Preparation and characterization of a biodegradable drug targeting system for anticancer drug delivery: microsphere-antibody conjugate

Targeted delivery of anticancer drugs is one of the most actively pursued goals in anticancer chemotherapy. A major disadvantage of anticancer drugs is their lack of selectivity for tumour tissue, which causes severe side effects and results in low cure rates. Any strategy by which a cytotoxic drug is targeted to the tumour, thus increasing the therapeutic index of the drug, is a way of improving cancer chemotherapy and minimizing systematic toxicity. This study covers the preparation of the gelatin microsphere (GM)-anti-bovine serum albumin (anti-BSA) conjugate for the development of a drug targeting approach for anticancer drug delivery. Microspheres of 5% (w/v) gelatin content were prepared by crosslinking with glutaraldehyde (GTA) at 0.05 and 0.50% (v/v) concentration. Microspheres were in the size range of 71-141?microm. The suitability of these microspheres as drug carriers for anticancer drug delivery was investigated in vitro by studying the release profiles of loaded methotrexate (MTX) and 5-fluorouracil (5-FU) and the cytotoxicities on cancer cell lines. The in vitro MTX release profiles (approximately 22-46% released in 24 h depending on the amount of GTA used) were much slower compared to 5-FU (approximately 42-91% released in 24 h). Both drugs demonstrated an initial fast release, which was followed by gradual, sustained drug release. The MTT cytotoxicity test results of GMs loaded with 5-FU and MTX showed approximately 54-70% and approximately 52-67% cytotoxicities in 4 days. In general, incorporation of MTX and 5-FU in microspheres enhanced the cytotoxic effect in a more prolonged manner compared to the free drugs. Gelatin micospheres were chemically conjugated to anti-BSA and the antigen-antibody activities were studied by immunofluorescence. Results indicated approximately 80% binding with conjugated anti-BSA and BSA-FITC. Based on their low cytotoxicity and the high antigen binding efficiencies, anti-BSA conjugated gelatin microspheres could be suitable targeted drug carrier systems for selective and long-term delivery of anticancer drugs to a specific body compartment (i.e. bladder cancer).     

Gelatin-methotrexate conjugate microspheres as a potential drug delivery system

Gelatin-methotrexate microspheres for intra-tumor administration have possibilities for minimizing systemic toxicities of methotrexate (MTX) and overcoming its resistance. Gelatin-MTX conjugates prepared by a carbodiimide reaction were crosslinked with glutaraldehyde to form microspheres (MTX:gelatin molar ratios of 2:1, 15:1, and 21:1). Microspheres were evaluated under in vitro tumor conditions at pH 6.5 and 37 degrees C with and without Cathepsin B (Cat B). Some microspheres were capped with an ethanolamine/cyanoborohydride procedure. SEM of broken microspheres revealed a hollow shell structure. Superficial Cat B degradation influenced some free MTX release but produced no conjugate fragment release. HPLC measured release of fragments (<10 kDa) was very little and release of free MTX was small. However, higher drug load microspheres released less free MTX than lower drug load, a substantial lag phase of free MTX release from capped microspheres changed to an initial rapid release in uncapped microspheres, and fragments were only released from uncapped microspheres. Opened unstable Schiff base crosslinks in uncapped microspheres may allow enzyme to produce conjugate fragments not observed in capped microspheres. Free MTX release may occur from dissolved uncrosslinked conjugate within the hollow microspheres. Important relationships and observations are described that will be useful for gelatin and perhaps other proteinaceous microspheres.     

5-Fluorouracil-loaded chitosan coated polylactic acid microspheres as biodegradable drug carriers for cerebral tumours

The development of injectable microspheres for anticancer drug delivery into the brain is a major challenge. The possibility of entrapping 5-fluorouracil (5-FU) in chitosan coated monodisperse biodegradable microspheres with a mean diameter of 10-25 um was demonstrated. An emulsion of 5-FU (in water) and polylactic acid (PLA) dissolved in acetone-dichloromethane mixture was poured into an aqueous solution of chitosan (or poly-vinyl alcohol) with stirring using a high-speed homogenizer, for the formation of microspheres. 5-FU recovery in microspheres ranged from 44-66% depending on the polymer and emulsification systems used for the preparation. Scanning electron microscopy revealed that the chitosan coated microspheres had less surface micropores compared to PVA based preparations. The drug release behaviour from microspheres suspended in phosphate buffered saline exhibited a biphasic pattern. The amount of drug release was much higher initially (approximately 25%), followed by a constant slow release profile for a 30 days period of study. This chitosan coated PLA/PLGA microsphere formulation may have potential for the targeted delivery of 5-FU to treat cerebral tumours.     

5-Fluorouracil-loaded chitosan coated polylactic acid microspheres as biodegradable drug carriers for cerebral tumours

The development of injectable microspheres for anticancer drug delivery into the brain is a major challenge. The possibility of entrapping 5-fluorouracil (5-FU) in chitosan coated monodisperse biodegradable microspheres with a mean diameter of 10-25um was demonstrated. An emulsion of 5-FU (in water) and polylactic acid (PLA) dissolved in acetone-dichloromethane mixture was poured into an aqueous solution of chitosan (or poly-vinyl alcohol) with stirring using a high-speed homogenizer, for the formation of microspheres. 5-FU recovery in microspheres ranged from 44-66% depending on the polymer and emulsification systems used for the preparation. Scanning electron microscopy revealed that the chitosan coated microspheres had less surface micropores compared to PVA based preparations. The drug release behaviour from microspheres suspended in phosphate buffered saline exhibited a biphasic pattern. The amount of drug release was much higher initially (25%),followed by a constant slow release profile for a 30 days period of study. This chitosan coated PLA/PLGA microsphere formulation may have potential for the targeted delivery of 5-FU to treat cerebral tumours.     

Dextran cross-linked gelatin microspheres as a drug delivery system.

This paper describes the use of oxidized dextran as a cross-linker for the preparation of gelatin microspheres. Microspheres were obtained by a thermal gelation method and their dissolution kinetic was examined. In order to find evidence of sugar mediated cross-linking, swelling tests and gelatin microspheres dissolution experiments were performed. The obtained results indicated that oxidized dextran can form a cross-linked gelatin network which can reduce the dissolution of gelatin. More interestingly, gelatin microspheres treated by both native and oxidized dextran slow down, even if to a different extent, the release of the antitumor drug TAPP-Br used as a model compound. Taken together, our results suggest that oxidized dextran could be an interesting means to cross-link gelatin microspheres allowing the use of this delivery formulation for controlled release of drugs.     

A novel method to prepare magnetite chitosan microspheres conjugated with methotrexate (MTX) for the controlled release of MTX as a magnetic targeting drug delivery system

The purpose of the present study is to develop a new method to prepare magnetite chitosan microspheres conjugated with methotrexate (MTX) for the controlled release of MTX as a magnetic targeting drug delivery system. MTX was first conjugated to the chitosan chain via a peptide bond and then a suspension cross-linking technique was used for the production of magnetic chitosan microspheres with glutaraldehyde as the cross-linker. The MTX-loading capacity of the magnetic chitosan microspheres was determined and drug release experiments were also carried out to discuss the MTX release behavior. All the data support that the magnetic chitosan-MTX microspheres prepared in this method would have great potential application in magnetic targeting drug delivery technology.     

Synthesis of hydrophilic intra-articular microspheres conjugated to ibuprofen and evaluation of anti-inflammatory activity on articular explants

The main limitation of current microspheres for intra-articular delivery of non-steroidal anti-inflammatory drugs (NSAIDs) is a significant initial burst release, which prevents a long-term drug delivery. In order to get a sustained delivery of NSAIDs without burst, hydrogel degradable microspheres were prepared by co-polymerization of a methacrylic derivative of ibuprofen with oligo(ethylene-glycol) methacrylate and poly(PLGA-PEG) dimethacrylate as degradable crosslinker. Microspheres (40–100 μm) gave a low yield of ibuprofen release in saline buffer (≈2% after 3 months). Mass spectrometry analysis confirmed that intact ibuprofen was regenerated indicating that ester hydrolysis occurred at the carboxylic acid position of ibuprofen. Dialysis of release medium followed by alkaline hydrolysis show that in saline buffer ester hydrolysis occurred at other positions in the polymer matrix leading to the release of water-soluble polymers (>6–8000 Da) conjugated with ibuprofen showing that degradation and drug release are simultaneous. By considering the free and conjugated ibuprofen, 13% of the drug is released in 3 months. In vitro, ibuprofen-loaded MS inhibited the synthesis of prostaglandin E2 in articular cartilage and capsule explants challenged with lipopolysaccharides. Covalent attachment of ibuprofen to PEG-hydrogel MS suppresses the burst release and allows a slow drug delivery for months and the cyclooxygenase-inhibition property of regenerated ibuprofen is preserved.     

5-氟脲嘧啶明胶微球化学交联团化研究

本文报告用乳化法制备的5－FU明胶微球，用成二醛在异丙醇介质中固化，得到的微球球形圆整，分散性良好，5－FU含量40％～47％，包封率80％～94％，体外释放试验72h累计释放90％左右。紫外光谱扫描发现5－FU可与甲醛结合，结合物在水中溶解度较大，可导致药物包封率大大降低，提示5－FU的微球或微囊均不宜用甲醛固化。     

Formulation factors in preparing BTM-chitosan microspheres by spray drying method

Chitosan (CTS) microspheres were prepared by a spray drying method using type-A gelatin and ethylene oxide-propylene oxide block copolymer as modifiers. Surface morphological characteristics and surface charges of prepared microspheres were investigated by using scanning electron microscopy (SEM) and microelectrophoresis. The particle shape, size and surface morphology of microspheres were significantly affected by the concentration of gelatin. Betamethasone disodium phosphate (BTM)-loaded microspheres demonstrated good drug stability (less 1% hydrolysis product), high entrapped efficiency (95%) and positive surface charge (37.5 mV). The in vitro drug release from the microspheres was related to gelatin content. Microspheres containing gelatin/CTS 0.4 approximately 0.6(w/w) had a prolong release pattern for 12 h. These formulation factors were correlated to particulate characteristics for optimizing BTM microspheres in pulmonary delivery.     

5-Fluorouracil-loaded microspheres prepared by spray-drying poly(D,L-lactide) and poly(lactide-co-glycolide) polymers: Characterization and drug release

5-Fluorouracil (5-FU), a hydrosoluble anti-neoplastic drug, was encapsulated in microspheres of poly(D,L-lactide) (PLA) and poly(lactide-co-glycolide) (PLGA) polymers using the spray-drying technique, in order to obtain small size microspheres with a significant drug entrapment efficiency. Drug-loaded microspheres included between 47?±?11 and 67?±?12?µg 5-FU?mg^?1 microspheres and the percentage of entrapment efficiency was between 52?±?12 and 74?±?13. Microspheres were of small size (average diameter: 0.9?±?0.4–1.4?±?0.8?µm microspheres without drug; 1.1?±?0.5–1.7?±?0.9?µm 5-FU-loaded microspheres) and their surface was smooth and slightly porous, some hollows or deformations were observed in microspheres prepared from polymers with larger T_g. A fractionation process of the raw polymer during the formation of microspheres was observed as an increase of the average molecular weight and also of T_g of the polymer of the microspheres. The presence of 5-FU did not modify the T_g values of the microspheres. Significant interactions between the drug and each one of the polymers did not take place and total release of the included drug was observed in all cases. The time needed for the total drug release (28–129?h) was in the order PLA?>?PLGA 75/25?>?PLGA 50/50. A burst effect (17–20%) was observed during the first hour and then a period of constant release rate (3.52?±?0.82–1.46?±?0.26?µg 5-FU?h^?1 per milligram of microspheres) up to 8 or 13?h, depending on the polymer, was obtained.     

Decreasing systemic toxicity via transdermal delivery of anticancer drugs

When used at a high dose, many anticancer drugs produce undesirable side effects including hepatotoxicity. Transdermal delivery bypasses first-pass metabolism, allowing the use of a lower dose of drug while decreasing systemic toxicity. In this review, we summarize various advanced technologies for improving anticancer drug delivery via the skin. This technology is discussed in the context of three anticancer drugs, 5-fluorouracil (5-FU), methotrexate (MTX) and 5-aminolevulinic acid (5-ALA). The use of a erbium:YAG (Er:YAG) laser for transdermal delivery of anticancer drugs is specifically highlighted in this review.     

5-Fluorouracil-loaded microspheres prepared by spray-drying poly(D,L-lactide) and poly(lactide-co-glycolide) polymers: characterization and drug release

5-Fluorouracil (5-FU), a hydrosoluble anti-neoplastic drug, was encapsulated in microspheres of poly(D,L-lactide) (PLA) and poly(lactide-co-glycolide) (PLGA) polymers using the spray-drying technique, in order to obtain small size microspheres with a significant drug entrapment efficiency. Drug-loaded microspheres included between 47 +/- 11 and 67 +/- 12 microg 5-FU mg(-1) microspheres and the percentage of entrapment efficiency was between 52 +/- 12 and 74 +/- 13. Microspheres were of small size (average diameter: 0.9 +/- 0.4-1.4 +/- 0.8 microm microspheres without drug; 1.1 +/- 0.5-1.7 +/- 0.9 microm 5-FU-loaded microspheres) and their surface was smooth and slightly porous, some hollows or deformations were observed in microspheres prepared from polymers with larger Tg. A fractionation process of the raw polymer during the formation of microspheres was observed as an increase of the average molecular weight and also of Tg of the polymer of the microspheres. The presence of 5-FU did not modify the Tg values of the microspheres. Significant interactions between the drug and each one of the polymers did not take place and total release of the included drug was observed in all cases. The time needed for the total drug release (28-129 h) was in the order PLA > PLGA 75/25 > PLGA 50/50. A burst effect (17-20%) was observed during the first hour and then a period of constant release rate (3.52 +/- 0.82-1.46 +/- 0.26 microg 5-FU h(-1) per milligram of microspheres) up to 8 or 13 h, depending on the polymer, was obtained.     

Ibuprofen-loaded ethylcellulose/polystyrene microspheres: An approach to get prolonged drug release with reduced burst effect and low ethylcellulose content

The aim of this study was to develop ethylcellulose microspheres for prolonged drug delivery with reduced burst effect. Ethylcellulose microspheres loaded with ibuprofen were prepared with and without polystyrene, which was used to retard drug release from ethylcellulose microspheres. Ibuprofen-loaded ethylcellulose microspheres with a polystyrene content of 0–25% were prepared by the solvent evaporation technique and characterized by drug loading, infrared spectroscopy, differential scanning calorimetry and scanning electron microscopy. The in vitro release studies were performed to study the influence of polystyrene on ibuprofen release from ethylcellulose microspheres. The microspheres showed 28–46% of drug loading and 80–92% of entrapment, depending on polymer/drug ratio. The infrared spectrum and thermogram showed stable character of ibuprofen in the microspheres and revealed an absence of drug polymer interaction. The prepared microspheres were spherical in shape and had a size range of 0.1–4μm. Ethylcellulose/polystyrene micro-spheres showed prolonged drug release and less burst effect when compared to microspheres prepared with ethylcellulose alone. Microspheres prepared with an ethylcellulose/polystyrene ratio of 80:20 gave a required release pattern for oral drug delivery. The presence of polystyrene above this ratio gave release over 24 h. To find out the mechanism of drug release from ethylcellulose/polystyrene microspheres, the data obtained from in vitro release were fitted in various kinetic models. High correlation was obtained in Higuchi and Korsmeyer-Peppas models. The drug release from ethylcellulose/polystyrene microspheres was found to be diffusion controlled.     

Intravascular tumour targeting of aclarubicin-loaded gelatin microspheres. Preparation,biocompatibility and biodegradability

This study is to evaluate the potential use of aclarubicin-loaded gelatin microspheres as an intravascular biodegradable drug delivery system for the regional cancer therapy. The diameter of the microspheres prepared by water in oil emulsion polymerization could be controlled by adjusting the stirring rate in the range of 10–50 μm: D(in μm)=?73.8 log(rpm)+262.7. The addition of proteolytic enzyme increased the in vitro aclarubicin release, but it did not change the amount of the initial burst release which reached about 45%. Microspheres injected intravenously into the mouse tail vein embolized only to the lung when observed by fluorescence microscopy. From histological examination following injection of gelatin microspheres into mouse femoral muscle, mild inflammation was observed from the appearance of neutrophils after 2 days and rapid repair process was confirmed thereafter. Biodegradation process of gelatin microspheres lodged on the pulmonary capillary bed was followed up by microscopic observation; degradation was taking place by about 36 hrs, followed by severe damage on the spherical shape and microspheres was no longer found 10 days after injection.     

Preparation and Characterization of Oxybenzone-Loaded Gelatin Microspheres for Enhancement of Sunscreening Efficacy

The objective of our present study was to prepare and evaluate gelatin microspheres of oxybenzone to enhance its sunscreening efficacy. The gelatin microspheres of oxybenzone were prepared by emulsion method. Process parameters were analyzed to optimize the formulation. The in vitro drug release study was performed in pH 7.4 using cellulose acetate membrane. Microspheres prepared using oxybenzone:gelatin ratio of 1:6 showed slowest drug release and those prepared with oxybenzone:gelatin ratio of 1:2 showed fastest drug release. The gelatin microspheres of oxybenzone were incorporated in aloe vera gel. Sun exposure method using sodium nitroprusside solution was used for in vitro sunscreen efficacy testing. The formulation C₅ containing oxybenzone-bearing gelatin microspheres in aloe vera gel showed best sunscreen efficacy. The formulations were evaluated for skin irritation test in human volunteers, sun protection factor, and minimum erythema dose in albino rats. These studies revealed that the incorporation of sunscreening agent–loaded microspheres into aloe vera gel greatly increased the efficacy of sunscreen formulation more than four times.     

Formulation and characterization of Paclitaxel, 5-FU and Paclitaxel + 5-FU microspheres

The purpose of this study was to compare the combination (Paclitaxel + 5-FU microspheres) with a single drug chemotherapy (Paclitaxel and 5-FU microspheres) against metastatic breast cancer cell line (MDA-MB 435 S). The physicochemical characteristics of the microspheres (i.e. encapsulation efficiency, particle size distribution, in vitro release, thermal characteristics) were studied. The results demonstrated that the encapsulation efficiency of Paclitaxel was high (90%) when the drug was encapsulated in poly(lactic-co-glycolic acid) (PLGA) microparticles with or without 5-fluorouracil (5-FU). However, the encapsulation efficiency of 5-FU was low (19%) and increased to 30% when the drug was encapsulated with Paclitaxel. The mean particle size of microspheres was 2.5microm and were spherical in shape. The in vitro release of both 5-FU and Paclitaxel from the microspheres was relatively fast initially followed by a slower and more controlled release. The cytotoxic activity of Paclitaxel microspheres was far greater compared to either the microspheres containing 5-FU + Paclitaxel or 5-FU alone. Overall results demonstrated that incorporation of Paclitaxel or 5-FU in microspheres enhances the cytotoxicity in more controlled manner compared to that of free drugs and also that careful consideration should be made when combining drugs acting in different phases of cell cycle.     

Surface modification of poly (l-lactic acid) microspheres for site-specific delivery of ketoprofen for chronic inflammatory disease

The purpose of this research was to investigate the potential of surface modified Poly (l-lactic acid) (PLA) microspheres as a carrier for site-specific delivery of anti-inflammatory drug, ketoprofen, for the treatment of rheumatoid arthritis. Microspheres were prepared by solvent evaporation method using 20% w/w PLA in methylene chloride and 100 mL of a 2.5% poly vinyl alcohol (PVA) solution. Formulations were optimized for several processing parameters like drug to polymer ratio, stirring rate and volume of preparation medium etc. The surface of PLA microspheres was modified with gelatin to impart fibronectin recognition. The microspheres were characterized by surface morphology, size distribution, encapsulation efficiency, and by in vitro drug release studies. The prepared microspheres were light yellow, discrete, and spherical. Formulation with optimum drug to polymer ratio exhibited smallest vesicle size (43.02), high drug encapsulation efficiency (81.11) and better process yield (83.45). The release of drug was extended up to 24 h with Higuchi pattern of drug release. The in vivo results showed that the gelatin modified formulation reduced paw edema at greater extent than pure drug and PLA microspheres and it could be a promising carrier system for controlled and site-specific delivery of ketoprofen with possible clinical applications.     

Preparation of gelatin microspheres containing lactic acid--effect of cross-linking on drug release

In this study, gelatin microspheres containing lactic acid were prepared by the polymerization technique using glutaraldehyde as the cross-linking agent. Dried microspheres were loaded by immersing them in an aqueous solution of lactic acid. In order to prepare microspheres with an appropriate drug release profile, the effect of time of cross-linking and the amount of cross-linking agent on the swelling properties of microspheres and their release profile were investigated. The microencapsulation efficiency, microspheres appearance, particle size, swelling ratio and drug release profile were also studied. Microspheres prepared with a larger amount of cross-linking agent, or after longer cross-linking time, showed a reduced swelling ratio in aqueous media. In vitro release pattern of lactic acid from gelatin microspheres showed a biphasic profile and the release rates were reduced upon increasing the amount of cross-liking agent and prolonging the cross-linking time.     

Novel hydrogel microspheres of chitosan and pluronic F-127 for controlled release of 5-fluorouracil

Hydrogel microspheres of chitosan (CS) and Pluronic F127 (PF-127) were prepared by the emulsion-crosslinking method employing glutaraldehyde (GA) as a crosslinker. 5-Fluorouracil (5-FU), an anticancer drug with good water solubility, was encapsulated into hydrogel microspheres. Various formulations were prepared by varying the ratio of CS and PF-127, % drug loading and amount of GA. Microspheres were characterized by Fourier transform infrared (FTIR) spectroscopy to confirm the absence of chemical interactions between drug, polymer and the crosslinking agent. Scanning electron microscopy (SEM) was performed to study the surface morphology of the microspheres. SEM showed that microspheres have smooth shiny surfaces. Particle size, as measured by laser light scattering technique, gave an average size ranging from 110 to 382?µm. Differential scanning calorimetry (DSC) and X-ray diffraction (X-RD) studies were performed to understand the crystalline nature of the drug after encapsulation into hydrogel microspheres. Encapsulation of the drug up to 86% achieved was measured by UV spectroscopy. Equilibrium swelling experiments were performed in distilled water. Diffusion coefficients (D) of water through microspheres were estimated by an empirical equation. In vitro release studies indicated the dependence of release rate on the extent of crosslinking, drug loading and the amount of PF-127 used to produce the microspheres; slow release was extended up to 24?h. The release data were also fitted to an empirical equation to compute the diffusional exponent (n), which indicated that the release mechanism followed the non-Fickian trend.     

Controlled-release injectable containing Terbinafine/PLGA microspheres for Onychomycosis Treatment

Controlled-release drug delivery systems based on biodegradable polymers have been extensively evaluated for use in localized drug delivery. In the present study, intralesionally injectable poly (lactide-co-glycolide) (PLGA) microspheres for controlled release of terbinafine hydrochloride (TH) was developed for treating fungal toe/finger nail infections. TH–PLGA microspheres were formulated using O/W emulsification and modified solvent extraction/evaporation technique. Microspheres were evaluated for particle size and size distribution, encapsulation efficiency, surface, and morphology. The in vitro drug release profile was studied in aqueous media as well as in 1% agar gel. Microspheres system was also evaluated in excised cadaver toe model, and extent of TH accumulation in nail bed, nail plate, and nail matrix was measured at different time points. Microspheres were found to provide consistent and sustained TH release. Intralesional administration of controlled-release microspheres can be a potential alternative mode of treating fungus-infected toe and/or finger nails. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 103:1178–1183, 2014