The drug encapsulation efficiency, in vitro drug release, cellular uptake and cytotoxicity of paclitaxel-loaded poly(lactide)-tocopheryl polyethylene glycol succinate nanoparticles

Paclitaxel is one of the most effective antineoplastic drugs. Its current clinical administration is formulated in Cremophor EL, which causes serious side effects. Nanoparticle (NP) technology may provide a solution for such poisonous adjuvant problems and promote a sustained chemotherapy, in which biodegradable polymers play a key role. Our group has successfully synthesized novel poly(lactide)-tocopheryl polyethylene glycol succinate (TPGS) (PLA-TPGS) copolymers of desired hydrophobic-hydrophilic balance for NP formulation of anticancer drugs. The present work is focused on effects of the PLA:TPGS composition ratio on drug encapsulation efficiency, in vitro drug release, in vitro cellular uptake and viability of the PLA-TPGS NP formulation of paclitaxel. The PLA-TPGS copolymers of various PLA:TPGS ratios were synthesized by the ring-opening polymerization method and characterized by GPC and (1)H NMR for their molecular structure. Paclitaxel-loaded PLA-TPGS NPs were prepared by a modified solvent extraction/evaporation method and characterized by laser light scattering for size and size distribution, scanning electron microscopy for surface morphology and zeta potential for surface charge. High performance liquid chromatography was used to measure the drug encapsulation efficiency and in vitro drug release profile. Cancer cell lines HT-29 and Caco-2 were used to image and measure the cellular uptake of fluorescent PLA-TPGS NPs. Cancer cell viability of the drug-loaded PLA-TPGS was measured by MTT assay. It was found that the PLA:TPGS composition ratio has little effects on the particle size and size distribution. However, the PLA-TPGS NPs of 89:11 PLA:TPGS ratio achieved the best effects on the drug encapsulation efficiency, the cellular uptake and the cancer cell mortality of the drug-loaded PLA-TPGS NPs. This research was also carried out in close comparison with the drug-loaded PLGA NPs.     

Folate-decorated poly(lactide-co-glycolide)-vitamin E TPGS nanoparticles for targeted drug delivery

Doxorubicin-loaded nanoparticles (NPs) of vitamin E TPGS-folate (TPGS-FOL) conjugate and doxorubicin-poly(lactide-co-glycolide)-vitamin E TPGS (DOX-PLGA-TPGS) conjugate were prepared by the solvent extraction/evaporation method for targeted chemotherapy of folate-receptor rich tumors. X-ray photoelectron spectroscopy demonstrated that folate was distributed on the NP surface while the drug molecules were entrapped in the NP matrix. The NPs were found of approximately 350nm diameter and exhibited a biphasic pattern of in vitro drug release. The cell uptake of the fluorescent NPs and the cell viability of the drug formulated in the NPs were quantitatively investigated, which were found dependent on the content of targeting TPGS-FOL conjugate. The NPs of 50% TPGS-FOL showed cellular uptake by MCF-7 cells 1.5 times higher and by C6 cells 1.7 times higher than the NPs with no TPGS-FOL component after 30min incubation. The MCF-7 cell viability was found decreased significantly from 50.8% for the drug-loaded NPs of no TPGS-FOL to 8.2% for those of 50% TPGS-FOL after incubation at 100microug concentration at 37 degrees C. The latter NPs also exhibited much lower IC(50) value than the DOX after 24h incubation, i.e., 19.4 vs. 43.7micror MCF-7 cells and 3.3 vs. >100micror C6 cells.     

Radiosensitization of paclitaxel, etanidazole and paclitaxel+etanidazole nanoparticles on hypoxic human tumor cells in vitro

Paclitaxel and etanidazole are hypoxic radiosensitizers that exhibit cytotoxic action at different mechanisms. The poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles containing paclitaxel, etanidazole and paclitaxel+etanidazole were prepared by o/w and w/o/w emulsification-solvent evaporation method. The morphology of the nanoparticles was investigated by scanning electron microscope (SEM). The drug encapsulation efficiency (EE) and release profile in vitro were measured by high-performance liquid chromatography (HPLC). The cellular uptake of nanoparticles for the human breast carcinoma cells (MCF-7) and the human carcinoma cervicis cells (HeLa) was evaluated by transmission electronic microscopy and fluorescence microscopy. Cell viability was determined by the ability of single cell to form colonies in vitro. The prepared nanoparticles were spherical shape with size between 80 and 150 nm. The EE was higher for paclitaxel and lower for etanidazole. The drug release was controlled over time. The cellular uptake of nanoparticles was observed. Co-culture of the two tumor cell lines with drug-loaded nanoparticles demonstrated that released drug effectively sensitized hypoxic tumor cells to radiation. The radiosensitization of paclitaxel+etanidazole nanoparticles was more significant than that of single drug-loaded nanoparticles.     

In vitro and in vivo evaluation of methoxy polyethylene glycol-polylactide (MPEG-PLA) nanoparticles for small-molecule drug chemotherapy

Methoxy polyethylene glycol-polylactide (MPEG-PLA) nanoparticles (NPs) were prepared by the nanoprecipitation method with particle size of 140+/-21nm in diameter and drug encapsulation efficiency of 87.6+/-3.1%. In vitro cytotoxicity of the drug formulated in the NPs was investigated with MCF-7 cancer cells in close comparison with that of Taxol((R)). The in vitro cytotoxicity with MCF-7 cells showed that the NP formulation could be 33.3, 10.7, 7.7 times more effective than Taxol((R)) after 24, 48, 72h culture at the same drug concentration of 1microg/ml. Confocal laser scanning microscopy (CLSM) visualized cellular internalization of the coumarin 6-loaded MPEG-PLA NPs. The in vitro results were further confirmed by the in vivo pharmacokinetic analysis with SD rats. The total area-under-the-curve (AUC(0-infinity)), which determines the therapeutic effects of a dose, was found to be 29,600+/-1,690ng-h/ml for the NP formulation, which is 3.09 times of 9,570+/-1,480ng-h/l for Taxol((R)) with 10mg/kg dose i.v. injection. The half-life (t(1/2)) of the drug formulated in the NPs was found to be 18.80+/-3.14h, which is 2.75 times of 6.84+/-1.39h for Taxol((R)). The distribution volume at steady state for the drug loaded in the NPs was 7.21+/-2.17l/kg, which was 2.93 times of 2.46+/-1.41l/kg for Taxol((R)). Our proof-of-concept in vitro and in vivo valuation shows that our MPEG-PLA NP formulation could have great advantages versus the original drug in small-molecule drug chemotherapy as well as in various applications in nanomedicine.     

Docetaxel-loaded nanoparticles based on star-shaped mannitol-core PLGA-TPGS diblock copolymer for breast cancer therapy

A star-shaped biodegradable polymer, mannitol-core poly(d,l-lactide-co-glycolide)-d-α-tocopheryl polyethylene glycol 1000 succinate (M-PLGA-TPGS), was synthesized in order to provide a novel nanoformulation for breast cancer chemotherapy. This novel copolymer was prepared by a core-first approach via three stages of chemical reaction, and was characterized by nuclear magnetic resonance, gel permeation chromatography and thermogravimetric analysis. The docetaxel-loaded M-PLGA-TPGS nanoparticles (NPs), prepared by a modified nanoprecipitation method, were observed to be near-spherical shape with narrow size distribution. Confocal laser scanning microscopy showed that the uptake level of M-PLGA-TPGS NPs was higher than that of PLGA NPs and PLGA-TPGS NPs in MCF-7 cells. A significantly higher level of cytotoxicity was achieved with docetaxel-loaded M-PLGA-TPGS NPs than with commercial Taxotere®, docetaxel-loaded PLGA-TPGS and PLGA NPs. Examination of the drug loading and encapsulation efficiency proved that star-shaped M-PLGA-TPGS could carry higher levels of drug than linear polymer. The in vivo experiment showed docetaxel-loaded M-PLGA-TPGS NPs to have the highest anti-tumor efficacy. In conclusion, the star-like M-PLGA-TPGS copolymer shows potential as a promising drug-loaded biomaterial that can be applied in developing novel nanoformulations for breast cancer therapy.     

Targeted delivery of paclitaxel using folate-decorated poly(lactide)-vitamin E TPGS nanoparticles

We synthesized nanoparticles (NPs) of the blend of two-component copolymers for targeted chemotherapy with paclitaxel used as model drug. One component is poly(lactide)-D-alpha-tocopheryl polyethylene glycol succinate (PLA-TPGS), which is of desired hydrophobic-lipophilic balance, and another is TPGS-COOH, which facilitates the folate conjugation for targeting. The nanoparticles of the two-copolymer blend at various component ratio were prepared by the solvent extraction/evaporation single emulsion method and then decorated by folate, which were characterized by laser light scattering (LLS) for particles' size and size distribution, zeta potential analyzer for surface charge, and X-ray photoelectron spectroscopy (XPS) for surface chemistry. The drug encapsulation efficiency (EE) and in vitro drug release were measured by high performance liquid chromatography (HPLC). The targeting effect was investigated in vitro by cancer cell uptake of coumarin-6-loaded NPs and further confirmed by cytotoxicity of cancer cells treated with the drug formulated in the NPs. We showed that the NP formulation has great advantages vs the pristine drug in achieving better therapeutic effect, which increased 8.68% for MCF-7 breast cancer cells, and that the folate-decoration can significantly promote targeted delivery of the drug into the corresponding cancer cells and thus enhance its therapeutic effect, which increased 24.4% for the NP formulation of 16.7% TPGS-COOH component and 31.1% for the NP formulation of 33.3% TPGS-COOH component after 24h treatment at the same 25 microg/ml paclitaxel concentration. The experiments on C6 glioma cells further confirmed these advantages.     

Synthesis,characterization, and in vitro evaluation of TRAIL-modified,cabazitaxel -loaded polymeric micelles for achieving synergistic anticancer therapy

Abstract

Combination therapy of two or more drugs has gradually become of outmost importance in cancer treatment. Cabazitaxel (CTX) is a taxoid drug and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of TNF superfamily. In this study, we prepared TRAIL-modified and CTX-loaded polymer micelle (TRAIL-M-CTX). This nanoparticle was self-assembled from biodegradable amphiphilic copolymers, monomethoxyl poly(ethylene glycol)–b-poly(DL-lactide) (mPEG-PLA) and COOH-PEG-PLA, via a nanoprecipitation method and were modified with the TRAIL protein, resulting in a particle size of 39.75 ± 0.17 nm in diameter and a drug encapsulation efficiency of 95.52 ± 1.69%. The successful coupling was confirmed by ¹H NMR, FTIR spectroscopy, and DLS article size measurement. Pharmacodynamic analysis in two human cancer cell lines with different TRAIL sensitivities showed that TRAIL-M-CTX has a significantly better anticancer efficacy than the individual CTX and TRAIL protein. Importantly, TRAIL-M-CTX showed synergistic effects against TRAIL-insensitive cells (MCF-7). A study of cellular uptake implied that the modified micelles were internalized into MCF-7 cells more effectively than unmodified micelles, owing to the coupled TRAIL protein. A cell cycle assay of MCF-7 cells revealed that TRAIL-M-CTX significantly increased the sub-G1 population compared with CTX or TRIAL, thus, facilitating cancer cell apoptosis. These results suggest that TRAIL-M-CTX micelles have potential as a cancer chemotherapy formulation.     

Nanoparticles of poly(lactide)-tocopheryl polyethylene glycol succinate (PLA-TPGS) copolymers for protein drug delivery

Nanoparticles (NPs) of poly(lactide)-tocopheryl polyethylene glycol succinate (PLA-TPGS) copolymers with various PLA:TPGS component ratios were prepared by the double emulsion technique for protein drug formulation with bovine serum albumin (BSA) as a model protein. Influence of the PLA:TPGS component ratio and the BSA loading level on the drug encapsulation efficiency (EE) and in vitro drug release behavior was investigated. The PLA-TPGS NPs achieved 16.7% protein drug loading and 75.6% EE, which exhibited a biphasic pattern of controlled protein release with higher initial burst for those NPs of more TPGS content. Furthermore, the released proteins retained good structural integrity for at least 35 days at 37 degrees C as indicated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and circular dichroism (CD) spectroscopy. Compared with other biodegradable polymeric NPs such as poly(D,L-lactide-co-glycolide) (PLGA) NPs, PLA-TPGS NPs could provide the encapsulated proteins a milder environment. Confocal laser scanning microscopy (CLSM) observation demonstrated the intracellular uptake of the PLA-TPGS NPs by NIH-3T3 fibroblast cells and Caco-2 cancer cells. This research suggests that PLA-TPGS NPs could be of great potential for clinical formulation of proteins and peptides.     

Dual docetaxel/superparamagnetic iron oxide loaded nanoparticles for both targeting magnetic resonance imaging and cancer therapy

Theragnostics polymer nanoparticles (NPs) loaded simultaneously with anticancer drug docetaxel (Dtxl) and superparamagnetic iron oxide (SPIO) nanocrystals were developed for both cancer therapy and ultrasensitive MRI. These multifunctional polymer vesicles were formed by carboxy-terminated poly(lactic-co-glycolic) acid using a single emulsion evaporation method. The active tumor-targeting single chain prostate stem cell antigen antibodies (scAb(PSCA)) were conjugated on the surface of polymer vesicles by using functional poly(ethylene glycol). The diameter of NPs was about 147 nm and the SPIO and drug encapsulation efficacy was 23% and 6.02%, respectively. Vibration simple magnetometer and X-ray diffraction proved that the superparamagnetic behavior of SPIO was not changed during NPs formation and modification. The NPs exhibited a triphasic drug release pattern in vitro over 30 days. Enhanced cellular uptake ability and antiproliferative effect of the targeted NPs in prostate cancer PC3 cell line by using the confocal laser scanning microscopy and cytotoxicity assay were observed. Moreover, the Prussian blue staining and the MRI assay in vitro demonstrated that the NPs have a high SPIO clustering effect. Therefore, these stable and tumor-targeting polymer NPs could be promising multifunctional vesicles for simultaneous targeting imaging, drug delivery and real time monitoring of therapeutic effect.     

Multifunctional poly(D,L-lactide-co-glycolide)/montmorillonite (PLGA/MMT) nanoparticles decorated by Trastuzumab for targeted chemotherapy of breast cancer

This paper continued our earlier work on the poly(D,L-lactide-co-glycolide)/montmorillonite nanoparticles (PLGA/MMT NPs), which were further decorated by human epidermal growth factor receptor-2 (HER2) antibody Trastuzumab for targeted breast cancer chemotherapy with paclitaxel as a model anticancer drug. Such a NP system is multifunctional, which formulates anticancer drugs with no harmful adjuvant, reduces the side effects of the formulated anticancer drug, promotes synergistic therapeutic effects, and achieves targeted delivery of the therapy. The paclitaxel-loaded PLGA/MMT NPs were prepared by a modified solvent extraction/evaporation technique, which were then decorated with Trastuzumab. The effects of the surface decoration on particle size and size distribution, surface morphology, drug encapsulation efficiency, as well as the drug release kinetics, were investigated. The NP formulation exhibited a biphasic drug release with a moderate initial burst followed by a sustained release profile. The surface decoration speeded the drug release. Surface chemistry analysis was conducted by X-ray photoelectron spectroscopy (XPS), which confirmed the presence of Trastuzumab on the NP surface. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed the stability of the antibody in the NP preparation process. Internalization of the coumarin-6-loaded PLGA/MMT NPs with or without the antibody decoration by both of Caco-2 colon adeno carcinoma cells and SK-BR-3 breast cancer cells was visualized by confocal laser scanning microscopy and quantitatively analyzed, which shows that the antibody decoration achieved significantly higher cellular uptake of the NPs. The results of in vitro cytotoxicity experiment on SK-BR-3 cells further proved the targeting effects of the antibody decoration. Judged by IC50 after 24h culture, the therapeutic effects of the drug formulated in the NPs with surface decoration could be 12.74 times higher than that of the bare NPs and 13.11 times higher than Taxol.     

Doxorubicin conjugated to D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS): conjugation chemistry, characterization, in vitro and in vivo evaluation

To develop a polymer-anticancer drug conjugate, D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) was employed as a carrier of doxorubicin (DOX) to enhance its therapeutic effects and reduce its side effects. Doxorubicin was chemically conjugated to TPGS. The molecular structure, drug loading efficiency, drug release kinetics and stability of the conjugate were characterized. The cellular uptake, intracellular distribution, and cytotoxicity were accessed by using MCF-7 breast cancer cells and C6 glioma cells as in vitro cell model. The conjugate showed higher cellular uptake efficiency and broader distribution within the cells. Judged by IC(50), the conjugate was found 31.8, 69.6, 84.1% more effective with MCF-7 cells and 43.9, 87.7, 42.2% more effective with C6 cells than the parent drug after 24, 48, 72 h culture, respectively. The in vivo pharmacokinetics and biodistribution were investigated after an i.v. administration at 5 mg DOX/kg body weight in rats. Promisingly, 4.5-fold increase in the half-life and 24-fold increase in the area-under-the-curve (AUC) of DOX were achieved for the TPGS-DOX conjugate compared with the free DOX. The drug level in heart, gastric and intestine was significantly reduced, which is an indication of reduced side effects. Our TPGS-DOX conjugate showed great potential to be a prodrug of higher therapeutic effects and fewer side effects than DOX itself.     

新型多肽聚酰胺-胺型靶向药物载体的载药性能及细胞吸收和毒性

背景：前期研究通过噬菌体展示体内筛选方法获得了一条NCI-H460非小细胞肺癌特异结合的多肽(Lung cancer targeting peptide, LCTP),将该多肽与修饰的聚酰胺-胺型(Polyamidoamine, PAMAM)树枝状高分子材料连接制备了纳米靶向药物载体PAMAM-Ac-FITC-LCTP,该载体在体内外对非小细胞肺癌NCI-H460具有很好的靶向性。 目的：在前期研究基础上,进一步研究PAMAM-Ac-FITC-LCTP靶向载体对阿霉素的包埋、释放及其细胞吸收和毒性性能。 方法：以筛选到的多肽LCTP为靶向剂,构建了PAMAM-Ac-FITC-LCTP靶向载体。采用物理包埋法将PAMAM-Ac- FITC-LCTP与阿霉素连接,通过体外透析实验观察载体对药物的缓释功能,共聚焦显微镜观察细胞对药物的吸收。以游离阿霉素作为对照,MTT法观察载体载药后对NCI-H460细胞的作用。 结果与结论：PAMAM-Ac-FITC-LCTP对阿霉素的最大包埋率为7.46%。载体对药物具有明显的缓释作用,离子浓度、pH和温度对药物的释放具有影响,说明PAMAM-Ac-FITC-LCTP主要是通过静电相互作用与阿霉素结合。PAMAM-Ac- FITC-LCTP/阿霉素短时间内较单独药物更高效进入NCI-H460细胞,而复合物24 h 的细胞毒性与阿霉素对细胞的毒性基本一致。以上结果说明PAMAM-Ac-FITC-LCTP可能是一个肿瘤治疗和诊断中很有用的药物靶向传输载体。     

Tamoxifen-loaded thiolated alginate-albumin nanoparticles as antitumoral drug delivery systems

Nanoparticles based on disulfide bond reduced bovine serum albumin and thiolated alginate (alginate-cysteine conjugate) have been prepared by coacervation method and have been loaded with tamoxifen (TMX). The TMX load into the nanoparticles was optimized (4-6 μg/mg NP) by freeze-drying the systems before the loading procedure. Maximum TMX release (45-52%) took place between 2 and 25 h. Cytotoxicity of unloaded nanoparticles in MCF-7 and HeLa cells was not observed, although a small decrease in viability took place at very high concentration. Cell uptake of nanoparticles occurred in both cell types and the presence of polysaccharide in the nanoparticle composition allowed a better interaction with cells. The administration of 10 μM TMX by TMX-nanoparticles was effective in both cellular lines, and the effect of the drug-loaded systems on MCF-7 cell cycle showed the efficacy of the TMX-loaded nanoparticles.     

Image-guided and tumor-targeted drug delivery with radiolabeled unimolecular micelles

Unimolecular micelles formed by dendritic amphiphilic block copolymers poly(amidoamine)–poly(l-lactide)-b-poly(ethylene glycol) conjugated with anti-CD105 monoclonal antibody (TRC105) and 1,4,7-triazacyclononane-N, N′, N-triacetic acid (NOTA, a macrocyclic chelator for ⁶⁴Cu) (abbreviated as PAMAM–PLA-b-PEG–TRC105) were synthesized and characterized. Doxorubicin (DOX), a model anti-cancer drug, was loaded into the hydrophobic core of the unimolecular micelles formed by PAMAM and PLA via physical encapsulation. The unimolecular micelles exhibited a uniform size distribution and pH-sensitive drug release behavior. TRC105-conjugated unimolecular micelles showed a CD105-associated cellular uptake in human umbilical vein endothelial cells (HUVEC) compared with non-targeted unimolecular micelles, which was further validated by cellular uptake in CD105-negative MCF-7 cells. In 4T1 murine breast tumor-bearing mice, ⁶⁴Cu-labeled targeted micelles exhibited a much higher level of tumor accumulation than ⁶⁴Cu-labeled non-targeted micelles, measured by serial non-invasive positron emission tomography (PET) imaging and confirmed by biodistribution studies. These unimolecular micelles formed by dendritic amphiphilic block copolymers that synergistically integrate passive and active tumor-targeting abilities with pH-controlled drug release and PET imaging capabilities provide the basis for future cancer theranostics.     

Self-assembled nanoparticles made from a new PEGylated poly(aspartic acid) graft copolymer for intravaginal delivery of poorly water-soluble drugs

New amphiphilic PEGylated poly(aspartic acid) graft copolymer (PASP-PEG-Ph) was synthesized as a nanocarrier for intravaginal drug delivery of poorly water-soluble drugs. PASP-PEG-Ph self-assembled into negatively charged spherically shaped nanoparticles in the presence of pH 4.5 and pH 7.0 vaginal fluid simulants with a diameter of approximately 200 nm as evidenced by Zeta-potentiometer, scanning electron microscope (SEM), dynamic light scattering (DLS) analysis. A significant number of stable NPs could be maintained at pH 4.5, 37 °C for 13 days. The PASP-PEG-Ph NP showed no significant cytotoxicity toward the T-cell line SupT1 and human vaginal epithelial cell line Vk2/E6E7 up to 1 mg/mL. The highest encapsulation efficiency of the model drug coumarin 6 (C6) by PASP-PEG-Ph was 92.0 ± 5.7%. The sustained release profile of the encapsulated C6 was demonstrated by an in vitro release study. An in vitro cellular uptake study revealed strong cellular uptake of the C6 loaded NP by SupT1 cells within 2 h.     

Theranostic liposomes of TPGS coating for targeted co-delivery of docetaxel and quantum dots

The aim of this work was to develop a new type of D-alpha-tocopheryl polyethylene glycol 1000 succinate mono-ester (TPGS) coated multi-functional (theranostic) liposomes, which contain both docetaxel and quantum dots (QDs) for cancer imaging and therapy. Non-targeting and folate receptor targeting TPGS coated theranostic liposomes were prepared by the solvent injection method and characterized for their particle size, polydispersity, zeta potential, surface chemistry and drug encapsulation efficiency. MCF-7 breast cancer cells of folate receptor overexpression were employed as an in vitro model to assess cellular uptake and cytotoxicity of the drug and QDs loaded liposomes. The mean particle size of the non-targeting and the targeting liposomes was found to be 202 and 210 nm, respectively. High resolution field emission transmission electron microscopy (FETEM) confirmed the presence of quantum dots in the peripheral hydrophobic membranes of the liposomes. The qualitative internalization of multi-functional liposomes by MCF-7 cells was visualized by confocal laser scanning microscopy (CLSM). The IC50 value, which is the drug concentration needed to kill 50% cells in a designated time period, was found to be 9.54 ± 0.76, 1.56 ± 0.19 and 0.23 ± 0.05 μg/ml for the commercial Taxotere(?), non-targeting and targeting liposomes, respectively after 24 h culture with MCF-7 cells. The targeting multi-functional liposomes showed greater efficacy than the non-targeting liposomes and thus great potential to improve the cancer imaging and therapy.     

Multifunctional nanoassemblies for vincristine sulfate delivery to overcome multidrug resistance by escaping P-glycoprotein mediated efflux

Multifunctional nanoassemblies (MNAs) were successfully developed for controlled delivery of water-soluble cationic vincristine sulfate (VCR) to overcome multidrug resistance (MDR). The incorporation of anionic small molecule of phosphatidylserine (PS) significantly enhanced the encapsulation efficiency of VCR in MNAs up to 94.4% by electrostatic interaction. Obvious sustained-release characteristics were found in VCR-loaded MNAs (VCR-MNAs) as the cumulative release of VCR was 83.2% at 96?h, and burst-release was effectively diminished. In vivo pharmacokinetics in rats following intravenous administration demonstrated that VCR-MNAs had higher AUC and longer t(1/2) than VCR solution (VCR-Sol). To investigate the MDR reversal effect and clarify the possible mechanism induced by MNAs, the cytotoxicity, cellular uptake and uptake mechanism experiments were performed in MCF-7 and P-glycoprotein over-expressing MCF-7/Adr cells, respectively. Compared with VCR-Sol, VCR-MNAs efficiently enhanced the cytotoxicity to 36.5-fold by increasing the cellular accumulation of VCR (12.6-fold higher) in MCF-7/Adr cells. The results of endocytosis inhibition experiment proved that VCR-MNAs were uptaken into the resistant cancer cells by clathrin- and caveolae-mediated endocytosis pathways, which escaped the efflux induced by P-gp transporter and thereby overcame the MDR of VCR.     

肝癌靶向载多烯紫杉醇聚多巴胺修饰琥珀酸酯-聚乳酸纳米粒子的研究

目的 建立一种简便的、具有肝癌靶向的聚多巴胺表面修饰聚合物纳米粒子以增加聚合物膜层和功能化的方法.方法 采用聚多巴胺修饰聚乙二醇1000-琥珀酸盐-聚乳酸纳米粒子(pD-TPGS-PLA/NPs),并用其包载多烯紫杉醇(DTX).为了达到靶向治疗肝癌的作用,再将半乳糖胺连接在pD-TPGS-PLA/NPs表面,从而通过配体-受体介导的作用提高DTX的递送效率.结果 聚多巴胺-琥珀酸酯-聚乳酸纳米粒子(pD-TPGS-PLA/NPs)和半乳糖胺-聚多巴胺-琥珀酸酯-聚乳酸纳米粒子(Gal-pD-TPGS-PLA/NPs)在粒径和形态学上与琥珀酸酯-聚乳酸纳米粒子(TPGS-PLA/NPs)有明显的不同.体外研究结果显示,TPGS-PLA/NPs、pD-TPGS-PLA/NPs和Gal-pD-TPGS-PLA/NPs对DTX有相似的释放行为.激光扫描共聚焦显微镜和流式细胞仪结果显示,HepG2细胞对于包载香豆素-6的Gal-pD-TPGS-PLA/NPs有较高地细胞摄取效率.相比于TPGS-PLA/NPs、pD-TPGS-PLA/NPs和临床应用的DTX药物多西他赛,载DTX的Gal-pD-TPGS-PLMNPs对HepG2细胞的生长有更强的抑制能力.体内抗肿瘤研究结果显示,在荷瘤裸鼠上注射包载DTX的Gal-pD-TPGS-PLA/NPs能有效减小肿瘤尺寸.结论 Gal-pD-TPGS-PLA/NPs可通过配体-受体识别与肝癌细胞发生特有的相互作用,有望成为一种极具潜力的靶向治疗肝癌的药物递送系统.     

Formulation of Docetaxel by folic acid-conjugated d-α-tocopheryl polyethylene glycol succinate 2000 (Vitamin E TPGS(2k)) micelles for targeted and synergistic chemotherapy

Although high efficacy has been showed, Paclitaxel and Docetaxel cause serious side effects due to the adjuvant used in their clinical formulation Taxol? and Taxotere?. We developed a micelle system with a newly synthesized TPGS(2k) polymer, which shows lower CMC of 0.0219 mg/ml compared with 0.2 mg/ml for traditional micelles with TPGS involved, to achieve sustained and controlled drug delivery with Docetaxel used as a model anti-cancer drug. The TPGS(2k) micelles were further conjugated to folic acid (FA) for targeted drug delivery. The Docetaxel-loaded TPGS(2k) micelles with and without FA conjugation were found of desired size and size distribution, high drug encapsulation efficiency and favorable drug release. In vitro studies using MCF-7 cancer cells demonstrated significantly the higher cellular uptake of the formulated drug for TPGS(2k) micelle formulation than that for Taxotere?. The targeting effects for the FA conjugated TPGS(2k) micelles are also demonstrated. The IC?? value, which is the drug concentration needed for 50% cell viability in the designated time period, is 103.4, 1.280 and 0.1480 μg/ml for MCF-7 cancer cells after 24, 48, and 72 h treatment respectively, which is greatly decreased to be 0.526, 0.251 and 0.233 μg/ml, i.e. a 99.5%, 80.4% decrease and 57.5% increase for the TPGS(2k) micelle formulation, and further decreased to be 0.1780, 0.1520 and 0.1140 μg/ml, i.e. a 99.8%, 88.1% and 23.0% decrease for the folic acid conjugated micelles, respectively. A synergistic effect between TPGS(2k) and Docetaxel is also achieved. The present work represents a new concept in the design of drug delivery systems--the carrier materials of the drug delivery system can also have therapeutic effects, which either modulate the side effects of, or promote a synergistic interaction with the formulated drug.     

Self-assembled nanoparticles based on hyaluronic acid-ceramide (HA-CE) and Pluronic® for tumor-targeted delivery of docetaxel

Hyaluronic acid-ceramide (HA-CE)-based self-assembled nanoparticles were developed for intravenous docetaxel (DCT) delivery. In this study, physicochemical properties, cellular uptake efficiency, and in vivo targeting capability of the nanoparticles developed were investigated. DCT-loaded nanoparticles composed of HA-CE and Pluronic 85 (P85) with a mean diameter of 110-140 nm were prepared and their morphological shapes were assessed using transmission electron microscopy (TEM). DCT release from nanoparticle was enhanced with increasing P85 concentrations in our in vitro model. Blank nanoparticles exhibited low cytotoxicity in U87-MG, MCF-7 and MCF-7/ADR cell lines. From cellular uptake studies, the nanoparticles developed enhanced the intracellular DCT uptake in the CD44-overexpressing cell line (MCF-7). The nanoparticles were shown to be taken up by the HA-CD44 interaction according to DCT and coumarin 6 (C6) cellular uptake studies. The multidrug resistance (MDR)-overcoming effects of DCT-loaded HA-CE/P85-based nanoparticles were also observed in cytotoxicity tests in MCF-7/ADR cells. Following the intravenous injection of DCT-loaded cyanine 5.5 (Cy5.5)-conjugated nanoparticles in MCF-7/ADR tumor-bearing mice, its in vivo targeting for CD44-overexpressing tumors was identified by non-invasive near-infrared (NIR) fluorescence imaging. These results indicate that the HA-CE-based nanoparticles prepared may be a promising anti-cancer drug delivery system through passive and active tumor targeting.