QbD Enabled Development and Evaluation of Pazopanib Loaded Nanoliposomes for PDAC Treatment.

Pancreatic ductal adenocarcinoma (PDAC) is one of the highly fatal types of cancer with high mortality/incidence. Considering the crucial role of vascular endothelial growth factor (VEGF) in PDAC progression, its inhibition can be a viable strategy for the treatment. Pazopanib, a second-generation VEGF inhibitor, is approved for the treatment of various oncological conditions. However, due to associated limitations like low oral bioavailability (14-39%), high inter/intra-subject variability, stability issues, etc., high doses (800 mg) are required, which further lead to non-specific toxicities and also contribute toward cancer resistance. Thus, to overcome these challenges, pazopanib-loaded PEGylated nanoliposomes were developed and evaluated against pancreatic cancer cell lines. The nanoliposomes were prepared by thin-film hydration method, followed by characterization and stability studies. This QbD-enabled process design successfully led to the development of a suitable pazopanib liposomal formulation with desirable properties. The % entrapment of PZP-loaded non-PEGylated and PEGylated nanoliposomes was found to be 75.2% and 84.9%, respectively, whereas their particle size was found to be 129.7 nm and 182.0 nm, respectively. The developed liposomal formulations exhibited a prolonged release and showed desirable physicochemical properties. Furthermore, these liposomal formulations were also assessed for in vitro cell lines, such as cell cytotoxicity assay and cell uptake. These studies confirm the effectiveness of developed liposomal formulations against pancreatic cancer cell lines. The outcomes of this work provide encouraging results and a way forward to thoroughly investigate its potential for PDAC treatment.

Photothermal therapy enhance the anti-mitochondrial metabolism effect of lonidamine to renal cell carcinoma in homologous-targeted nanosystem.

Renal cell carcinoma (RCC) is a common malignant tumor of the urinary system with poor prognosis. Therapeutic drugs for RCC can easily develop resistance or have unignorable toxicity or limited efficiency. Here, the thermosensitive mitochondrial metabolism-interfering anticancer drug lonidamine (LND) was combined with the photothermal material polydopamine (PDA) to treat RCC. To delivery drugs accurately to RCC site, LND and PDA were loaded in stellate mesoporous silica nanoparticles (MSNs) with a large surface area and cloaked with RCC membranes (MLP@M). The results showed that MLP@M exhibited excellent tumor targeting ability. The synergistic effects of LND and PDA in MLP@M were greatly enhanced when triggered by an 808 nm laser. Moreover, the antiproliferative and tumor suppressing abilities were enhanced with good biocompatibility after MLP@M + laser treatment. Additionally, 80% of RCC tumor-bearing mice treated with MLP@M + laser did not relapse. Our study provides a potential therapeutic approach for RCC treatment.

Role and Regulation of Mechanotransductive HIF-1α Stabilisation in Periodontal Ligament Fibroblasts.

Orthodontic tooth movement (OTM) creates compressive and tensile strain in the periodontal ligament, causing circulation disorders. Hypoxia-inducible factor 1α (HIF-1α) has been shown to be primarily stabilised by compression, but not hypoxia in periodontal ligament fibroblasts (PDLF) during mechanical strain, which are key regulators of OTM. This study aimed to elucidate the role of heparan sulfate integrin interaction and downstream kinase phosphorylation for HIF-1α stabilisation under compressive and tensile strain and to which extent downstream synthesis of VEGF and prostaglandins is HIF-1α-dependent in a model of simulated OTM in PDLF. PDLF were subjected to compressive or tensile strain for 48 h. In various setups HIF-1α was experimentally stabilised (DMOG) or destabilised (YC-1) and mechanotransduction was inhibited by surfen and genistein. We found that HIF-1α was not stabilised by tensile, but rather by compressive strain. HIF-1α stabilisation had an inductive effect on prostaglandin and VEGF synthesis. As expected, HIF-1α destabilisation reduced VEGF expression, whereas prostaglandin synthesis was increased. Inhibition of integrin mechanotransduction via surfen or genistein prevented stabilisation of HIF-1α. A decrease in VEGF expression was observed, but not in prostaglandin synthesis. Stabilisation of HIF-1α via integrin mechanotransduction and downstream phosphorylation of kinases seems to be essential for the induction of VEGF, but not prostaglandin synthesis by PDLF during compressive (but not tensile) orthodontic strain.

Multifunctional Nanoparticles Loading with Docetaxel and GDC0941 for Reversing Multidrug Resistance Mediated by PI3K/Akt Signal Pathway.

The polylactic-co-glycolic acid polyethylene glycol conjugated with cell penetrating peptide R7 (PLGA-PEG-R7)/polysulfadimethoxine-folate nanoparticles loaded with docetaxel (DTX) and GDC0941 (R7/PSD-Fol NPs) were prepared to overcome multidrug resistance (MDR) and enhance the antitumor activity. First, polysulfadimethoxine-folate was synthesized to construct the R7/PSD-Fol NPs. The R7/PSD-Fol NPs were prepared with the abilities of effective entrapment and drug loading. Due to the pH-sensitive effect of PSD-folate, the releasing of DTX and GDC0941 from the R7/PSD-Fol NPs was lower in pH 7.4 buffer solution than that in pH 5.0 buffer solution. The half maximal inhibitory concentration (IC50) of MCF-7 and resistant to doxorubicin (MCF-7/Adr) cells illustrated the cytotoxicity of R7/PSD-Fol nanoparticles by using the MTT method. The uptake of R7/PSD-Fol NPs was visualized by using the fluorescence of Rh-123 to detect the targeting effect of folate on the surface of R7/PSD-Fol NPs. The results of the cell apoptosis and the depolarization of mitochondrial membrane potential (MMP) were adopted to show the cytotoxicity of the R7/PSD-Fol NPs on MCF-7/Adr cells. The Western blot revealed the inhibition of PI3K/Akt pathway in MCF-7/Adr cells induced by R7/PSD-Fol NPs. Finally, both in vivo distribution and in vivo antitumor showed the R7/PSD-Fol NPs displayed the better distribution at tumor site and the stronger suppression of tumor growth in the tumor bearing nude mice compared with control group. It was concluded that R7/PSD-Fol NPs loaded with DTX and GDC0941 could overcome MDR and enhance the antitumor effect further.

[Alteration mechanisms of oxidative stress at periodontal tissues of rats in a simulated periodontitis and elaborate methods of their correction].

INTRODUCTION: one of the peroxidation stress mechanisms is inducible NO synthase (iNOS) expression involved in the pathogenesis of periodontitis. AIM: to access the influence of isoform NO synthase (NOS) on alteration mechanisms of oxidative stress at periodontal tissues of 50 mature rats in a simulated periodontitis (SP). MATERIALS AND METHODS: a SP at rats was induced by a high-carbohydrate, high-fat (HCHF) diet. Тreated SP rat groups were intragastrically administered with selective neuronal NOS (nNOS) inhibitor 7-nitroindazole, selective inducible NOS (iNOS) inhibitor aminoguanidine, and nitric oxide synthase substrate L-arginine. Oxidative stress level in the homogenated soft periodontal tissues was evaluated by TBARS (thiobarbituric acid reactive substances) level before and after 1,5 hours of incubation. Antioxidant response was evaluated by the increase in concentration of TBARS for incubation, аnd by antioxidant enzyme activity - superoxide dismutase and catalase. RESULTS: nNOS activity increase in a SP considerably limits oxidative stress activation at periodontal tissues, decreases antioxidant response, but heightens catalase activity. iNOS functional activity stimulates oxidative stress at periodontal tissues of rats, decreases antioxidant response. L-arginine in a MS effectively repaired antioxidant response at periodontal tissues that probably will give positive result at complex treatment of periodontitis and MS generally. CONCLUSIONS: in the near future, the appropriate regulation of NO activity by using NOS-active agents may provide a novel strategy for the periodontal disease prevention and correction in a MS.

Hypoxia triggers a HIF-mediated differentiation of peripheral blood mononuclear cells into osteoclasts.

BACKGROUND: The source and mechanisms leading to osteoclast (OC) generation during tooth movement are not clearly understood. We hypothesized that during tooth movement, OC differentiate from peripheral blood mononuclear cells (PBMNC) downstream of the global hypoxia-inducible transcription factor hypoxia-inducible factor (HIF)-1α. OBJECTIVE: The objective of this study was to demonstrate up-regulation of OC growth factors from osteoblasts (OB) and subsequent conversion of PBMNC into functional OC under hypoxic stress. MATERIAL AND METHODS: Human primary PBMNC were cocultured with/without OB and subjected to either hypoxia (2.5% O2) or normoxia (21% O2) over 14 days. Levels of HIF, vascular endothelial growth factor (VEGF) and receptor activator for nuclear factor kappa-ß ligand (RANKL) were measured. Conversion of PBMNC into OC was measured using resorption and TRAP assays. RESULTS: Functional OC were only observed in response to hypoxia during coculture of PBMNC and OB and only after up-regulation of HIF, VEGF and RANKL in the hypoxic conditions. YC-1, a HIF inhibitor, reduced OC formation in response to hypoxia. CONCLUSION: Hypoxia triggers the differentiation of PBMNC into functional OC in the presence of OB in a HIF-dependent manner as would occur during orthodontic loading of the periodontal ligament space.

Development of EGFR-targeted polymer blend nanocarriers for combination paclitaxel/lonidamine delivery to treat multi-drug resistance in human breast and ovarian tumor cells.

Multi-drug resistant (MDR) cancer is a significant clinical obstacle and is often implicated in cases of recurrent, nonresponsive disease. Targeted nanoparticles were made by synthesizing a poly(D,L-lactide-co-glycolide)/poly(ethylene glycol)/epidermal growth factor receptor targeting peptide (PLGA/PEG/EGFR-peptide) construct for incorporation in poly(epsilon-caprolactone) (PCL) nanoparticles. MDR was induced in a panel of nine human breast and ovarian cancer cell lines using hypoxia. EGFR-targeted polymer blend nanoparticles were shown to actively target EGFR overexpressing cell lines, especially upon induction of hypoxia. The nanoparticles were capable of sustained drug release. Combination therapy with lonidamine and paclitaxel significantly improved the therapeutic index of both drugs. Treatment with a nanoparticle dose of 1 µM paclitaxel/10 µM lonidamine resulted in less than 10% cell viability for all hypoxic/MDR cell lines and less than 5% cell viability for all normoxic cell lines. Comparatively, treatment with 1 µM paclitaxel alone was the approximate IC50 value of the MDR cells while treatment with lonidamine alone had very little effect. The PLGA/PEG/EGFR-peptide delivery system actively targets a MDR cell by exploiting the expression of EGFR. This system treats MDR by inhibiting the Warburg effect and promoting mitochondrial binding of pro-apoptotic Bcl-2 proteins (lonidamine), while hyperstabilizing microtubules (paclitaxel). This nanocarrier system actively targets a MDR associated phenotype (EGFR receptor overexpression), further enhancing the therapeutic index of both drugs and potentiating the use of lonidamine/paclitaxel combination therapy in the treatment of MDR cancer.

Inhibition of CCL2 by bindarit alleviates diabetes-associated periodontitis by suppressing inflammatory monocyte infiltration and altering macrophage properties.

Diabetes-associated periodontitis (DP) aggravates diabetic complications and increases mortality from diabetes. DP is caused by diabetes-enhanced host immune-inflammatory responses to bacterial insult. In this study, we found that persistently elevated CCL2 levels in combination with proinflammatory monocyte infiltration of periodontal tissues were closely related to DP. Moreover, inhibition of CCL2 by oral administration of bindarit reduced alveolar bone loss and increased periodontal epithelial thickness by suppressing periodontal inflammation. Furthermore, bindarit suppressed the infiltration of proinflammatory monocytes and altered the inflammatory properties of macrophages in the diabetic periodontium. This finding provides a basis for the development of an effective therapeutic approach for treating DP.

Increase nitric oxide synthase activity in parotid glands from rats with experimental periodontitis.

OBJECTIVE: In this study we investigated the activity of the nitric oxide synthase (NOS) in parotid glands from rats with experimental periodontitis and controls. METHODS: Periodontitis was produced by a ligature placed around the cervix of the two lower first molar. Experiments were carried out 22 days after the ligature. RESULTS: Ligation caused an increase in parotid NOS activity. The selective blocker of the inducible isoform of the enzyme partially inhibited its activity in parotid glands from rat with ligature. In controls, the activity was partially inhibited by the antagonists of the selective neural and endothelial isoforms. NOS activity in rats with ligature was cyclic adenosine monophosphate (cAMP)-dependent while in controls it was calcium-dependent. Prostaglandin E2 concentration was increased in parotid gland from rats with ligature. The inhibitor of prostaglandin production, FR 122047, diminished both, prostaglandin production and NOS activity. In rats with ligature unstimulated amylase released is increased. Both, prostaglandin and NOS were involved in the increment of amylase release. CONCLUSION: It can be concluded that in parotid glands from ligated rats, prostaglandin E2 production is increased and, through cAMP accumulation, activates the inducible NOS isoform. The increment of nitric oxide production participates in the increase in basal amylase release.

Biomimetic oral targeted delivery of bindarit for immunotherapy of atherosclerosis.

Monocyte chemoattractant protein-1 (MCP-1) plays an important role in the development of atherosclerosis. However, the application of bindarit (a specific synthetic inhibitor of MCP-1) in atherosclerosis has not been confirmed due to the non-specific distribution profile in vivo. Herein, based on the recruitment of monocytes into atherosclerotic plaques, we successfully delivered bindarit into the interior of atherosclerotic plaques through a yeast-derived microcapsule (YC) mediated biomimetic approach. In this biomimetic approach, bindarit was firstly assembled with polyethyleneimine to form the positively charged nanoparticles (BIN/PEI NPs) via multiple intermolecular forces, and then the obtained BIN/PEI NPs were packed into YCs by electrostatic force-mediated spontaneous deposition. Through an oral adsorption routine similar to yeasts, bindarit loaded YCs (BIN/YCs) were distributed into peripheral blood monocytes after oral administration, and then their targeted delivery to atherosclerotic plaques was successfully performed through monocyte transportation. Correspondingly, oral delivery of bindarit loaded YCs afforded notably potentiated efficacies for inhibiting the MCP-1 and further reducing the recruitment of monocytes into atherosclerotic plaques, and thus presented a good efficacy in preventing the formation of atherosclerotic plaques. These results demonstrated that a 'Trojan horse'-like YC mediated nanomedicine delivery strategy is expected to realize the application of certain potential anti-inflammatory drugs in the treatment of atherosclerosis and is of great significance for the development of novel strategies for atherosclerosis treatment.

Clinicopathological and immunohistochemical evaluation of lonidamine-entrapped lipid-polymer hybrid nanoparticles in treatment of benign prostatic hyperplasia: An experimental rat model.

Benign prostatic hyperplasia (BPH) is a progressive proliferative disease, the incidence of which is constantly increasing due to aging of population. In this research, a hexokinase-II enzyme inhibiting agent, lonidamine - the use of which is limited in BPH treatment due to high hepatic toxicity observed after three months of treatment - was selected as an active agent, based on its mechanism of action in treating BPH. The aim of this study was to evaluate in vivo therapeutic efficacy and hepatic toxicity of lipid-polymer hybrid nanoparticles of lonidamine in a rat BPH model created in rat prostates. After local injections of hybrid nanoparticles of lonidamine were administered to the rat prostates, hyperplasic structures of prostates were evaluated in terms of prostatic index values, immunohistochemical evaluations, and histopathological findings. Liver blood enzyme values were also determined to specify hepatic toxicity. Apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) reaction and histopathological methods to determine intravital degenerative destruction in liver. Through this study, lonidamine-loaded hybrid nanoparticles were found to reduce the hepatic toxicity and increase therapeutic efficiency of lonidamine. Therefore, lonidamine-entrapped hybrid nanoparticles may provide a promising, and very safe, drug delivery strategy in the treatment of BPH.

Coadministration of chemotherapy and PI3K/Akt pathway treatment with multistage acidity/CathB enzyme-responsive nanocarriers for inhibiting the metastasis of breast cancer.

As the principal reason for the inducement of high mortality, tumor metastasis is regulated by different pathways owing to its complexity and multistep process. In order to inhibit the proliferation and metastasis of human breast cancer simultaneously, controlling the codelivery of chemotherapeutics and pathway inhibitors precisely has been considered as a high-potential strategy to accurately eliminate tumor metastasis. In this study, polymer PLGA-p-PEI-DA was synthesised and automatically assembled into a cascade "trinity" response drug delivery system, i.e., PPP-DA/NPs (PLGA: poly(lactin-co-glycolic acid), PEI: polyethyleneimine, DA: 2,3-dimethylmaleic anhydride). In the tumor microenvironment, PPP-DA/NPs could remove the outer DA molecules via the pH-sensitive hydrolysis of ß-carboxylic amide bonded with DA and PEI. Then, PPP-DA/NPs were broken up owing to the enzymatically cleavable GFLGF (Gly-Phe-Leu-Gly-Phe) linker. The structure of the polymer and the properties of PPP-DA/NPs were evaluated in detail. Moreover, studies on the antitumor metastasis efficiency and antitumor mechanism of PPP-DA/NPs were carried out in detail. As demonstrated in this study, PPP-DA/NPs could reverse the potential in pH 6.8 PBS and showed elevated cellular uptake efficiency. Moreover, PPP-DA/NPs exhibited strong antitumor metastasis ability in vitro and in vivo. The tumor inhibiting rate (TIR) of PPP-DA/NPs (68.4%) was significantly higher than that of docetaxel (DTX) (5.9%). The antitumor mechanistic studies confirmed that PPP-DA/NPs could down-regulate the expressions of Akt, MMP-9 and pro-caspase-3/9 protein, as indicated by western blot analysis. This multifunctional drug delivery system (DDS) is highly selective and effective in inhibiting tumor metastasis, which shows a great potential in inventing smart nanocarriers for targeted tumor-metastasis therapy.

Second-Generation SYK Inhibitor Entospletinib Ameliorates Fully Established Inflammation and Bone Destruction in the Cherubism Mouse Model.

Cherubism is a craniofacial disorder characterized by maxillary and mandibular bone destruction. Gain-of-function mutations in the SH3-domain binding protein 2 (SH3BP2) are responsible for the excessive bone resorption caused by fibrous inflammatory lesions. A homozygous knock-in (KI) mouse model for cherubism (Sh3bp2KI/KI ) develops autoinflammation resulting in systemic bone destruction. Although administration of the TNF-α blocker etanercept to neonatal Sh3bp2KI/KI mice prevented the disease onset, this therapy was not effective for adult Sh3bp2KI/KI mice or human cherubism patients who already had lesions. Because genetic ablation of spleen tyrosine kinase (SYK) in myeloid cells rescues Sh3bp2KI/KI mice from inflammation, we examined whether SYK inhibitor administration can improve fully developed cherubism symptoms in adult Sh3bp2KI/KI mice. Entospletinib (GS-9973) was intraperitoneally injected into 10-week-old Sh3bp2KI/KI mice every day for 6 weeks. Treatment with GS-9973 improved facial swelling and histomorphometric analysis of lung and liver tissue showed that GS-9973 administration significantly reduced inflammatory infiltrates associated with decreased levels of serum TNF-α. Micro-computed tomography (µCT) analysis showed that GS-9973 treatment reduced bone erosion in mandibles, calvariae, and ankle and elbow joints of Sh3bp2KI/KI mice compared to Sh3bp2KI/KI mice treated with dimethyl sulfoxide (DMSO). Taken together, the results demonstrate that administration of the SYK inhibitor ameliorates an already established cherubism phenotype in mice, suggesting that pharmacological inhibition of SYK may be a treatment option for cherubism patients with active disease progression. © 2018 American Society for Bone and Mineral Research.

[Therapeutic effect of enhancer of Zeste homolog 2 inhibitor GSK343 on periodontitis by regulating macrophage differentiation].

OBJECTIVE: To explore the therapeutic effect of enhancer of Zeste homolog 2 (EZH2) inhibitor GSK343 on periodontitis by regulating microphage differentiation. METHODS: Macrophage RAW264.7 cells were divided into the blank (A group), control (B group), lipopolysaccharide (LPS) stimulation (C group), and LPS+GSK343 (D group) groups. Phenotype transformations was determined through Western blot analysis and enzyme-linked immunosorbent assay by detecting the differentiation of phenotypic biological markers, including tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS), interleukin-10 (IL-10), and Arginase-1 (Arg-1). Metergasis was identified by performing a phagocytosis test on Escherichia coli (E. coli). RESULTS: Macrophage RAW264.7 cells produced classical phenotypic biomarkers (M1) TNF-α and iNOS under LPS stimulation. The expression levels of IL-10 and Arg-1 increased after adding GSK343 into the culture medium. GSK343 also induced the conversion of M1 macrophages into M2 macrophages. Macrophage RAW264.7 cells exerted a phagocytic effect on E. coli, and this effect was enhanced after adding LPS into the culture medium. GSK343 regulated the macrophage RAW264.7 phagocytosis of E. coli. CONCLUSIONS: GSK343 possibly participates in the regulation of macrophage differentiation and, consequently, in the latent treatment of periodontitis.

Ras Signaling Regulates Stem Cells and Amelogenesis in the Mouse Incisor.

The role of Ras signaling during tooth development is poorly understood. Ras proteins-which are activated by many upstream pathways, including receptor tyrosine kinase cascades-signal through multiple effectors, such as the mitogen-activated protein kinase (MAPK) and PI3K pathways. Here, we utilized the mouse incisor as a model to study how the MAPK and PI3K pathways regulate dental epithelial stem cells and amelogenesis. The rodent incisor-which grows continuously throughout the life of the animal due to the presence of epithelial and mesenchymal stem cells-provides a model for the study of ectodermal organ renewal and regeneration. Utilizing models of Ras dysregulation as well as inhibitors of the MAPK and PI3K pathways, we found that MAPK and PI3K regulate dental epithelial stem cell activity, transit-amplifying cell proliferation, and enamel formation in the mouse incisor.

OCLI-023, a Novel Pyrimidine Compound, Suppresses Osteoclastogenesis In Vitro and Alveolar Bone Resorption In Vivo.

An abnormal increase in osteoclast differentiation and activation results in various bone-resorptive diseases, including periodontitis, rheumatoid arthritis, and osteoporosis. Chemical compounds containing pyrimidine ring have been shown to regulate a variety of biological processes. Therefore, in order to identify an antiresorptive agent, we synthesized a series of pyrimidine ring-containing chemical compounds, and found that OCLI-023 suppressed the differentiation and activation of osteoclasts in vitro. OCLI-023 directly inhibited receptor activator of nuclear factor-κB ligand (RANKL)-induced differentiation of bone marrow macrophages into osteoclasts, without a cytotoxic response. OCLI-023 also downregulated the RANKL-induced mRNA expression of osteoclast markers as well as inhibited the formation of actin rings and resorption pits. OCLI-023 attenuated the RANKL-induced activation of c-Jun N-terminal kinase and nuclear factor kappa-light-chain-enhancer of activated B cell signaling pathways. In a mouse model of periodontitis, ligature induced an increase of distance between cementoenamel junction (CEJ) and alveolar bone crest (ABC) in the second molar, and OCLI-023 significantly reduced it. Histological analysis showed ligature-induced increase of osteoclast numbers was also significantly reduced by OCLI-023. These data demonstrated the inhibitory effect of OCLI-023 on osteoclast differentiation and activity of osteoclasts in vitro, as well as on ligature-induced bone loss in vivo, and OCLI-023 can be proposed as a novel anti-resorptive compound.

In vivo synergistic antitumor effect and safety of siRNA and lonidamine dual-loaded hierarchical targeted nanoparticles.

Based on development of nano-delivery system, co-delivery of chemotherapeutic drug and small interfering RNA (siRNA) has exerted a promising advantage in cancer therapy. In this work, the superiority of synergistic therapy and safety of the hierarchical targeted co-delivery system loaded with siRNA and lonidamine (LND) were evaluated. The in vivo tumor accumulation ability and cancer growth inhibition effect of the polymer-blend nanocarriers were evaluated by a H22 subcutaneous sarcoma model. Moreover, hematoxylin and eosin (H&E) staining and transferase-mediated dUTP nick end-labeling (TUNEL) staining of tumor sections from each group were compared to assess the therapeutic efficacy. The dual-loaded nanocarriers had better tumor accumulation ability, remarkably inhibited growth of solid tumor in a synergistic manner, even significantly decreased hepatotoxicity of LND, and had good in vivo biocompatibility whereas LND alone showed serious hepatotoxicity. We believed that the dual-loaded hierarchical targeted delivery system with high effectiveness and biocompatibility would provide a promising approach for cancer combination therapy.

Dual-pH responsive micelle platform for co-delivery of axitinib and doxorubicin.

While the complicated pathogenesis of cancer results in limited therapeutic efficacy of current mono-drug treatment, combination therapy by multiple drugs is becoming increasingly attractive due to the decreased side effects and synergistic anti-cancer activities. The recently emerging modality is the co-delivery of traditional chemotherapeutics and anti-angiogenesis agents. Although nanocarriers are frequently utilized for the co-delivery of different drugs, there are still concerns regarding their implementations. Most of the nanocarriers cannot release drugs separately into their different targeted sites of action. Therefore, we have developed a micellar platform for the co-delivery of an antiangiogenesis agent, axitinib (AXI) and a DNA intercalator, doxorubicin (DOX). Our results showed that this cross-linked micelle (DA-CM) could release AXI and DOX in tumor extracellular environment and intracellular lysosome compartments, respectively, in response to the dual pH stimulus. Notably, DA-CM exhibited remarkably improved tumor accumulation, cell internalization, tumor spheroids penetration and cytotoxicity. Ultimately, DA-CM reduced the number of immature vessels within xenograft tumors, demonstrating an effective antiangiogenesis effect. Meanwhile, they inhibited tumor growth by 88%. Our co-delivery micellar system with the dual-pH responsive feature might hold great promises for the combinatory cancer therapy.

Coupling of ascorbate and nitric oxide dynamics in vivo in the rat hippocampus upon glutamatergic neuronal stimulation: a novel functional interplay.

BACKGROUND: Ascorbate and neuronal-derived nitric oxide (NO) play regulatory roles in the brain that tare dependent on their compartmentalization and diffusion. Glutamatergic activation triggers both ascorbate fluxes toward extracellular medium and NO production. The information on the profiles of change in time and space upon glutamatergic activation is scarce and yet this knowledge is important for the understanding of ascorbate and NO functions in vivo, in particular in the case of a coupled interaction between both dynamics. HYPOTHESIS: NO produced upon NMDA receptor activation is a modulator of ascorbate release to the extracellular space. METHODS: In this work, carbon fiber microelectrodes for simultaneous measurements of these substances in the hippocampus were used to collect information about ascorbate and NO dynamic profiles in real time. RESULTS: Glutamate stimulation evoked transient ascorbate and NO signals with high degree of spatial and temporal correlation between them. Combined experiments encompassing direct stimulus with NO and inhibitors of glutamate uptake and nNOS provided additional evidence supporting the modulator role of NO in the release of ascorbate to the extracellular space. CONCLUSIONS: The coupling between NO and ascorbate upon glutamatergic activation points to a functional impact on the activities of both compounds and, although the precise molecular mechanism needs to be clarified, such a coupling lays the foundations for new regulatory mechanisms in the brain.

Effects of synthetic peptido-leukotrienes on bone resorption in vitro.

Peptido-leukotrienes are short-lived organic molecules known to have potent biological effects as mediators of inflammation, hypersensitivity and respiratory disorders. However, little is known concerning their effects on bone cells. We have shown previously that stromal cells isolated from a human giant cell tumor secrete 5-HETE (5-hydroxyeicosatetraenoic acid) and the peptido-leukotrienes, also known as the cysteinyl leukotrienes LTC4, LTD4, and LTE4. These eicosanoids were shown to stimulate the multinucleated giant cells obtained from these tumors to form resorption lacunae on sperm whale dentine. Here, we show that the peptido-leukotrienes also stimulate isolated avian osteoclast-like cells to form resorption lacunae and to increase their content of tartrate-resistant acid phosphatase. LTD4 increased 45Ca release from murine calvarial bone organ cultures, but not from fetal rat long bone cultures. Isolated avian osteoclast-like cells were chosen to perform receptor binding studies, as this population is the most homogeneous source of osteoclasts available. After the precursors had fused to form multinucleated cells, receptor binding assays were performed. Scatchard analysis of saturation binding data showed a single class of binding sites, with a dissociation constant (Kd) of 0.53 nM and a receptor density of 5,200 receptors per cell. Competition binding studies showed receptor specificity using a specific LTD4 receptor antagonist ZM 198,615. These data show that the peptido-leukotrienes activate highly enriched populations of isolated avian osteoclast-like cells, and also that specific LTD4 receptors are present in this cell population.