific publishing. MATERIALS AND METHODS: An exemplary bibliometric analysis of urologic journals from 3 different continents was conducted between 2002 and 2012. Based on the ISI Web of Knowledge Journal Citation Reports, 2 urologic journals with similar impact factors (IFs) in 2013 were selected from Europe ('British Journal of Urology International', 'World Journal of Urology'), Asia ('International Journal of Urology', 'Asian Journal of Andrology') and North America ('Urologic Oncology-Seminars and Original Investigations', 'Urology'). The home continent of the journal, the workplace continental affiliation of the last author, article type (clinical, experimental or review) as well as the IF were documented. RESULTS: Most authors published their manuscripts in journals from the same continent in which they worked. However, a significant increase in cross-continental publishing was apparent from 2002 to 2012. Asians publishing in North America increased from 17% in 2002 to 35% in 2012. Europeans also increased the number of articles they published in North American journals, while publications from North American authors were shifted towards both European and Asian journals. Experimental and clinical articles showed significant increases in cross-continental publishing, while review publishing showed no significant change. The average IF for authors from all 3 continents increased from 2002 to 2012 (p < 0.001). The largest increase in the IF was found for Asian authors (0.11 per year). CONCLUSIONS: Cross-continental publication significantly increased during the period from 2002 to 2012. The impact that the Asian authors have experienced was found to be gradually impacting the North American and European colleagues.