In vitro, ex vivo, and in vivo evaluation of the effect of saturated fat acid chain length on the transdermal behavior of ibuprofen-loaded microemulsions.

In this study, the effect of the saturated fatty acid (FA) chain length in the oil phase on the behavior of Ibuprofen (IBU)-loaded transdermal microemulsion (ME) was evaluated in vitro, ex vivo, and in vivo. Three oils classified as long (LFA), medium (MFA), and short (SFA) chain length oils, Cremophor RH40 (surfactant) and Transcutol P (cosurfactant) were selected after experimental optimization. The physicochemical properties of ME were characterized, including IBU solubility in excipients, pseudo-ternary phase diagram construction, particle size, zeta potential, viscosity, and stability. Permeation flux and residual amount of IBU ex vivo using Franz cell system occurred in the following order: MFA-based ME > LFA-based ME > SFA-based ME, which correlated well with the results of confocal scanning laser microscopy study and the in vivo retention study. The results of in vitro cytotoxicity study and skin irritation tests measured by differential scanning calorimetry were ranked in the following order: LFA-based ME > MFA-based ME > SFA-based ME. Moreover, MFA-based ME has the highest analgesic activity among all the treatment groups. MFA was found to be an optimal oil phase with appropriate FA chain length for IBU-loaded transdermal ME, which exhibited excellent physicochemical properties, low toxicity, and good permeability profile.

Pluronic mixed micelles overcoming methotrexate multidrug resistance: in vitro and in vivo evaluation.

A Pluronic polymeric mixed micelle delivery system was developed in this study by using Pluronic P105 and F127 block copolymers to encapsulate the antitumor compound, methotrexate (MTX). The MTX-loaded Pluronic P105/F127 mixed micelle exhibited the spherical shape with about 22 nm in diameter, high encapsulation efficiency (about 85%) and pH-dependent in vitro drug release. In this study, A-549 and KBv cell lines were selected as multidrug resistance tumor cell models, while H-460 and KB cell lines were chosen as sensitive tumor cells. The MTX-loaded Pluronic P105/F127 mixed micelle exhibited significant higher in vitro cytotoxicity in multidrug resistant tumor cells than that of control (MTX injection) mainly because of higher cellular uptake of MTX. The pharmacokinetic studies indicated that the Pluronic micelles significantly prolonged systemic circulation time of MTX compared to MTX injection. Moreover, a much stronger antitumor efficacy in KBv tumor xenografts nude mice was observed in the MTX-loaded Pluronic P105/F127 mixed micelle group, than MTX. Collectively, Pluronic P105/F127 mixed micelles could significantly enhance the antitumor activity of MTX and might be a promising drug delivery platform for multidrug resistance modulation.

Integrin-facilitated transcytosis for enhanced penetration of advanced gliomas by poly(trimethylene carbonate)-based nanoparticles encapsulating paclitaxel.

The treatment of cerebral tumor, especially advanced gliomas, represents one of the most formidable challenges in oncology. In this study, integrin-mediated poly(trimethylene carbonate)-based nanoparticulate system (c(RGDyK)-NP) was proposed as a delivery vehicle for enhancing drug penetration and chemotherapy of malignant gliomas. Following the recognition by integrin proteins on cell surface, c(RGDyK)-NP could be energy-dependently internalized by human U87MG glioma cells through a multiple endocytic pathway. The tumor penetration, homing specificity and anticancer efficacy of PTX-loaded c(RGDyK)-NP (c(RGDyK)-NP/PTX) were performed on the 3D glioma spheroids, the U87MG glioma cells and the intracranial glioma mice model, respectively. Compared with conventional nanoparticles (NP/PTX) and Taxol, c(RGDyK)-NP/PTX showed the strongest penetration and accumulation into 3D glioma spheroids, an obvious microtubule stabilization effect to U87MG glioma cells, a significant homing specificity to malignant glioma in vivo, and an extended median survival time in the intracranial glioma-bearing mice. Furthermore, preliminary in vivo subacute toxicity was also evaluated by measuring the histopathology, blood cell counts and clinical biochemistry parameters, and the results revealed no obvious subacute toxicity to hematological system, major organs or tissues were observed post successive intravenous injection of c(RGDyK)-NP. Therefore, our results suggested that cyclic RGD-conjugated PEG-PTMC nanoparticle could be a promising vehicle for enhancing the penetration and cxhemotherapy of high-grade malignant gliomas.

Pluronic P105/F127 mixed micelles for the delivery of docetaxel against Taxol-resistant non-small cell lung cancer: optimization and in vitro, in vivo evaluation.

The aim of this work was to establish a novel polymeric mixed micelle composed of Pluronic P105 and F127 copolymers loaded with the poorly soluble antitumor drug docetaxel (DTX) against Taxol-resistant non-small cell lung cancer. A central composite design was utilized to optimize the preparation process, helping to improve drug solubilization efficiency and micelle stability. Prepared by a thin-film hydration method, the average size of the optimized mixed micelle was 23 nm, with a 92.40% encapsulation ratio and a 1.81% drug-loading efficiency. The optimized formulation showed high storage stability in lyophilized form, with 95.7% of the drug content remaining after 6 months' storage at 4°C. The in vitro cytotoxicity assay showed that the IC50 values for Taxotere(®) and mixed micelles were similar for A549, while on A549/Taxol cell lines, DTX-loaded P105/F127 mixed micelles showed a superior hypersensitizing effect; their IC50 value (0.059 µg/mL) was greatly reduced compared to those of Taxotere injections (0.593 µg/mL). The in vivo pharmacokinetic study showed that the mixed-micelle formulation achieved a 1.85-fold longer mean residence time in circulation and a 3.82-fold larger area under the plasma concentration-time curve than Taxotere. In addition, therapeutic improvement of mixed micelles in vivo against A549/Taxol was obtained. The tumor inhibition rate of the micelles was 69.05%, versus 34.43% for Taxotere (P < 0.01). Therefore, it could be concluded from the results that DTX-loaded P105/F127 mixed micelles might serve as a potential antitumor drug delivery system to overcome multidrug resistance in lung cancer.

Solid tumor penetration by integrin-mediated pegylated poly(trimethylene carbonate) nanoparticles loaded with paclitaxel.

Limited penetration of antineoplastic agents is one of the contributing factors for chemotherapy failure of many solid tumors. In order to enhance drug penetration into solid cancer, especially, into the avascular regions inside tumors, we proposed cyclic RGD peptide functionalized PEGylated poly(trimethylene carbonate) nanoparticles (c(RGDyK)-NP). By integrin-mediated transcytosis and enhanced drug permeation, c(RGDyK)-NP could access the neoplastic cells distant from blood vessels, and consequently, avoiding the capability of cancer regeneration from these tumor cells. In the present study, the solid tumor penetration, homing specificity and anticancer efficacy were evaluated both on the ex vivo 3D tumor spheroids and on the subcutaneous xenograft mice model. In comparison with conventional nanoparticles (NP/PTX) and Taxol, c(RGDyK)-NP/PTX showed the strongest penetration and accumulation into 3D tumor spheroids, a marked tumor-homing specificity in vivo and the greatest tumor growth inhibitory effect in vitro and in vivo. Histochemistry analysis revealed that no obvious histopathological abnormalities or lesions were observed in major organs after intravenous administration with the treatment doses. In conclusion, cyclic RGD peptide-conjugated PEG-PTMC nanoparticle could facilitate drug penetration and accumulation in tumor tissues and may be a promising vehicle for enhancing the chemotherapy of solid cancers.

Anti-glioblastoma efficacy and safety of paclitaxel-loading Angiopep-conjugated dual targeting PEG-PCL nanoparticles.

Therapeutic effect of glioma is often limited due to low permeability of delivery systems across the Blood-Brain Barrier (BBB) and poor penetration into the tumor tissue. In order to overcome the two barriers, we proposed Angiopep-conjugated PEG-PCL nanoparticles (ANG-PEG-NP) as a dual targeting drug delivery system for glioma treatment basing on low density lipoprotein receptor related protein (LRP) receptor not only over-expressed on BBB but also on glioma cells. This system could transport across BBB through LRP-mediated transcytosis and then targeted glioma via LRP-mediated endocytosis. In this study, we evaluated the preliminary availability and safety of ANG-PEG-NP for glioma treatment. The penetration, distribution, and accumulation into 3D glioma spheroid and in vivo glioma region of ANG-PEG-NP were obviously higher than that of plain PEG-PCL nanoparticles (PEG-NP). The anti-glioblastoma efficacy of paclitaxel (PTX) loading ANG-PEG-NP was significantly enhanced as compared to that of Taxol and PEG-NP. Preliminary safety results showed that no acute toxicity to hematological system, liver, kidney and brain tissue was observed after intravenous administration with a dose of 100 mg/kg blank ANG-PEG-NP per day for a week. Results indicate that Angiopep-conjugated dual targeting PEG-PCL nanoparticle is a potential brain targeting drug delivery system for glioma treatment.

The brain targeting mechanism of Angiopep-conjugated poly(ethylene glycol)-co-poly(ε-caprolactone) nanoparticles.

In order to evaluate the potential and mechanism of Angiopep-conjugated poly(ethylene glycol)-co-poly(ε-caprolactone)nanoparticles (ANG-PEG-NP) as brain targeting drug delivery system, Rhodamine B isothiocyanate (RBITC) was used as a fluorescent probe molecule to label ANG-PEG-NP through covalent bonding. The brain transcytosis across the blood-brain barrier (BBB) and brain delivery in mice of RBITC labeled ANG-PEG-NP were investigated in this paper. Results showed that ANG-PEG-NP enhanced significantly the uptake by BCECs compared with that of PEG-NP through caveolae- and clathrin-mediated endocytosis, involving a time-dependent, concentration-dependent and energy-dependent mode. The transport of ANG-PEG-NP across the in vitro BBB model was significantly increased than that of PEG-NP. After injection a dose of 100 mg/kg RBITC labeled ANG-PEG-NP or PEG-NP in mouse caudal vein, the brain coronal section showed a higher accumulation of ANG-PEG-NP in the cortical layer, lateral ventricle, third ventricles and hippocampus than that of PEG-NP. By using an excess of free LRP ligand (Angiopep-2 and/or Aprotinin) as a specific receptor inhibitor, it was evidenced that the uptake by BCECs in vitro, transport across in vitro BBB model and penetration into brain tissue in vivo of RBITC labeled ANG-PEG-NP could be inhibited significantly, which demonstrated the brain targeting mechanism of Angiopep-conjugated poly(ethylene glycol)-co-poly(ε-caprolactone)nanoparticles might be a LRP receptor mediated transcytosis process. Understanding these issues is important for the future development of ANG-PEG-NP as a brain targeting drug delivery system for neurodegenerative disorders including glioma and Alzheimer's disease.

Self-assembled carboxymethyl poly (L-histidine) coated poly (ß-amino ester)/DNA complexes for gene transfection.

Biomaterials coated polymer/DNA complexes are developed as an efficient non-viral gene delivery system. It is able to circumvent the changes of various biophysical properties of the biomaterials and the corresponding polymer/DNA nanoparticles with covalent linkage. In the present study, we introduced pH-sensitive carboxymethyl poly (l-histidine) (CM-PLH) and poly (ß-amino ester) (PbAE) as functional biomaterials to form CM-PLH/PbAE/DNA core-shell ternary complexes system based on electrostatically adsorbed coatings for gene efficient delivery and transfection. The preparation of the complexes was performed self-assembly in 25 mm sodium acetate buffer solution at pH 5.2. The complexes kept stable nano-size, behaving good condensation capacity and low toxicity, even provided a higher transfection efficiency than the binary complexes (PbAE/DNA without CM-PLH) and transfected up to (89.6 ± 4.45) % in HEK293 and (57.1 ± 2.10) % in B16-F10 in vitro. The ternary complexes significantly enhanced their cellular uptake and endosomal escape which were proved by the results that the complexes could evade the endosomal lumen and localize in the nucleus of treated cells visualized under Fluorescence Confocal Microscopy (FCM). The aforementioned results indicated that CM-PLH with pH-sensitive imidazole groups played an important role in enhancing the endosomal escape and transfection efficiency. The in vivo gene transfection confirmed that the ternary complexes with pGL3-promoter as led to effectively deposit at the tumor site by the EPR effect and shown 4 fold higher luciferase expression in B16-F10 tumor than the binary complexes. Consequently, CM-PLH/PbAE/DNA ternary complexes system exhibited significant improvements in transfection efficiency in comparison with non-coated PbAE/DNA both in vitro and in vivo, highlighting their functional prospect. Our approach and the gene delivery system fabrication could potentially be useful for effective gene delivery and therapies to targeted cells.

Self-aggregated pegylated poly (trimethylene carbonate) nanoparticles decorated with c(RGDyK) peptide for targeted paclitaxel delivery to integrin-rich tumors.

Cyclic RGD peptide-decorated polymeric micellar-like nanoparticles (MNP) based on PEGylated poly (trimethylene carbonate) (PEG-PTMC) were prepared for active targeting to integrin-rich cancer cells. An amphiphilic diblock copolymer, α-carboxyl poly (ethylene glycol)-poly (trimethylene carbonate) (HOOC-PEG-PTMC), was synthesized by ring-opening polymerization. The c(RGDyK) ligand, a cyclic RGD peptide that can bind to the integrin proteins predominantly expressed on the surface of tumor cells with high affinity and specificity, was conjugated to the NHS-Activated PEG terminus of the copolymer. The c(RGDyK)-functionalized PEG-PTMC micellar nanoparticles encapsulating PTX (c(RGDyK)-MNP/PTX) was fabricated by the emulsion/solvent evaporation technique and characterized in terms of morphology, size and zeta potential. Cellular uptake of c(RGDyK)-MNP/PTX was found to be higher than that of MNP/PTX due to the integrin protein-mediated endocytosis effect. In vitro cytotoxicity, cell apoptosis and cell cycle arrest studies also revealed that c(RGDyK)-MNP/PTX was more potent than those of MNP/PTX and Taxol. Pharmacokinetic study in rats demonstrated that the polymeric micellar nanoparticles significantly enhanced the bioavailability of PTX than Taxol. In vivo multispectral fluorescent imaging indicated that c(RGDyK)-MNP/PTX had high specificity and efficiency in tumor active targeting. Therefore, the results demonstrated that c(RGDyK)-decorated PEG-PTMC MNP developed in this study could be a potential vehicle for delivering hydrophobic chemotherapeutic agents to integrin-rich tumors.

PEGylated poly(trimethylene carbonate) nanoparticles loaded with paclitaxel for the treatment of advanced glioma: in vitro and in vivo evaluation.

The aim of this study was to investigate the antitumor effect of paclitaxel (PTX)-loaded poly(ethylene glycol)-poly(trimethylene carbonate) (MPEG-PTMC) nanoparticles (NP) against gioblastoma multiforme (GMB). PTX-loaded NP (NP/PTX) were prepared with synthesized MPEG-PTMC by the emulsion/solvent evaporation technique. In vitro physiochemical characterization of those NP/PTX showed satisfactory encapsulation efficiency and loading capacity and size distribution. Cytotoxicity assay revealed that encapsulation in nanoparticles did not compromise the antitumor efficacy of PTX against U87MG cells. Pharmacokinetic study in rats demonstrated that the polymer micellar nanoparticles significantly enhanced the bioavailability of PTX than Taxol. In intracranial xenograft tumor-bearing mice, the accumulation of nanoparticles in tumor tissues increased distinctly after 12 h post i.v. More importantly, in vivo anti-tumor effect exhibited the median survival time of NP/PTX treated mice (27 days) was significantly longer than those of mice treated with Taxol (24 days), physiological saline (21 days) and blank MPEG-PTMC NP (21 days). Therefore, our results suggested that PTX-loaded MPEG-PTMC nanoparticles significantly enhanced the anti-glioblastoma activity of PTX and may be a potential vehicle in the treatment of high-grade glioma.

Angiopep-conjugated poly(ethylene glycol)-co-poly(ε-caprolactone) nanoparticles as dual-targeting drug delivery system for brain glioma.

Dual-targeting nanoparticle drug delivery system was developed by conjugating Angiopep with PEG-PCL nanoparticles (ANG-NP) through bifunctional PEG to overcome the limitations of low transport of chemotherapeutics across the Blood-brain barrier (BBB) and poor penetration into tumor tissue. ANG-NP can target the low-density lipoprotein receptor-related protein (LRP) which is over-expressed on the BBB and glioma cells. Compared with non-targeting nanoparticles, a significantly higher amount of rhodamine isothiocyanate-labeled dual-targeting nanoparticles were endocytosed by U87 MG cells. The antiproliferative and cell apoptosis assay of paclitaxel-loaded ANG-NP (ANG-NP-PTX) demonstrated that ANG-NP-PTX resulted in enhanced inhibitory effects to U87 MG glioma cells. The transport ratios across the BBB model in vitro were significantly increased and the cell viability of U87 MG glioma cells after crossing the BBB was obviously decreased by ANG-NP-PTX. Enhanced accumulation of ANG-NP in the glioma bed and infiltrating margin of intracranial U87 MG glioma tumor-bearing in vivo model were observed by real time fluorescence image. In conclusion, Angiopep-conjugated PEG-PCL nanoparticles were prospective in dual-targeting drug delivery system for targeting therapy of brain glioma.

Enhanced anti-glioblastoma efficacy by PTX-loaded PEGylated poly(ɛ-caprolactone) nanoparticles: In vitro and in vivo evaluation.

The aim of this work was to investigate the anti-tumor effect of paclitaxel (PTX)-loaded methoxy poly(ethylene glycol)-poly(ɛ-caprolactone) nanoparticles (MPEG-NP/PTX) against glioblastoma multiforme (GBM). MPEG-NP/PTX was prepared by the emulsion and evaporation technique with particle size of 72.5±2.2nm and did not change remarkably during the period of 21-day storage at 4°C. The drug-loading coefficient and encapsulation ratio of optimized formulation were 8.2±0.6% and 90.4±2.3%, respectively. The in vitro release behavior exhibits a biphase release manner and was affected by PEG segment. In vitro cytotoxicity was assessed using C6 cell lines and was compared to Taxol and PTX-loaded poly(ɛ-caprolactone) conventional nanoparticles (NP/PTX). Cell viability assay against C6 cells exhibited higher or at least comparable cytotoxicity than that of Taxol and NP/PTX. More importantly, in vivo real-time fluorescence imaging analysis in intracranial C6 glioblastoma bearing mice showed that the methoxy poly(ethylene glycol)-poly(ɛ-caprolactone) nanoparticles (MPEG-NP) displayed much stronger fluorescence signal and 3-fold larger Area-Under-Curve (AUC) than poly(ɛ-caprolactone) conventional nanoparticles (NP) in tumor-bearing brain. Furthermore, in vivo anti-glioblastoma effect exhibited the mean survive time of MPEG-NP/PTX (28 days) was much longer than those of Taxol injection (20 days) and NP/PTX (23 days). Therefore, MPEGylated poly(ɛ-caprolactone) nanoparticles significantly enhanced the anti-glioblastoma activity of PTX and might be considered a promising drug delivery system against advanced glioblastoma.