Osteopontin is a pleiotropic cytokine that is involved in several diseases including multiple sclerosis. Secreted osteopontin is cleaved by few known proteases, modulating its pro-inflammatory activities. Here we show by in vitro experiments that secreted osteopontin can be processed by extracellular proteasomes, thereby producing fragments with novel chemotactic activity. Furthermore, osteopontin reduces the release of proteasomes in the extracellular space. The latter phenomenon seems to occur in vivo in multiple sclerosis, where it reflects the remission/relapse alternation. The extracellular proteasome-mediated inflammatory pathway may represent a general mechanism to control inflammation in inflammatory diseases.

Osteopontin is a proinflammatory cytokine and plays a pathogenetic role in multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), by recruiting autoreactive T cells into the central nervous system. Osteopontin functions are modulated by thrombin cleavage generating N- and C-terminal fragment, whose individual roles are only partly known. Published data are difficult to compare since they have been obtained with heterogeneous approaches. Interestingly, thrombin cleavage of osteopontin unmasks a cryptic domain of interaction with α 4 β 1 integrin that is the main adhesion molecule involved in lymphocyte transmigration to the brain and is the target for natalizumab, the most potent drug preventing relapses. We produced recombinant osteopontin and its N- and C-terminal fragments in an eukaryotic system in order to allow their posttranslational modifications. We investigated, in vitro, their effect on human cells and in vivo in EAE. We found that the osteopontin cleavage plays a key role in the function of this cytokine and that the two fragments exert distinct effects both in vitro and in vivo. These findings suggest that drugs targeting each fragment may be used to fine-tune the pathological effects of osteopontin in several diseases.

Aim: To develop an innovative delivery system for temozolomide (TMZ) in solid lipid nanoparticles (SLN), which has been preliminarily investigated for the treatment of melanoma. Materials and Methods: SLN-TMZ was obtained through fatty acid coacervation. Its pharmacological effects were assessed and compared with free TMZ in in vitro and in vivo models of melanoma and glioblastoma. Results: Compared to the standard free TMZ, SLN-TMZ exerted larger effects, when cell proliferation of melanoma cells, and neoangiogeneis were evaluated. SLN-TMZ also inhibited growth and vascularization of B16-F10 melanoma in C57/BL6 mice, without apparent toxic effects. Conclusion: SLN could be a promising strategy for the delivery of TMZ, allowing an increased stability of the drug and thereby its employment in the treatment of aggressive malignacies.

Background: Ailanthone (Aila) is a natural active compound isolated from the Ailanthus altissima,-inhibitory effect against several cancer cell lines. Advanced bladder cancer is a common disease characterized by a frequent onset of resistance to cisplatin-based therapy. The cisplatin (CDDP) resistance is accompanied by an increase in Nrf2 protein expression which contributes to conferring resistance. Recently, we demonstrated a crosstalk between Nrf2 and YAP. YAP has also been demonstrated to play an important role in chemoresistance of bladder cancer. Purpose: We analyzed the antitumor effect of Aila in sensitive and CDDP-resistant bladder cancer cells and the molecular mechanisms involved in Aila activity. Study design: Sensitive and CDDP-resistant 253J B-V and 253J bladder cancer cells, and intrinsically CDDP-resistant T24 bladder cancer cells were used. Cells were treated with diverse concentrations of Aila and proliferation, cell cycle, apoptosis and gene expressions were determined. Methods: Aila toxicity and proliferation were determined by MTT and colony forming methods, respectively. Cell cycle was determined at cytofluorimeter by PI staining method Apoptosis was detected using Annexin V and PI double staining followed by quantitative flow cytometry. Expressions of Nrf2, Yap, c-Myc, and housekeeping genes were determined by western blot with specific antibodies. Cell migration was detected by wound healing and Boyden chamber analysis. Results: Aila inhibited growth of sensitive and CDDP-resistant bladder cancer cells with the same effectiveness, and reduced cell migration with higher effectiveness in CDDP resistant cells. Interestingly, Aila strongly reduced Nrf2 expression in all cell lines. Cell cycle analysis revealed an accumulation of Aila-treated cells in G0/G1 phase. Moreover, Aila significantly reduced YAP and c-Myc protein expression. The random and the oriented migration were strongly inhibited by Aila treatment, in particular in CDDP-resistant cells. Conclusions: Aila, inhibited proliferation and invasiveness of bladder cancer cells. Its high effectiveness in CDDP resistant cells could be related to the inhibition of Nrf2 , YAP and c-Myc expressions. Aila could represent a new tool to overcome CDDP resistance in bladder cancer.

Background: Solid lipid nanoparticles (SLNs) are attractive drug delivery systems for lipophilic molecules like curcumin (CURC) with low chemical stability. Methods: A simple, innovative, and cold-operating method, named “cold dilution of microemulsion” is developed by the authors to produce SLNs. An oil-in-water microemulsion (µE), whose disperse phase consisted of a solution of trilaurin in a partially water-miscible solvent, was prepared after mutually saturating solvent and water. Trilaurin SLNs precipitated following solvent removal upon water dilution of the µE. After SLN characterization (mean size, Zeta potential, CURC entrapment efficiency, and over time stability), they were tested for in vitro cytotoxicity studies on pancreatic adenocarcinoma cell lines and for in vivo preliminary biodistribution studies in Wistar healthy rats. Results: CURC loaded SLNs (SLN-CURC) had mean diameters around 200 nm, were negatively charged, stable over time, and able to entrap CURC up to almost 90%, consequently improving its stability. SLN-CURC inhibited in vitro pancreatic carcinoma cell growth in concentration-dependent manner. Their in vivo intravenous administration suggested a possible long circulation. Conclusions: These results, according to a concomitant study on chitosan-coated SLNs, confirm the possibility to apply the developed SLN-based delivery systems as a means to entrap CURC, to improve both its water dispersibility and chemical stability, facilitating its application in therapy.

No abstract

Strigolactones (SLs) are carotenoid-derived plant hormones that exhibit anti-cancer activities. We previously demonstrated that two SL analogues, MEB55 and ST362, inhibit the growth and survival of various cancer cell lines. However, these compounds have low aqueous solubility and stability at physiological pH. Here, we generated SL-loaded glutathione/pH-responsive nanosponges (GSH/pH-NS) to selectively deliver SLs to prostate cancer cells and enhance their therapeutic efficacy. The SLs were readily incorporated into the GSH/pH-NS. The drug loading efficiency was 13.9% for MEB55 and 15.4% for ST362, and the encapsulation efficiency was 88.7% and 96.5%, respectively. Kinetic analysis revealed that release of MEB55 and ST362 from the GSH/pH-NS was accelerated at acidic pH and in the presence of a high GSH concentration. Evaluation of the effects of MEB55- and ST362-loaded GSH/pH-NS on the growth of DU145 (high GSH) and PC-3 (low GSH) prostate cancer cells revealed that the GSH/pH-NS inhibited the proliferation of DU145 cells to a greater extent than free MEB55 or ST362 over a range of concentrations. These findings indicate GSH/pH-NS are efficient tools for controlled delivery of SLs to prostate cancer cells and may enhance the therapeutic efficacy of these compounds.

Doxorubicin (DOX) is an anthracycline widely used in cancer therapy and in particular in breast cancer treatment. The treatment with DOX appears successful, but it is limited by a severe cardiotoxicity. This work evaluated the in vitro and in vivo anticancer effect of a new formulation of β-cyclodextrin nanosponges containing DOX (BNS-DOX). The BNS-DOX effectiveness was evaluated in human and mouse breast cancer cell lines in vitro in terms of effect on cell growth, cell cycle distribution, and apoptosis induction; and in vivo in BALB-neuT mice developing spontaneous breast cancer in terms of biodistribution, cancer growth inhibition, and heart toxicity. BNS-DOX significantly inhibited cancer cell proliferation, through the induction of apoptosis, with higher efficiency than free DOX. The breast cancer growth in BALB-neuT mice was inhibited by 60% by a BNS-DOX dose five times lower than the DOX therapeutic dose, with substantial reduction of tumor neoangiogenesis and lymphangiogenesis. Biodistribution after BNS-DOX treatment revealed a high accumulation of DOX in the tumor site and a low accumulation in the hearts of mice. Results indicated that use of BNS may be an efficient strategy to deliver DOX in the treatment of breast cancer, since it improves the anti-cancer effectiveness and reduces cardiotoxicity.