Heparin Immobilized Porous PLGA Microspheres for Angiogenic Growth Factor Delivery

Purpose Heparin immobilized porous poly(d,l-lactic-co-glycolic acid) (PLGA) microspheres were prepared for sustained release of basic fibroblast growth factor (bFGF) to induce angiogenesis.Materials and Methods Porous PLGA microspheres having primary amine groups on the surface were prepared using an oil-in-water (O/W) single emulsion method using Pluronic F-127 as an extractable porogen. Heparin was surface immobilized via covalent conjugation. bFGF was loaded into the heparin functionalized (PLGA-heparin) microspheres by a simple dipping method. The bFGF loaded PLGA-heparin microspheres were tested for in vitro release and in vivo angiogenic activity.Results PLGA microspheres with an open-porous structure were formed. The amount of conjugated amine group onto the microspheres was 1.93 ± 0.01 nmol/mg-microspheres, while the amount of heparin was 95.8 pmol/mg-microspheres. PLGA-heparin microspheres released out bFGF in a more sustained manner with a smaller extent of initial burst than PLGA microspheres, indicating that surface immobilized heparin controlled the release rate of bFGF. Subcutaneous implantation of bFGF loaded PLGA-heparin microspheres in mice significantly induced the formation of new vascular microvessels.Conclusions PLGA microspheres with an open porous structure allowed significant amount of heparin immobilization and bFGF loading. bFGF loaded PLGA-HP microspheres showed sustained release profiles of bFGF in vitro, demonstrating reversible and specific binding of bFGF to immobilized heparin. They also induced local angiogenesis in vivo in an animal model.     

Injectable Sustained-Release Depots of PLGA Microspheres for Insoluble Drugs Prepared by hot-Melt Extrusion

Purpose

Progesterone (PRG) was selected as a model drug to develop a long-acting injection system for poorly water-soluble drugs.

Methods

Microspheres with high density-low porosity were prepared by hot-melt extrusion (HME) combined with wet-milling as the representative formulation, and a microcrystal suspension was also studied as a comparison. The morphology, particle size and distribution, polymorphism, drug distribution, density and porosity were characterized by scanning electron microscopy, laser diffraction particle size analyzer, power X-ray diffraction and DSC respectively. The in vivo performance of the different formulations within 7 days after intramuscular injection was evaluated in male SD rats.

Results

The drug-loading rate of the microspheres could be as high as 40%. The average initial burst release of the microspheres (PLGA lactide:glycolide = 75:25) was only 6.7% much lower than that of the microsuspension (25.7%) and a sustained release was exhibited for at least 7 days. The release mechanism was speculated to be as follows. The microspheres are a drug depot with drug microcrystals in the PLGA matrix which is a layer by layer honeycomb structure.

Conclusions

Microspheres prepared by HME combined with wet-milling could achieve a long-term sustained release effect as a novel long-acting formulation strategy.

     

PLGA微球控释系统的突释及其控制

针对目前限制PLGA微球控释系统临床应用的突释问题，重点介绍了近年来国内外有关的研究进展，包括突释现象、突释原因、影响因素和控制突释的方法和技术。     

罗哌卡因-醋酸地塞米松PLGA微球的制备及体外释药特性研究

目的 制备罗哌卡因 醋酸地塞米松聚乳酸羟基乙酸共聚物（PLGA）微球（简称微球）并研究其体外释药特性。方法以PLGA为载体,采用W1/O/W2双重乳化 溶剂挥发法制备微球,研究实验过程中有机相PLGA浓度、外水相/有机相体积比、内水相体积、外水相聚乙烯醇（PVA）浓度几项因素变化对罗哌卡因 醋酸地塞米松PLGA微球粒径、表面形态﹑载药量﹑包封率和突释行为的影响。结果有机相PLGA浓度在制备微球的过程中是一个关键性因素。随着PLGA浓度增加,微球粒径增大,载药量﹑包封率明显提高,突释降低;外水相/有机相体积比增大,微球粒径增大, 载药量﹑包封率明显提高,微球表面更加光滑﹑微孔减少,突释降低;随着内水相体积增加使得微球表面的微孔明显增多,突释增加,载药量﹑包封率降低;当外水相PVA浓度由0.5%增加到2%,微球粒径变小,突释效应增加。通过优化条件制备的微球形状为球形,外观光滑圆整,粒径分布均匀,其中＞90%分布在20～70 μm。罗哌卡因载药量（7.48±0.33）%,包封率（70.97±2.36）%;醋酸地塞米松载药量（1.52±0.16）%,包封率（57.30±1.17）%。结论采用W1/O/W2双重乳化 溶剂挥发法成功制备罗哌卡因加醋酸地塞米松PLGA微球;以优化工艺制备的微球,在体外具有明显的缓释行为,释药曲线呈典型S形三阶段模式。     

两种方法制备肺靶向地塞米松磷酸钠微球的比较

目的:研究两种制备方法所制备的肺靶向地塞米松磷酸钠微球的特性,确定两种方法是否适合用于制备该微球。方法:对油／水型乳化-溶剂挥发法的工艺进行优化,以优化后的油／水型乳化-溶剂挥发法和水／油／水型乳化-溶剂挥发法制备微球,考察微球的大小、载药量、体外释药等性质。结果:两法所制的微球的平均粒径相近,载药量和体外释药特性各不相同。结论:两种方法都适合用于制备肺靶向的地塞米松微球。从载药量方面分析,优化后的油／水型乳化-溶剂挥发法较好。     

有源植入医疗器械核磁共振成像安全性评价方法研究

目的：在同样标准、尺度下进行有源植入医疗器械“MR兼容”性评价，确保有源植入医疗器械的使用安全和有效。方法：本文依据ISO/TS 10974：2018标准，结合中国食品药品检定研究院的工作，针对标准中要求的各个试验的试验依据、试验方法、接收准则进行了研究。结果：标准共涉及6项已知或可预见的风险，分别为温升、振动、力、力矩、非预期的刺激以及器械故障。为了评价这些风险，本文介绍了和6项风险相对应的9个试验项目，以此评价有源植入医疗器械的核磁兼容性。结论：目前国内针对有源植入医疗器械核磁兼容性的评价刚刚起步，一些检测手段和方法处在借鉴和摸索之中，本文系统性介绍了评价项目的考虑角度和方法，为国内从事有源植入物核磁兼容性设计及评价者提供参考。     

A Novel in Vitro Delivery System for Assessing the Biological Integrity of Protein upon Release from PLGA Microspheres

Purpose. The development of a novel in vitro system is required to assess the stability and release kinetics of a protein microsphere formulation used for drug delivery to the brain. Methods. Microspheres containing lysozyme as model protein were prepared using a (w/o/w) emulsion-solvent evaporation process. Both the active and total (active + inactive) encapsulation efficiencies and release profiles were determined. The biologic activity of lysozyme was measured using bacterial cell lysis; total protein content was measured using a ¹²⁵I-radiolabel. A novel in vitro apparatus was developed to determine kinetics over a sustained time period (>30 days). Results. The microencapsulation technique allowed an entrapment of active lysozyme at 80 ± 4% and a sustained (>42 days) in vitro release. The kinetics study showed that the novel in vitro system was able to detect the release of low amounts (ng) of protein. To improve the stability of the protein within microspheres and allow the release of biologically active lysozyme, a basic additive ( Mg(OH)₂ ) was successfully encapsulated. Conclusions. This novel in vitro system was appropriate to study protein microsphere release kinetics. In addition, the model is cost-effective and mimes brain physiological conditions more closely than previous models.     

PLGA微球的修饰及其应用

微球作为新型的药物载体系统已广泛用于临床研究,高分子化合物材料聚乳酸-羟基乙酸(pdy lactic-co-glycolic acid,PLGA)因其良好的生物相容性和生物可降解性备受关注。近年来,PLGA微球的研究一直是热门,针对其释放缺陷出现了很多复合修饰方法,主要包括环糊精、壳聚糖、聚乳酸、明胶、泊洛沙姆、聚乙烯亚胺等高分子材料的联用、针对末端基团进行化学修饰以及制备成核壳型微球,在保证包封率的情况下大大降低突释,改善药物释放曲线,从而在药物传递、基因治疗、影像诊断、组织工程等领域得到了广泛的应用。     

Bioadhesive Fluorescent Microspheres as Visible Carriers for Local Delivery of Drugs. II: Uptake of Insulin-Loaded PCEFB/PLGA Microspheres by the Gastrointestinal Tract

Uptake of novel inherently fluorescent microspheres composed of a luminescent polyanhydride, poly[p-(carboxyethylformamido)- benzoic anhydride] (PCEFB), and poly(lactide-co-glycolide) (PLGA) (2:1, weight ratio) by the gastrointestinal tract was evaluated by fluorescent microscopy. Oral efficiency of the incorporated insulin also was determined by measuring reduction of plasma glucose levels after feeding diabetic rats with a single dose of the microspheres. We found that PCEFB/PLGA microspheres could adhere to the intestinal epithelium and traverse the absorptive cells. A large number of the spheres were observed in spleen, whereas few were detected in liver within the evaluated period of time. Apparent reduction of the plasma glucose levels was observed over a span of 6 h postfeeding. The unique properties of the delivery system such as biodegradability, bioadhesivity, and inherently luminescent characteristics render it an ideal “visible” tracer for monitoring oral fate of polymeric microspheres.     

Adsorption of Salmon Calcitonin to PLGA Microspheres

Purpose. The interaction of salmon calcitonin (sCT) and poly (d,l-lactide-co-glycolide) was detected during preparation and evaluation of microspheres. The purpose of this study was to quantitate the extent and nature of the interaction. Methods. Blank microspheres were prepared by an aqueous emulsification solvent extraction technique. Adsorption studies were carried out at six concentrations of sCT and three concentrations of microspheres. Adsorption isotherms were constructed using the Langmuir and Freundlich treatments. Results. Adsorption at 1 mg/ml sCT concentration resulted in almost complete depletion of the peptide from the adsorption medium with the time to reach maximum adsorption decreasing with increasing microsphere concentration. At sCT concentrations below 100 µg/ml, a true equilibrium occurred in 1 hour or less while at higher concentrations (up to 350 µg/ml), a transient equilibrium was reached in 1 to 2 hours, followed by further adsorption of the peptide. The adsorption followed the Langmuir isotherm at concentrations below 200 µg/ml, indicating formation of a monolayer. Multilayer interaction, described by the Freundlich isotherm, occurred at higher concentrations and resulted in complete depletion of sCT from the adsorption medium. The affinity constant during monolayer formation was 0.09 and the plateau surface concentration was 5.1 µg/mg. The multilayer peptide-peptide adsorption showed a lower affinity (0.025) but higher capacity (24 µg/mg) than the monolayer peptide-polymer adsorption. Conclusions. The results show that poly (d,l-lactide-co-glycolide) microspheres have a high adsorption capacity for sCT which must be considered in formulating a controlled delivery product of this peptide.     

lontophoretic Devices for Drug Delivery

Transdermal drug delivery of ionized drugs can be enhanced by iontophoresis. Drug in the ionic form, contained in some reservoir, can be phoresed through the skin with a small current across two electrodes, one above the reservoir and one at a distal skin location. Positive ions can be introduced from the positive pole, or negative ions from the negative pole. The design and development of iontophoretic devices are rather simple. Some of the principles of operation and the advantages/disadvantages and clinical implications associated with these devices are outlined in this review.     

Carboplatin-Loaded PLGA Microspheres for Intracerebral Implantation: In Vivo Characterization

Abstract

Carboplatin is a potent anticancer agent that has shown efficacy in clinical trials against malignant glioma, one of the most deadly cancers in humans. However, a high systemic dose is required to achieve an effective concentration in the brain because of the presence of the blood-brain barrier (BBB). Such a high dose can cause many side effects. Local delivery of antitumor agents to the brain using injectable and biodegradable microspheres is a new strategy for the treatment of malignant glioma. This method is able to bypass the BBB and allows maximal local exposure and minimal systemic exposure to avoid the severe side effects of carboplatin. Delivering sustained-release microspheres directly to the tumor site could also control local tumor recurrence and improve survival. In the present studies, carboplatin-loaded microspheres were delivered intracerebrally in rats. No signs of systemic or neurologic toxicity associated with the microspheres implanted in the rat brain were observed. The in vivo release of carboplatin followed apparently zero-order release kinetics up to 30 days. The surface characteristics of the microspheres retrieved from the rat brains changed with the progress of polymer biodegradation. Implantation of the microspheres evoked a transient and localized inflammatory reaction that was well tolerated by the animals.     

Stromal-Derived Factor-1 Alpha-Loaded PLGA Microspheres for Stem Cell Recruitment

Purpose  

Stromal-derived factor-1 alpha (SDF-1α) is a chemoattractant that has been investigated for treating various diseases, with the goal of recruiting endogenous stem cells to the site of injury. Biodegradable PLGA microspheres were investigated as a means to deliver SDF-1α in a sustained-release manner.     

Carboplatin-Loaded PLGA Microspheres for Intracerebral Implantation: In Vivo Characterization

Carboplatin is a potent anticancer agent that has shown efficacy in clinical trials against malignant glioma, one of the most deadly cancers in humans. However, a high systemic dose is required to achieve an effective concentration in the brain because of the presence of the blood-brain barrier (BBB). Such a high dose can cause many side effects. Local delivery of antitumor agents to the brain using injectable and biodegradable microspheres is a new strategy for the treatment of malignant glioma. This method is able to bypass the BBB and allows maximal local exposure and minimal systemic exposure to avoid the severe side effects of carboplatin. Delivering sustained-release microspheres directly to the tumor site could also control local tumor recurrence and improve survival. In the present studies, carboplatin-loaded microspheres were delivered intracerebrally in rats. No signs of systemic or neurologic toxicity associated with the microspheres implanted in the rat brain were observed. The in vivo release of carboplatin followed apparently zero-order release kinetics up to 30 days. The surface characteristics of the microspheres retrieved from the rat brains changed with the progress of polymer biodegradation. Implantation of the microspheres evoked a transient and localized inflammatory reaction that was well tolerated by the animals.     

Development of Biodegradable Starch Microspheres for Intranasal Delivery

Domperidone microspheres for intranasal administration were prepared by emulsification crosslinking technique. Starch a biodegradable polymer was used in preparation of microspheres using epichlorhydrine as cross-linking agent. The formulation variables were drug concentration and polymer concentration and batch of drug free microsphere was prepared for comparisons. All the formulations were evaluated for particle size, morphological characteristics, percentage drug encapsulation, equilibrium swelling degree, percentage mucoadhesion, bioadhesive strength, and in vitro diffusion study using nasal cell. Spherical microspheres were obtained in all batches with mean diameter in the range of above 22.8 to 102.63 μm. They showed good mucoadhesive property and swelling behaviour. The in vitro release was found in the range of 73.11% to 86.21%. Concentration of both polymer and drug affect in vitro release of drug.     

Lysozyme Release and Polymer Erosion Behavior of Injectable Implants Prepared from PLGA-PEG Block Copolymers and PLGA/PLGA-PEG Blends

Purpose

We evaluated the controlled release of lysozyme from various poly(D,L-lactic-co-glycolic acid) (PLGA) 50/50-polyethylene glycol (PEG) block copolymers relative to PLGA 50/50.

Methods

Lysozyme was encapsulated in cylindrical implants (0.8 mm diameter) by a solvent extrusion method. Release studies were conducted in phosphate buffered saline +0.02% Tween 80 (PBST) at 37°C. Lysozyme activity was measured by a fluorescence-based assay. Implant erosion was evaluated by kinetics of polymer molecular weight decline, water uptake, and mass loss.

Results

Lysozyme release from an AB15 di-block copolymer (15% 5 kDa PEG, PLGA 28 kDa) was very fast, whereas an AB10 di-block copolymer (with 10% 5 kDa PEG, PLGA 45 kDa) and ABA10 tri-block copolymer (with 10% 6 kDa PEG, PLGA 27 kDa) showed release profiles similar to PLGA. We achieved continuous lysozyme release for up to 4 weeks from AB10 and ABA10 by lysozyme co-encapsulation with the pore-forming and acid-neutralizing MgCO₃, and from AB15 by co-encapsulation of MgCO₃ and blending AB15 with PLGA. Lysozyme activity was mostly recovered during 4 weeks.

Conclusions

These block co-polymers may have utility either alone or as PLGA blends for the controlled release of proteins.     

PLGA/Liposome Hybrid Nanoparticles for Short-Chain Ceramide Delivery

Purpose

Rapid premature release of lipophilic drugs from liposomal lipid bilayer to plasma proteins and biological membranes is a challenge for targeted drug delivery. The purpose of this study is to reduce premature release of lipophilic short-chain ceramides by encapsulating ceramides into liposomal aqueous interior with the aid of poly (lactic-coglycolicacid) (PLGA).

Methods

BODIPY FL labeled ceramide (FL-ceramide) and BODIPY-TR labeled ceramide (TR-ceramide) were encapsulated into carboxy-terminated PLGA nanoparticles. The negatively charged PLGA nanoparticles were then encapsulated into cationic liposomes to obtain PLGA/liposome hybrids. As a control, FL-ceramide and/or TR ceramide co-loaded liposomes without PLGA were prepared. The release of ceramides from PLGA/liposome hybrids and liposomes in rat plasma, cultured MDA-MB-231 cells, and rat blood circulation was compared using fluorescence resonance energy transfer (FRET) between FL-ceramide (donor) and TR-ceramide (acceptor).

Results

FRET analysis showed that FL-ceramide and TR-ceramide in liposomal lipid bilayer were rapidly released during incubation with rat plasma. In contrast, the FL-ceramide and TR-ceramide in PLGA/liposome hybrids showed extended release. FRET images of cells revealed that ceramides in liposomal bilayer were rapidly transferred to cell membranes. In contrast, ceramides in PLGA/liposome hybrids were internalized into cells with nanoparticles simultaneously. Upon intravenous administration to rats, ceramides encapsulated in liposomal bilayer were completely released in 2 min. In contrast, ceramides encapsulated in the PLGA core were retained in PLGA/liposome hybrids for 4 h.

Conclusions

The PLGA/liposome hybrid nanoparticles reduced in vitro and in vivo premature release of ceramides and offer a viable platform for targeted delivery of lipophilic drugs.     

Inhibition of Peptide Acylation in PLGA Microspheres with Water-soluble Divalent Cationic Salts

Purpose  To test the potential of water-soluble divalent cationic salts to inhibit acylation of octreotide encapsulated in poly(D,L-lactic-co-glycolic acid)-star (PLGA) microspheres. Methods  The divalent cationic salts, calcium chloride and manganese chloride, previously shown to disrupt peptide sorption, were introduced in PLGA microspheres prepared by the double emulsion-solvent evaporation method. Peptide stability was monitored by reversed-phase high performance liquid chromatography (RP-HPLC) and identified by liquid chromatography coupled with mass spectrometry (LC-MS) during microsphere degradation under physiological conditions for 4 weeks. Microsphere morphology and salt content were examined by scanning electron microscopy (SEM) and inductively coupled plasma-optical emission spectroscopy (ICP-OES), respectively. Results  Addition of divalent cationic salts solely to the organic phase did not provide acylation inhibition. However, addition of the salt inhibitors to both the primary emulsion and the outer water phase resulted in improved drug and salt encapsulation efficiency as well as significantly decreased salt leaching and octreotide acylation. After 28 days, the extent of acylation inhibition afforded by divalent cations was > 58% relative to 13% for the NaCl control group. Conclusions  Water-soluble divalent cationic salts represent a suitable class of stabilizer of peptide acylation in PLGA microspheres and this study provides an important formulation approach to maximize stabilizer potency.