Hydrophobically modified glycol chitosan nanoparticles as carriers for paclitaxel.

Self-assembled nanoparticles based on hydrophobically modified glycol chitosan (HGC) were prepared as a carrier for paclitaxel. HGC conjugates were prepared by chemically linking 5beta-cholanic acid to glycol chitosan chains using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide chemistry. In phosphate-buffered saline (PBS; pH 7.4), the synthesized HGC conjugates formed nano-sized particles with a diameter of 200 nm and exhibited high thermodynamic stability as reflected by their low critical aggregation concentration (0.03 mg/ml). Paclitaxel was efficiently loaded into HGC nanoparticles up to 10 wt.% using a dialysis method. The paclitaxel-loaded HGC (PTX-HGC) nanoparticles were 400 nm in diameter and were stable in PBS for 10 days. These PTX-HGC nanoparticles also showed sustained release of the incorporated of paclitaxel (80% of the loaded dose was released in 8 days at 37 degrees C in PBS). Owing to sustained release, the PTX-HGC nanoparticles were less cytotoxic to B16F10 melanoma cells than free paclitaxel formulated in Cremophor EL. Injection of PTX-HGC nanoparticles into the tail vein of tumor-bearing mice prevented increases in tumor volume for 8 days. Finally, PTX was less toxic to the tumor-bearing mice when formulated in HGC nanoparticles than when formulated with Cremophor EL.     

紫杉醇纳米粒子的制备及其应用

背景:紫杉醇临床用剂型紫素易引起过敏反应,因此研制新的紫杉醇新剂型就显得十分有意义。目的:研制紫杉醇新剂型,观察其在动物模型上治疗肿瘤的效果。方法:合成具有自主知识产权的生物可降解材料医用聚己内酯。采用溶剂替代法制备载紫杉醇纳米粒子,对其粒径、形态、紫杉醇含量、体外释放等进行测定。选用TA2系实验小鼠,建立乳腺癌动物模型,随机分为5组,分别局部注射生理盐水、紫素、低剂量、中剂量及高剂量紫杉醇纳米粒子进行治疗。结果与结论:实验制备的紫杉醇纳米粒子平均粒径约为153.54nm,包埋率为87.25%,紫杉醇含量19.06%。体外可恒定释放30d以上。2周药物治疗显示,各治疗组均不同程度上抑制了肿瘤的生长,其中紫杉醇纳米粒子中、高剂量组的抑瘤率明显高于紫素治疗组(P<0.01)。提示紫杉醇纳米粒子可缓释药物,中剂量组和高剂量组对小鼠乳腺癌的抑瘤率高于紫素组。     

Paclitaxel nanoparticles for the potential treatment of brain tumors.

Despite the advances in tumor therapy, patients with primary brain tumors and brain metastases have a very poor prognosis. Low responses to chemotherapy are mainly attributed to impermeability of the blood-brain barrier to cytotoxic agents. Paclitaxel has been shown to be active against gliomas and various brain metastases. However, its use in treatment of brain tumors is limited due to low blood-brain barrier permeability and serious side effects associated with administration of the paclitaxel solvent, Cremophor EL. Lack of paclitaxel brain uptake is thought to be associated with the p-glycoprotein (p-gp) efflux transporter. In this work, paclitaxel (PX) was entrapped in novel cetyl alcohol/polysorbate nanoparticles. Paclitaxel nanoparticles (PX NPs) were characterized by means of size, short-term stability, drug entrapment efficiency, and release profile. The PX NP cytotoxicity profile was monitored using two different cell lines, U-118 and HCT-15. Brain uptake of PX NPs was evaluated using an in situ rat brain perfusion model. The results suggest that entrapment of paclitaxel in nanoparticles significantly increases the drug brain uptake and its toxicity toward p-glycoprotein expressing tumor cells. It was hypothesized that PX NPs could mask paclitaxel characteristics and thus limit its binding to p-gp, which consequently would lead to higher brain and tumor cell uptake of the otherwise effluxed drug.     

Polymeric micelles of poly(2-ethyl-2-oxazoline)-block-poly(epsilon-caprolactone) copolymer as a carrier for paclitaxel.

Polymeric micelles based on amphiphilic block copolymers of poly(2-ethyl-2-oxazoline) (PEtOz) and poly(epsilon -caprolactone) (PCL) were prepared in an aqueous phase. The loading of paclitaxel into PEtOz-PCL micelles was confirmed by 1H-NMR spectra. Paclitaxel was efficiently loaded into PEtOz-PCL micelles using dialysis method, and the loading content of paclitaxel in micelles was in the range 0.5-7.6 wt.% depending on the block composition of block copolymers, organic solvent used in the dialysis, and feed weight ratio of paclitaxel to block copolymer. The higher the content of hydrophobic block in the block copolymers, the higher the loading efficiency of micelles for paclitaxel. When acetonitrile was used as solvent, a higher drug loading efficiency was obtained than with THF. The loading efficiency decreased with increasing feed weight ratio of paclitaxel to block copolymer from 0.1:1 to 0.2:1. The hydrodynamic diameters of paclitaxel-loaded micelles were in the range 18.3-23.4 nm with narrow size distribution. The hemolysis test of PEtOz-PCL performed in vitro indicated that the toxicity of PEtOz-PCLs to lipid membrane was not significant compared with Tween 80, and was comparable to that observed with Cremophore EL. The proliferation inhibition activity of paclitaxel-loaded micelles for KB human epidermoid carcinoma cells was also evaluated in vitro. Paclitaxel-entrapped polymeric micelles exhibited comparable activity to that observed with Cremophore EL-based paclitaxel formulations in inhibiting the growth of KB cells.     

In vitro degradation of nanoparticles prepared from polymers based on DL-lactide, glycolide and poly(ethylene oxide)

Nanoparticles of poly(DL-lactic acid) (PDLLA), poly(DL-lactic-co-glycolic acid) (PLGA) and poly(ethylene oxide)-PLGA diblock copolymer (PEO-PLGA) were prepared by the salting-out method. The in vitro degradation of PDLLA, PLGA and PEO-PLGA nanoparticles in PBS (pH 7.4) at 37 degrees C was studied. The particle size, molecular weight of the polymers and the amount of lactic and glycolic acids formed were followed in time. PDLLA nanoparticles gradually degraded over a period of 2 years and retain their size during that period. A faster degradation was observed for PLGA nanoparticles, which was nearly complete after 10 weeks. PLGA nanoparticles retained their size during that period. In PEO-PLGA nanoparticles, the ester bond connecting the PEO and the PLGA segments was preferentially cleaved, which led to a relatively fast decrease in molecular weight and to (partial) aggregation, as multimodal size distributions were observed. PEO-PLGA nanoparticles were almost completely degraded within 8 weeks.     

Development of nonionic surfactant/phospholipid o/w emulsion as a paclitaxel delivery system.

Paclitaxel is an anticancer agent with low aqueous solubility. More extensive clinical use of this drug is somewhat delayed due to lack of appropriate delivery vehicles. An attempt was made to adopt an o/w emulsion as the drug carrier which incorporated paclitaxel in the triacylglycerol stabilized by a mixed-emulsifier system. A suitable formulation was found in this study: 0.75 mg/ml paclitaxel, 10% (w/v) oil blend, 4% (w/v) EPC, 3% (w/v) Tween 80 in 2.25% (w/v) glycerol solution. The formulated emulsion has very good stability when stored at 4 degrees C, and the paclitaxel containment efficiency can be maintained above 95% and the mean emulsion diameter around 150 nm for at least 3 months. Paclitaxel-emulsion displayed cytotoxicity against HeLa cells with IC50 at 30 nM. The average life span of ascitic-tumor-bearing mice was prolonged significantly by the treatment of paclitaxel-emulsion (P<0.05). The formulated emulsion is a promising carrier for paclitaxel and other lipophilic drugs.     

Stable paclitaxel formulations in oily contrast medium.

Stable paclitaxel/Lipiodol solutions as well as emulsions were developed for the treatment of solid tumors including hepatocellular carcinoma. Paclitaxel could be dissolved in Lipiodol, an oily contrast medium, but precipitated out and formed aggregates with time. Paclitaxel precipitation was due to the inter- and intra-molecular hydrogen bonding of paclitaxel molecules. Time-dependent paclitaxel aggregation was completely prevented by adding small amounts of additional solvents, which are miscible with Lipiodol. It was also notable that paclitaxel helped in stabilizing the water-in-oil (w/o) type emulsion of Lipiodol and Iopamiro. The stability, physical properties and in vitro drug release profiles of the stable paclitaxel solutions and emulsions were characterized. When the stable oily paclitaxel solution was used for the treatment of B16F10 melanoma in C57BL/6 mice, the malignant cells were eradicated completely in 2 weeks, whereas the solid tumor grew rapidly and metastasized to the thigh and to other organs in the control group. Also, the mice survived for more than 1 year after the paclitaxel treatment, whereas all of those in the control group died in 40 days.     

Paclitaxel poliglumex (XYOTAX, CT-2103): a macromolecular taxane.

Paclitaxel poliglumex (PPX) is an innovative macromolecular taxane designed to increase the therapeutic index of paclitaxel. This large macromolecular conjugate of paclitaxel and poly-L-glutamic acid accumulates in tumor tissues by taking advantage of the enhanced permeability of tumor vasculature and lack of lymphatic drainage. Preclinical studies in animal tumor models demonstrate that PPX is more effective than standard paclitaxel and is associated with prolonged tumor exposure to active drug while minimizing systemic exposure. Phase 1 and 2 clinical studies with PPX showed encouraging outcomes compared to standard taxanes with reduced neutropenia and alopecia and allowed a more convenient administration schedule without the need for routine premedications. Human pharmacokinetic data are consistent with prolonged tumor exposure to active drug and a limited systemic exposure. Three phase 3 trials in nonsmall cell lung cancer (STELLAR 2, 3, and 4) have completed enrollment and results are expected in 2005.     

紫杉醇磁性脂质体制备及其性质的研究

目的制备紫杉醇磁性脂质体。方法采用薄膜超声法制备紫杉醇磁性脂质体;用透射电镜考察脂质体形态;高效液相色谱法测定紫杉醇磁性脂质体紫杉醇含量及组织分布。结果制得紫杉醇磁性脂质体外观、形态良好,平均粒径600-800 nm,包封率在90%以上。紫杉醇磁性脂质体体内、外具有良好得磁场响应性。应用紫杉醇磁性脂质体加磁场方式给药可以使紫杉醇有效的聚集到靶部位。结论紫杉醇磁性脂质体在磁场的作用下可以有效提高组织内化疗药物浓度。     

The effect of paclitaxel-loaded nanoparticles with radiation on hypoxic MCF-7 cells

BACKGROUND AND OBJECTIVE: The inability of radiotherapy to eradicate completely certain human tumours may be due to the presence of resistant hypoxic cells. Several studies have confirmed the radiosensitizing effect of paclitaxel, a microtubular inhibitor. The object of this study was to evaluate the physicochemical characteristics of paclitaxel-loaded nanoparticles, and determine the ability of the released paclitaxel to radiosensitize hypoxic human breast carcinoma cells (MCF-7) with respect to radiation dose. METHODS: The poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles containing paclitaxel were prepared by o/w emulsification-solvent evaporation method. The morphology of the paclitaxel-loaded nanoparticles was investigated by scanning electron microscopy. The drug encapsulation efficiency (EE) and in vitro release profile were measured by high-performance liquid chromatography. Cell cycle was evaluated by flow cytometry. Cell viability was measured by the ability of single cells to form colonies in vitro. RESULTS: The prepared nanoparticles were spherical with diameter between 200 and 800 nm. The EE was 85.5%. The drug release pattern was biphasic with a fast release rate followed by a slow one. Co-culture of human breast carcinoma cells (MCF-7) with paclitaxel-loaded nanoparticles demonstrated that released paclitaxel retained its bioactivity to block cells in the G2/M phase of the cell cycle and effectively sensitized hypoxic MCF-7 cells to radiation with radiosensitivity shown to be dependent of radiation dose at levels of dosages studied. The sensitizer enhancement ratio for paclitaxe-loaded nanoparticles at 10% survival is approximately 1.4. CONCLUSION: This work has demonstrated that paclitaxel can be effectively released from a biodegradable PLGA nanoparticle delivery system while maintaining potent combined cytotoxic and radiosensitizing abilities for hypoxic tumour cells.     

Transport of paclitaxel (Taxol) across the blood-brain barrier in vitro and in vivo

Paclitaxel concentrations in the brain are very low after intravenous injection. Since paclitaxel is excluded from some tumors by p-glycoprotein (p-gp), the same mechanism may prevent entry into the brain. In vitro, paclitaxel transport was examined in capillaries from rat brains by confocal microscopy using BODIPY Fl-paclitaxel. Western blots and immunostaining demonstrated apical expression of p-gp in isolated endothelial cells, vessels, and tissue. Secretion of BODIPY Fl-paclitaxel into capillary lumens was specific and energy-dependent. Steady state luminal fluorescence significantly exceeded cellular fluorescence and was reduced by NaCN, paclitaxel, and SDZ PSC-833 (valspodar), a p-gp blocker. Leukotriene C(4) (LTC(4)), an Mrp2-substrate, had no effect. Luminal accumulation of NBDL-cyclosporin, a p-gp substrate, was inhibited by paclitaxel. In vivo, paclitaxel levels in the brain, liver, kidney, and plasma of nude mice were determined after intravenous injection. Co-administration of valspodar led to increased paclitaxel levels in brains compared to monotherapy. Therapeutic relevance was proven for nude mice with implanted intracerebral human U-118 MG glioblastoma. Whereas paclitaxel did not affect tumor volume, co-administration of paclitaxel (intravenous) and PSC833 (peroral) reduced tumor volume by 90%. Thus, p-gp is an important obstacle preventing paclitaxel entry into the brain, and inhibition of this transporter allows the drug to reach sensitive tumors within the CNS.     

紫杉醇可降解乙交酯/丙交酯共聚物材料对人脐动脉平滑肌细胞的作用(英文)

背景：冠脉支架术后较高的早期急性再闭塞及晚期再狭窄已成为研究热点,而生物降解材料携带对抑制再狭窄有效的药物——紫杉醇金属涂层支架有望解决这一问题。目的：评价含药物紫杉醇可降解支架材料乙交酯/丙交酯共聚物（PLLGA）对人脐动脉平滑肌细胞的作用。设计、时间及地点：对比观察的细胞学实验,于2003-07/2005-07在吉林大学公共卫生学院毒理实验室完成。材料：含紫杉醇可降解材料PLLGA由中国科学院长春应用化学研究所提供,左旋丙素酯与聚乙交酯的比例为9∶1。方法：选取人脐动脉原代平滑肌细胞培养,得到传代细胞,然后与含1,2,3g紫杉醇PLLGA可降解材料一起培养,以金属支架组和不含紫杉醇只含PLLGA组为对照。主要观察指标：分别在培养0,24,48,72h后在相差显微镜下观察不同材料对细胞生长情况的影响。结果：PLLGA对照组及金属支架组对平滑肌细胞生长无影响,含1,2,3g紫杉醇的降解材料组细胞生长状态不佳,至培养72h,与PLLGA对照组及金属支架组比较差异有非常显著性意义（P〈0.01）。结论：PLLGA可作为抑制平滑肌细胞生长的血管内支架材料。     

Preparation and in vitro anticancer activity of wheat germ agglutinin (WGA)-conjugated PLGA nanoparticles loaded with paclitaxel and isopropyl myristate.

The purpose of this study was to develop a novel lectin-conjugated isopropyl myristate (IPM)-incorporated PLGA nanoparticle system (NP) for the local delivery of paclitaxel to the lungs. Wheat germ agglutinin (WGA) was conjugated onto preformed IPM- and paclitaxel-loaded PLGA NPs by a two-step carbodiimide method following comparative uptake studies of Concanavalin A, Ricinus communis-120 and WGA on A549, H1299 and CCL-186 cells. WIT-NP with mean diameter of 331 nm and zeta potential of -4.3 mV were prepared with yield of 66% and paclitaxel encapsulation efficiency of 61%. Particle size was expanded by surface conjugation with WGA, while zeta potential was reduced by the addition of IPM and WGA. In vitro paclitaxel release profile was not affected by WGA but initial drug release was enhanced by adding IPM into the formulation. The WIT-NP showed a burst-release of about 32% of the paclitaxel load within the first 5 h followed by a slow zero-order release of another 7% of the drug load in the next 115 h. Compared with the clinical paclitaxel formulation, paclitaxel-loaded nanoparticles without IPM or WGA, or paclitaxel-loaded nanoparticles with only IPM or WGA, the WIT-NP had superior in vitro cytotoxicity against A549 and H1299 cells. IC50 for WIT-NP after 5 and 24 h incubation with A549 cells were not significantly different (15.5 and 15 microM, respectively) whereas the clinical formulation was not cytotoxic after 5 h but had IC50 of 14 microM after 24 h incubation. WIT-NP exhibited stronger cell-killing effect because of more efficient cellular uptake via WGA-receptor-mediated endocytosis and IPM-facilitated release of paclitaxel from the NPs.     

Preparation of stable insulin-loaded nanospheres of poly(ethylene glycol) macromers and N-isopropyl acrylamide.

A series of nanospheres composed of temperature-sensitive poly(N-isopropylacrylamide), poly(ethylene glycol) 400 dimethacrylate, and poly(ethylene glycol) 1000 methacrylate was prepared by a thermally-initiated free radical dispersion polymerization method. Insulin was loaded into the nanoparticles by equilibrium partitioning. The loading capacity of insulin into the nanoparticles was 2.1% (2.1 mg insulin/100 mg nanoparticles). The stability of the loaded insulin at elevated temperatures was investigated by reverse phase high pressure liquid chromatography. The nanoparticles were able to protect the loaded insulin, as more than 80% of the loaded insulin could still be detected compared to 0% for the control (0.1% insulin solution in PBS) when heated to 80 °C for 5 h. The stability of the loaded insulin at high shear stress (289 1/s) was also investigated. No significant loss of insulin was detected both from nanoparticles loaded with insulin sample and the control (0.1% insulin solution in PBS). The results showed that shear stress alone did not have a major effect on insulin denaturation. The ability of the nanoparticles to protect the insulin from high temperature and high shear stress made the system a good candidate as a carrier for insulin for fluidized bed coating technology.     

抑制PI3K/Akt信号通路提高膀胱癌化疗效果的实验研究

目的:探讨通过抑制Akt信号通路提高抗癌药物紫杉醇对膀胱癌T-24的杀伤作用。方法:采用MTT法检测Akt抑制剂鱼藤素、紫杉醇单独以及两药联合应用对T-24细胞的增殖抑制率,采用流式细胞术(FCM)检测药物单独或联合作用对细胞周期的影响。结果:联合鱼藤素能够显著提高紫杉醇对T-24细胞的增殖抑制率(P<0.01),协同治疗指数<1,具有协同治疗作用;FCM结果显示联合鱼藤素使细胞阻滞在G0/G1期的比例增加,S期比例减少,与对照组比较,差异有统计学意义(P<0.05)。结论:抑制Akt信号通路能够显著提高紫杉醇对膀胱癌T-24细胞的杀伤作用。     

载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒抑制人肝癌细胞HepG2的作用

背景：临床上应用的紫杉醇注射剂毒性大,过敏反应发生率高。目的：研制载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒,观察其对人肝癌细胞HepG2的抑制及诱导细胞凋亡的作用。方法：采用MTT法检测0,3.125,6.25,12.5,25,50,100mg/L载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子或紫杉醇作用后人肝癌细胞HepG2的生长;观察25mg/L载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子或紫杉醇作用后人肝癌细胞HepG2的形态变化,并观察0,12.5,25,50mg/L载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子作用后细胞的凋亡情况。结果与结论：在3.125-100mg/L质量浓度范围内,载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子与紫杉醇均能明显抑制HepG2细胞的生长,且载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子显示出明显的缓释作用,随着作用时间的增加其抑制率显著增加,72h时抑制效果最好,但紫杉醇此现象不明显。载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子或紫杉醇作用后,细胞出现典型的凋亡形态,且随着载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子作用时间的增加这一现象更加典型;12.5,25,50mg/L载紫杉醇聚乳酸聚乙醇酸共聚物纳米粒子可明显诱导细胞凋亡,且有明显的量效和时效关系,质量浓度越高、时间越长效果越明显。     

Preparation and characterization of protein-loaded N-trimethyl chitosan nanoparticles as nasal delivery system.

In this study, the potential of N-trimethyl chitosan (TMC) nanoparticles as a carrier system for the nasal delivery of proteins was investigated. TMC nanoparticles were prepared by ionic crosslinking of TMC solution (with or without ovalbumin) with tripolyphosphate, at ambient temperature while stirring. The size, zeta-potential and morphology of the nanoparticles were investigated as a function of the preparation conditions. Protein loading, protein integrity and protein release were studied. The toxicity of the TMC nanoparticles was tested by ciliary beat frequency measurements of chicken embryo trachea and in vitro cytotoxicity assays. The in vivo uptake of FITC-albumin-loaded TMC nanoparticles by nasal epithelia tissue in rats was studied by confocal laser scanning microscopy. The nanoparticles had an average size of about 350 nm and a positive zeta-potential. They showed a loading efficiency up to 95% and a loading capacity up to 50% (w/w). The integrity of the entrapped ovalbumin was preserved. Release studies showed that more than 70% of the protein remained associated with the TMC nanoparticles for at least 3 h on incubation in PBS (pH 7.4) at 37 degrees C. Cytotoxicity tests with Calu-3 cells showed no toxic effects of the nanoparticles, whereas a partially reversible cilio-inhibiting effect on the ciliary beat frequency of chicken trachea was observed. In vivo uptake studies indicated the transport of FITC-albumin-associated TMC nanoparticles across the nasal mucosa. In conclusion, TMC nanoparticles are a potential new delivery system for transport of proteins through the nasal mucosa.     

Paclitaxel releasing films consisting of poly(vinyl alcohol)-graft-poly(lactide-co-glycolide) and their potential as biodegradable stent coatings.

Although substantial progress in catheter and stent design has contributed to the success of percutaneous transluminal angioplasty (PTA) of atherosclerotic disease, the incidence of restenosis caused by in-stent neointimal hyperplasia remains a serious problem. Therefore, stents with a non-degradable polymer coating showing controlled release of active ingredients have become an attractive option for the site-specific delivery of anti-restenotic agents. Biodegradable coatings using polyesters, namely poly(lactic-co-glycolic acid) (PLGA) and different poly(vinyl alcohol)-graft-poly(lactic-co-glycolic acid) (PVA-g-PLGA) as paclitaxel-eluting stent coating materials were investigated here to evaluate their influence on the release kinetic. Whereas PLGA showed sigmoid release behavior, the paclitaxel release from PVA-g-PLGA films was continuous over 40 days without initial drug burst. Wide angle X-ray diffraction confirmed that paclitaxel is dissolved in the polymer matrix. Paclitaxel crystallization can be observed at a drug load of > or =10%. The effect of drug loading on polymer degradation was studied in films prepared from PVA300-g-PLGA30 with paclitaxel loadings of 5% and 15% over a time period of 6 weeks. The results suggest a surface-like erosion mechanism in films. A model stent (Jostent peripheral) coated with Parylene N, a poly(p-xylylene) (PPX) derivate, was covered with a second layer of PVA300-g-PLGA15, and PVA300-g-PLGA30 by using airbrush method. Morphology of coated stents, and film integrity after expansion from 3.12 to 5 mm was investigated by scanning electron microscopy (SEM). The devices resisted mechanical stress during stent expansion and merit further investigation under in vivo conditions.     

On the design of in situ forming biodegradable parenteral depot systems based on insulin loaded dialkylaminoalkyl-amine-poly(vinyl alcohol)-g-poly(lactide-co-glycolide) nanoparticles

The feasibility to generate in situ forming parenteral depot systems from insulin loaded dialkylaminoalkyl-amine-poly(vinyl alcohol)-g-poly(lactide-co-glycolide) nanoparticles, was investigated. Biodegradable nanoparticles formed polymeric semi-solid depots upon injection into isotonic phosphate buffered saline (PBS) with no additional initiators. Nanoparticles (NP) prepared from the different amine-modified polyesters displayed a pronounced positive ζ-potential of > 25 mV. Diethylaminopropyl-amine-poly(vinyl alcohol)-g-poly(lactide-co-glycolide) (DEAPA(68)-PVAL-g-PLGA(1:20)), diethylaminoethyl-amine-poly(vinyl alcohol)-g-poly(lactide-co-glycolide) (DEAEA(33)-PVAL-g-PLGA(1:20)), and dimethylaminopropyl-amine-poly(vinyl alcohol)-g-poly(lactide-co-glycolide) (DMAPA(33)-PVAL-g-PLGA(1:20)), formed in situ depots by an ion-mediated aggregation with subsequent fusion of nanoparticles, related to a decreased glass transition temperature in the presence of PBS.Moreover, two factors, namely, polymer and insulin-nanocomplex concentration, were evaluated using a response surface design with respect to nanoparticles formation and insulin loading. Nanoparticles and implants were investigated by atomic force microscopy (AFM). The in vitro release from implants loaded with 2% insulin was carried out in a flow trough cell and quantified by high performance liquid chromatography (HPLC). The release showed a triphasic profile with an initial burst, pore diffusion and diffusion from the swollen matrix over more than two weeks. Insulin distribution in the implants during the release was followed by confocal laser scanning microscopy (CLSM). These findings combined with the protection of the model peptide against competitive macromolecules and the possibility to get dry powders by lyophilization make these nanoparticles-based depots suitable candidates for the design of controlled release devices for bioactive macromolecules.     

Paclitaxel therapy potentiates cold hyperalgesia in streptozotocin-induced diabetic rats through enhanced mitochondrial reactive oxygen species production and TRPA1 sensitization

Diabetes comorbidities include disabling peripheral neuropathy (DPN) and an increased risk of developing cancer. Antimitotic drugs, such as paclitaxel, are well known to facilitate the occurrence of peripheral neuropathy. Practitioners frequently observe the development or co-occurrence of enhanced DPN, especially cold sensitivity, in diabetic patients during chemotherapy. Preclinical studies showed that reactive oxygen species (ROS) and cold activate transient receptor potential ankyrin-1 (TRPA1) cation channels, which are involved in cold-evoked pain transduction signaling in DPN. Additionally, paclitaxel treatment has been associated with an accumulation of atypical mitochondria in the sensory nerves of rats. We hypothesized that paclitaxel might potentiate cold hyperalgesia by increasing mitochondrial injuries and TRPA1 activation. Thus, the kinetics of paclitaxel-induced cold hyperalgesia, mitochondrial ROS production, and TRPA1 expression were evaluated in dorsal root ganglia of normoglycemic and streptozotocin-induced diabetic rats. In diabetic rats, paclitaxel significantly enhanced cold hyperalgesia in comparison to normoglycemic paclitaxel-treated control rats. These effects were prevented by N-acetyl-cysteine, a reducing agent, and by HC030031, an antagonist of TRPA1. In diabetic and control rats, paclitaxel treatment was associated with an accumulation of atypical mitochondria and a 2-fold increase in mitochondrial ROS production. Moreover, mRNA levels of glutathione peroxidase 4 and glutathione-S-reductase were significantly lower in diabetic groups treated with paclitaxel. Finally, TRPA1 gene expression was enhanced by 45% in diabetic rats. Paclitaxel potentiation of cold hyperalgesia in diabetes may result from the combination of increased mitochondrial ROS production and poor radical detoxification induced by paclitaxel treatment and diabetes-related overexpression of TRPA1.