- Email: [email protected]
chitosan nanoparticles by nucleation and ionic crosslinking in micro emulsions
Abstracts / Journal of Controlled Release 152 (2011) e1–e132
application in medicine for quickly diminishing inflammation and other biomedical applications, such as cellular imaging and biosensors. Acknowledgments The work was supported by Shanghai Leading Academic Disciplines (S30109), NSFC (20871081), Science and Technology Commission of Shanghai Municipality (10QH1401000, 10DZ0500100), and Shanghai Key Laboratory of Bio-Energy Crops (09DZ2271800). References [1] M. Vallet-Regi, A. Ra mila, R.P. del Real, J. Pe´rez-Pariente, A new property of MCM-41: drug delivery systemChem. Mater. 13 (2001) 308–311. [2] J. Wang, Q. Xiao, H. Zhou, P. Sun, Z. Yuan, B. Li, D. Ding, A. Shi, T. Chen, Budded, mesoporous silica hollow spheres: hierarchical structure controlled by kinetic selfassembly, Adv. Mater. 18 (2006) 3284–3288. [3] M. Grün, K.K. Unger, A. Matsumoto, K. Tsutsumi, Novel pathways for the preparation of mesoporous MCM-41 materials: control of porosity and morphology, Microporous Mesoporous Mater. 27 (1999) 207–216.
doi:10.1016/j.jconrel.2011.08.108
Preparation and characterization of aspirin/chitosan nanoparticles by nucleation and ionic crosslinking in micro emulsions Shuping Jin, Lei Feng, Xinghai Yu Key Laboratory of Resources and Environmental Chemistry of West China, Department of Chemistry, Hexi University, Zhangye 734000, China E-mail address: [email protected] (S. Jin). Abstract summary Chitosan nanoparticles loaded with aspirin (aspirin/chitosan) were prepared by a process of nucleation and ionic crosslinking in micro emulsions for medical and pharmaceutical applications. The nanoparticles were characterized by Fourier transform infrared spectroscopy, field emission scanning electron microscopy, transmission electron microscopy, dynamic laser light scattering and X-ray powder diffraction, respectively. In vitro release profiles under different conditions were determined using an UV/Vis spectrophotometer and the results showed that the nanoparticles displayed an excellent drug-controlled release behavior. Keywords: Chitosan, Nanoparticle, Nucleation in micro emulsion, Controlled drug release, Conformation
Introduction Aspirin is an old drug, which falls into the category of nonsteroidal anti-inflammatory drugs and is used extensively as a painkiller. In recent years, studies showed that aspirin can reduce the risk of heart attack, stroke and colorectal cancer [1]. Chitosan nanoparticles have been widely investigated as protein delivery systems [2–4]. Many approaches have been developed to prepare chitosan beads including the water in oil method, the emulsiondroplet coalescence technique and spray drying process. The solution behavior of chitosan has been studied extensively. The gross conformation of chitosan in solution may be a spherical shape, random coil, and rod shape, which can be manipulated by two types of parameters: structure parameters such as molecular weight and degree of acetylation, and solution parameters such as ionic strength, solvent, temperature and pH of solution [5]. The aim of this study was to prepare chitosan nanoparticles loaded with aspirin by exploiting the change in conformation of chitosan in aqueous solutions with pH. Experimental methods Aspirin-loaded chitosan nanoparticles were obtained by the process described as follows: a certain amount of chitosan was added into acetic acid aqueous solution (2%, w/v) under stirring till the mixture changed into a solution. Certain amounts of Span-80 and Tween-80 were dispersed into the above solution and stirred for 0.5 h to obtain a stable, transparent and homogeneous micro emulsion. Aspirin powder was added into the above emulsion. The pH of the system was adjusted by adding 1 M NaOH aqueous solution under stirring till pH 7.2, which was measured using a PHS-3B model pH meter. Finally, trisodium citrate was added and the mixture was stirred for 3 h. Results and discussion Fig. 1 displays the FTIR spectra of chitosan, aspirin and aspirin/ chitosan. The absorptions appearing at about 1625–1580, 1590–1570, 1525–1470 and 1460–1440 cm− 1 (spectrum b) are usually used for characterizing the presence of benzene. In the spectrum c and inset, two small shoulders appearing at about 1481.92 cm − 1 and 1463.67 cm− 1 correspond to the CC stretching vibration of the benzene residue, which indicates that aspirin was loaded into the chitosan nanoparticles. It is also evident that a weak absorption appearing at 754.96 cm− 1 corresponds to the stretching vibration of four adjoining CH bonds from benzene.
c Transmittance (%)
Fig. 2. Controlled release kinetics of IBU-encapsulated at different pH.
e39
b
a
3500
3000
2500
2000
1500
1000
500
Wavenumbers (cm-1) Fig. 1. FTIR spectra of the chitosan (a), aspirin (b) and aspirin/chitosan (c).
The phase transition mechanism of chitosan in aqueous solution driven by pH is proposed as shown in Scheme 1. Initially, at low pH, protonation of amine groups (NH3+) on chitosan brings about strong
e40
Abstracts / Journal of Controlled Release 152 (2011) e1–e132
electrostatic repulsion along the chitosan chain resulting in a random coil (Scheme 1a). At high pH, the disappearance of NH3+ accompanied by a formation of hydrogen bonds and hydrophobic aggregation results in a change of the conformation of chitosan from a loose coil to a hyper-coiled form (Scheme 1b) and nanoparticles form. Finally, a certain amount of crosslinker would bring about a more compact globular conformation (Scheme 1c). The formation of nanoparticles in pH 7.2 aqueous solution can be investigated by field emission scanning electron microscopy, transmission electron microscopy and dynamic laser light scattering, and the results are shown in Figs. 2, 3 and 4 respectively. The aggregation shown in Fig. 2 may be attributed to higher concentrations after centrifugation. Fig. 4 displays the uniform distribution of nanoparticle diameters, which was measured at about 88 nm.
Fig. 5 displays the in vitro release behavior of aspirin from aspirin/ chitosan nanoparticles in saline, a glucose solution for injection and deionized water at 37 °C, respectively. As shown in this figure, the release rate in saline is faster than that in glucose solution and deionized water. This may be attributed to NaCl, which would destroy the hydration sheath surrounding the nanoparticle and then penetrating, from the solution to the particle. Electrostatic attraction between NH3+ and COO− would be partly replaced by Na+ and COO− resulting in a decrease in the degree of crosslinking of chitosan. Hydrogen bonds among chitosan chains would be destroyed markedly resulting in a change of the conformation of chitosan from a compact globular conformation to a loose coil with subsequent release of aspirin.
a
b
Intensity
100
c
Scheme 1. The molecular conformations of chitosan (a: Random coil model in acid solution; b: Globular model in pH 7.2 solution; c: Compact globular model after adding crosslinker).
50
0 50
100
Diameter (nm) Fig. 4. The diameter distribution of aspirin/ chitosan in deionized water after ultrasonic dispersion.
100 90 80
Freleased (%)
70 60 50 40 30 Fig. 2. FESEM image of crosslinked aspirin/chitosan in pH 7.2 solution.
20 10 0 -50
0
50
100 150 200 250 300 350 400
Time (min) Fig. 5. The release profiles of aspirin in saline (○), glucose solution for injection (◊) and deionized water (+) from aspirin/ chitosan at 37 °C.
Conclusion Chitosan nanoparticles loaded with aspirin were prepared by a process of nucleation and ionic crosslinking in a micro emulsion and displayed an excellent controlled drug release behavior in glucose solution.
100 nm Fig. 3. TEM image of crosslinked aspirin/chitosan in dilute aqueous solution.
The measurements of X-ray powder diffraction indicate that the change in the crystalline structure of chitosan did not occur during the nucleation process. However the decrease in crystallinity of aspirin is dramatic and it is almost amorphous in the nanoparticle.
Acknowledgment This article was supported by President Fund KJ2009008 from Hexi University. References [1] J.F. Neault, M. Naoui, M. Manfait, H.A. Tajmir-Riahi, Aspirin–DNA interaction studied by FTIR and laser Raman difference spectroscopyFEBS Lett. 382 (1996) 26–30. [2] S.T. Lim, G.P. Martin, D.J. Berry, M.B. Brown, Preparation and evaluation of the in vitro drug release properties and mucoadhesion of novel microspheres of hyaluronic acid and chitosan, J. Control. Release 66 (2000) 281–292. [3] K.H. Min, K. Park, Y.S. Kim, S.M. Bae, S. Lee, H. Gon Jo, R.W. Park, I.S. Kim, S.Y. Jeong, K. Kim, I.C. Kwon, Hydrophobically modified glycol chitosan nanoparticles-encapsulated
Abstracts / Journal of Controlled Release 152 (2011) e1–e132
60 40 20
-20
2 D
D
1
0 C
Introduction Intracranial glioma is one of the most serious malignant tumors. It takes 44% among all intracranial tumors. Its clinical treatment is surgery plus chemotherapy and radiotherapy. In order to protect the brain, only part of the glioma tumor bed can be cut off. Therefore it always recurs in a few months and has high mortality rate. The efficacy of chemotherapy to glioma is very low because very few drugs can pass the blood–brain barrier (BBB). Therefore, exploring a new therapy for malignant glioma is one of the challenges in the field of neurosurgery. Paclitaxel (PTX) is a well-known anti-cancer agent, suitable for the ovary, breast, lung, head and neck, esophagus, bladder, endometrium, testis, and other malignant tumors. But it is seldom used for glioma because of its extremely low water solubility, serious allergic reactions, and problems for passing the BBB. Our group has long been interested in developing functional biodegradable polymers for anticancer drug delivery [1–6]. Anticancer drug molecules (e.g., PTX) are attached to an amphiphilic block copolymer (e.g., PEG-PLA) to form a polymer–drug conjugate (e.g., PEG–PLA–PTX). Because of the amphiphilicity of the conjugate, after self-assembling in aqueous medium, the anticancer drug PTX is trapped in the core part of the micelles formed and gets well protected, and the PEG segments constitute the corona part of the micelles and remain soluble in water. Therefore, conjugate micelles are water soluble after freeze-drying and can be injected as common
80
B
Keywords: C6 glioma, Wistar rats, Paclitaxel, Pro-drug, EPR effect, BBB
a
A
Abstract summary An intracranial C6 glioma model was constructed using Wistar rats. Paclitaxel (PTX) was conjugated to biodegradable diblock copolymer PEG–PLA to form water-soluble PEG–PLA–PTX micelles. The tumor-bearing rats were treated with blank PEG–PLA micelles, commercial Taxol, and PEG–PLA–PTX micelles, respectively, via tail vein injection. With the same equivalent PTX dose, the PEG–PLA–PTX micelles displayed enhanced inhibition ability to tumor growth as shown by the body weight change, survival time and tumor image size. This improved therapeutic effect was ascribed to the EPR effect of the PEG–PLA–PTX micelles. Fluorescent imaging of the brain slice further confirmed that the PEG–PLA–PTX micelles can pass the BBB and remain in the brain even 5 days post drug administration.
Cell culture and animal models. C6 glioma cells were first cultured and grown in RPMI1640 containing 10% FBS, 0.03% l-glutamine, 100 units/mL penicillin, and 100 μg/mL streptomycin in 5% CO2 at 37 ° C. Test animals were Wistar male rats, 20 rats for comparison and 50 for constructing tumor models. The tumor was inoculated into the intracranial part of the Wistar rats with the help of a Universal Stereotaxic Instrument from Japan to ensure desired position (1 mm in front of the coronal suture and 3 mm right to the sagittal suture) and depth (from 5 to 6.5 mm under the skull surface, diameter
F
Zhanfeng Wang1,2,a, Xiuli Hu1, Jun Yue1,3, Xiabin Jing1 State Key Laboratory of Polymer Physics and Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China 2 The First Hospital of Jilin University, Changchun 130021, China 3 Graduate School of Chinese Academy of Sciences, Beijing 100049, China a Present address: Affiliate Hospital of Beihua University, Jilin 132011, China. E-mail address: [email protected] (X. Jing). 1
E
Experimental study on biodegradable polymer–paclitaxel conjugate micelles for chemotherapy of C6 glioma
Experimental methods Preparation of PEG–PLA–PTX conjugates and micelles. Diblock copolymer PEG–PLA was used for paclitaxel conjugation. The terminal hydroxyl groups of PEG–PLA were first converted to carboxyl groups by reacting with malic anhydride. PTX was attached to the PLA end via the ester formation between this end COOH group and the OH group on PTX. By changing the block lengths of PEG and PLA, the paclitaxel content in the conjugates can be adjusted in a range of 8– 15 wt.%. The sample used in the present study had a composition of PEG5000–PLA2200–PTX850 and a PTX content of ca. 10 wt.%. The conjugate micelles were prepared by emulsification plus solvent evaporation, followed by freeze-drying. The micelles obtained were of spherical shape and had an average size of ca. 62 nm.
Change of body weight (g/week)
doi:10.1016/j.jconrel.2011.08.109
water-based formulations. In the present study, PEG–PLA–PTX micelles are used to treat C6 glioma in rat models to examine their efficacy to glioma, paying special attention to the possibility of the conjugate micelles passing through the BBB.
Different experimental groups
b
Mean survivial time of tumor mice (days)
camptothecin enhance the drug stability and tumor targeting in cancer therapy, J. Control. Release 127 (2008) 208–218. [4] Y. Zhang, M.R. Huo, J.P. Zhou, D. Yu, Y.P. Wu, Potential of amphiphilically modified low molecular weight chitosan as a novel carrier for hydrophobic anticancer drug: synthesis, characterization, micellization and cytotoxicity evaluation, Carbohydr. Polym. 77 (2009) 231–238. [5] Q. Gao, A.J.W., Effects of molecular weight, degree of acetylation and ionic strength on surface tension of chitosan in dilute solution, Carbohydr. Polym. 64 (2006) 29–36.
e41
60 50 40 30 20 10 0 A
B
C
D1
D2
Different experimental groups
Fig. 1. Change of rat body weight (a) and mean survival time (b) of tumor-bearing rats after various treatments: A. control for tumor-bearing rats; B. blank micelles group (180 mg/kg); C. Taxol group (20 mg/kg); D1. PTX-micelle group (eq. 10 mg/kg); D2. PTX-micelle group (eq. 20 mg/kg); E. healthy rats; F. healthy rats injected with the blank micelles.
application in medicine for quickly diminishing inflammation and other biomedical applications, such as cellular imaging and biosensors. Acknowledgments The work was supported by Shanghai Leading Academic Disciplines (S30109), NSFC (20871081), Science and Technology Commission of Shanghai Municipality (10QH1401000, 10DZ0500100), and Shanghai Key Laboratory of Bio-Energy Crops (09DZ2271800). References [1] M. Vallet-Regi, A. Ra mila, R.P. del Real, J. Pe´rez-Pariente, A new property of MCM-41: drug delivery systemChem. Mater. 13 (2001) 308–311. [2] J. Wang, Q. Xiao, H. Zhou, P. Sun, Z. Yuan, B. Li, D. Ding, A. Shi, T. Chen, Budded, mesoporous silica hollow spheres: hierarchical structure controlled by kinetic selfassembly, Adv. Mater. 18 (2006) 3284–3288. [3] M. Grün, K.K. Unger, A. Matsumoto, K. Tsutsumi, Novel pathways for the preparation of mesoporous MCM-41 materials: control of porosity and morphology, Microporous Mesoporous Mater. 27 (1999) 207–216.
doi:10.1016/j.jconrel.2011.08.108
Preparation and characterization of aspirin/chitosan nanoparticles by nucleation and ionic crosslinking in micro emulsions Shuping Jin, Lei Feng, Xinghai Yu Key Laboratory of Resources and Environmental Chemistry of West China, Department of Chemistry, Hexi University, Zhangye 734000, China E-mail address: [email protected] (S. Jin). Abstract summary Chitosan nanoparticles loaded with aspirin (aspirin/chitosan) were prepared by a process of nucleation and ionic crosslinking in micro emulsions for medical and pharmaceutical applications. The nanoparticles were characterized by Fourier transform infrared spectroscopy, field emission scanning electron microscopy, transmission electron microscopy, dynamic laser light scattering and X-ray powder diffraction, respectively. In vitro release profiles under different conditions were determined using an UV/Vis spectrophotometer and the results showed that the nanoparticles displayed an excellent drug-controlled release behavior. Keywords: Chitosan, Nanoparticle, Nucleation in micro emulsion, Controlled drug release, Conformation
Introduction Aspirin is an old drug, which falls into the category of nonsteroidal anti-inflammatory drugs and is used extensively as a painkiller. In recent years, studies showed that aspirin can reduce the risk of heart attack, stroke and colorectal cancer [1]. Chitosan nanoparticles have been widely investigated as protein delivery systems [2–4]. Many approaches have been developed to prepare chitosan beads including the water in oil method, the emulsiondroplet coalescence technique and spray drying process. The solution behavior of chitosan has been studied extensively. The gross conformation of chitosan in solution may be a spherical shape, random coil, and rod shape, which can be manipulated by two types of parameters: structure parameters such as molecular weight and degree of acetylation, and solution parameters such as ionic strength, solvent, temperature and pH of solution [5]. The aim of this study was to prepare chitosan nanoparticles loaded with aspirin by exploiting the change in conformation of chitosan in aqueous solutions with pH. Experimental methods Aspirin-loaded chitosan nanoparticles were obtained by the process described as follows: a certain amount of chitosan was added into acetic acid aqueous solution (2%, w/v) under stirring till the mixture changed into a solution. Certain amounts of Span-80 and Tween-80 were dispersed into the above solution and stirred for 0.5 h to obtain a stable, transparent and homogeneous micro emulsion. Aspirin powder was added into the above emulsion. The pH of the system was adjusted by adding 1 M NaOH aqueous solution under stirring till pH 7.2, which was measured using a PHS-3B model pH meter. Finally, trisodium citrate was added and the mixture was stirred for 3 h. Results and discussion Fig. 1 displays the FTIR spectra of chitosan, aspirin and aspirin/ chitosan. The absorptions appearing at about 1625–1580, 1590–1570, 1525–1470 and 1460–1440 cm− 1 (spectrum b) are usually used for characterizing the presence of benzene. In the spectrum c and inset, two small shoulders appearing at about 1481.92 cm − 1 and 1463.67 cm− 1 correspond to the CC stretching vibration of the benzene residue, which indicates that aspirin was loaded into the chitosan nanoparticles. It is also evident that a weak absorption appearing at 754.96 cm− 1 corresponds to the stretching vibration of four adjoining CH bonds from benzene.
c Transmittance (%)
Fig. 2. Controlled release kinetics of IBU-encapsulated at different pH.
e39
b
a
3500
3000
2500
2000
1500
1000
500
Wavenumbers (cm-1) Fig. 1. FTIR spectra of the chitosan (a), aspirin (b) and aspirin/chitosan (c).
The phase transition mechanism of chitosan in aqueous solution driven by pH is proposed as shown in Scheme 1. Initially, at low pH, protonation of amine groups (NH3+) on chitosan brings about strong
e40
Abstracts / Journal of Controlled Release 152 (2011) e1–e132
electrostatic repulsion along the chitosan chain resulting in a random coil (Scheme 1a). At high pH, the disappearance of NH3+ accompanied by a formation of hydrogen bonds and hydrophobic aggregation results in a change of the conformation of chitosan from a loose coil to a hyper-coiled form (Scheme 1b) and nanoparticles form. Finally, a certain amount of crosslinker would bring about a more compact globular conformation (Scheme 1c). The formation of nanoparticles in pH 7.2 aqueous solution can be investigated by field emission scanning electron microscopy, transmission electron microscopy and dynamic laser light scattering, and the results are shown in Figs. 2, 3 and 4 respectively. The aggregation shown in Fig. 2 may be attributed to higher concentrations after centrifugation. Fig. 4 displays the uniform distribution of nanoparticle diameters, which was measured at about 88 nm.
Fig. 5 displays the in vitro release behavior of aspirin from aspirin/ chitosan nanoparticles in saline, a glucose solution for injection and deionized water at 37 °C, respectively. As shown in this figure, the release rate in saline is faster than that in glucose solution and deionized water. This may be attributed to NaCl, which would destroy the hydration sheath surrounding the nanoparticle and then penetrating, from the solution to the particle. Electrostatic attraction between NH3+ and COO− would be partly replaced by Na+ and COO− resulting in a decrease in the degree of crosslinking of chitosan. Hydrogen bonds among chitosan chains would be destroyed markedly resulting in a change of the conformation of chitosan from a compact globular conformation to a loose coil with subsequent release of aspirin.
a
b
Intensity
100
c
Scheme 1. The molecular conformations of chitosan (a: Random coil model in acid solution; b: Globular model in pH 7.2 solution; c: Compact globular model after adding crosslinker).
50
0 50
100
Diameter (nm) Fig. 4. The diameter distribution of aspirin/ chitosan in deionized water after ultrasonic dispersion.
100 90 80
Freleased (%)
70 60 50 40 30 Fig. 2. FESEM image of crosslinked aspirin/chitosan in pH 7.2 solution.
20 10 0 -50
0
50
100 150 200 250 300 350 400
Time (min) Fig. 5. The release profiles of aspirin in saline (○), glucose solution for injection (◊) and deionized water (+) from aspirin/ chitosan at 37 °C.
Conclusion Chitosan nanoparticles loaded with aspirin were prepared by a process of nucleation and ionic crosslinking in a micro emulsion and displayed an excellent controlled drug release behavior in glucose solution.
100 nm Fig. 3. TEM image of crosslinked aspirin/chitosan in dilute aqueous solution.
The measurements of X-ray powder diffraction indicate that the change in the crystalline structure of chitosan did not occur during the nucleation process. However the decrease in crystallinity of aspirin is dramatic and it is almost amorphous in the nanoparticle.
Acknowledgment This article was supported by President Fund KJ2009008 from Hexi University. References [1] J.F. Neault, M. Naoui, M. Manfait, H.A. Tajmir-Riahi, Aspirin–DNA interaction studied by FTIR and laser Raman difference spectroscopyFEBS Lett. 382 (1996) 26–30. [2] S.T. Lim, G.P. Martin, D.J. Berry, M.B. Brown, Preparation and evaluation of the in vitro drug release properties and mucoadhesion of novel microspheres of hyaluronic acid and chitosan, J. Control. Release 66 (2000) 281–292. [3] K.H. Min, K. Park, Y.S. Kim, S.M. Bae, S. Lee, H. Gon Jo, R.W. Park, I.S. Kim, S.Y. Jeong, K. Kim, I.C. Kwon, Hydrophobically modified glycol chitosan nanoparticles-encapsulated
Abstracts / Journal of Controlled Release 152 (2011) e1–e132
60 40 20
-20
2 D
D
1
0 C
Introduction Intracranial glioma is one of the most serious malignant tumors. It takes 44% among all intracranial tumors. Its clinical treatment is surgery plus chemotherapy and radiotherapy. In order to protect the brain, only part of the glioma tumor bed can be cut off. Therefore it always recurs in a few months and has high mortality rate. The efficacy of chemotherapy to glioma is very low because very few drugs can pass the blood–brain barrier (BBB). Therefore, exploring a new therapy for malignant glioma is one of the challenges in the field of neurosurgery. Paclitaxel (PTX) is a well-known anti-cancer agent, suitable for the ovary, breast, lung, head and neck, esophagus, bladder, endometrium, testis, and other malignant tumors. But it is seldom used for glioma because of its extremely low water solubility, serious allergic reactions, and problems for passing the BBB. Our group has long been interested in developing functional biodegradable polymers for anticancer drug delivery [1–6]. Anticancer drug molecules (e.g., PTX) are attached to an amphiphilic block copolymer (e.g., PEG-PLA) to form a polymer–drug conjugate (e.g., PEG–PLA–PTX). Because of the amphiphilicity of the conjugate, after self-assembling in aqueous medium, the anticancer drug PTX is trapped in the core part of the micelles formed and gets well protected, and the PEG segments constitute the corona part of the micelles and remain soluble in water. Therefore, conjugate micelles are water soluble after freeze-drying and can be injected as common
80
B
Keywords: C6 glioma, Wistar rats, Paclitaxel, Pro-drug, EPR effect, BBB
a
A
Abstract summary An intracranial C6 glioma model was constructed using Wistar rats. Paclitaxel (PTX) was conjugated to biodegradable diblock copolymer PEG–PLA to form water-soluble PEG–PLA–PTX micelles. The tumor-bearing rats were treated with blank PEG–PLA micelles, commercial Taxol, and PEG–PLA–PTX micelles, respectively, via tail vein injection. With the same equivalent PTX dose, the PEG–PLA–PTX micelles displayed enhanced inhibition ability to tumor growth as shown by the body weight change, survival time and tumor image size. This improved therapeutic effect was ascribed to the EPR effect of the PEG–PLA–PTX micelles. Fluorescent imaging of the brain slice further confirmed that the PEG–PLA–PTX micelles can pass the BBB and remain in the brain even 5 days post drug administration.
Cell culture and animal models. C6 glioma cells were first cultured and grown in RPMI1640 containing 10% FBS, 0.03% l-glutamine, 100 units/mL penicillin, and 100 μg/mL streptomycin in 5% CO2 at 37 ° C. Test animals were Wistar male rats, 20 rats for comparison and 50 for constructing tumor models. The tumor was inoculated into the intracranial part of the Wistar rats with the help of a Universal Stereotaxic Instrument from Japan to ensure desired position (1 mm in front of the coronal suture and 3 mm right to the sagittal suture) and depth (from 5 to 6.5 mm under the skull surface, diameter
F
Zhanfeng Wang1,2,a, Xiuli Hu1, Jun Yue1,3, Xiabin Jing1 State Key Laboratory of Polymer Physics and Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China 2 The First Hospital of Jilin University, Changchun 130021, China 3 Graduate School of Chinese Academy of Sciences, Beijing 100049, China a Present address: Affiliate Hospital of Beihua University, Jilin 132011, China. E-mail address: [email protected] (X. Jing). 1
E
Experimental study on biodegradable polymer–paclitaxel conjugate micelles for chemotherapy of C6 glioma
Experimental methods Preparation of PEG–PLA–PTX conjugates and micelles. Diblock copolymer PEG–PLA was used for paclitaxel conjugation. The terminal hydroxyl groups of PEG–PLA were first converted to carboxyl groups by reacting with malic anhydride. PTX was attached to the PLA end via the ester formation between this end COOH group and the OH group on PTX. By changing the block lengths of PEG and PLA, the paclitaxel content in the conjugates can be adjusted in a range of 8– 15 wt.%. The sample used in the present study had a composition of PEG5000–PLA2200–PTX850 and a PTX content of ca. 10 wt.%. The conjugate micelles were prepared by emulsification plus solvent evaporation, followed by freeze-drying. The micelles obtained were of spherical shape and had an average size of ca. 62 nm.
Change of body weight (g/week)
doi:10.1016/j.jconrel.2011.08.109
water-based formulations. In the present study, PEG–PLA–PTX micelles are used to treat C6 glioma in rat models to examine their efficacy to glioma, paying special attention to the possibility of the conjugate micelles passing through the BBB.
Different experimental groups
b
Mean survivial time of tumor mice (days)
camptothecin enhance the drug stability and tumor targeting in cancer therapy, J. Control. Release 127 (2008) 208–218. [4] Y. Zhang, M.R. Huo, J.P. Zhou, D. Yu, Y.P. Wu, Potential of amphiphilically modified low molecular weight chitosan as a novel carrier for hydrophobic anticancer drug: synthesis, characterization, micellization and cytotoxicity evaluation, Carbohydr. Polym. 77 (2009) 231–238. [5] Q. Gao, A.J.W., Effects of molecular weight, degree of acetylation and ionic strength on surface tension of chitosan in dilute solution, Carbohydr. Polym. 64 (2006) 29–36.
e41
60 50 40 30 20 10 0 A
B
C
D1
D2
Different experimental groups
Fig. 1. Change of rat body weight (a) and mean survival time (b) of tumor-bearing rats after various treatments: A. control for tumor-bearing rats; B. blank micelles group (180 mg/kg); C. Taxol group (20 mg/kg); D1. PTX-micelle group (eq. 10 mg/kg); D2. PTX-micelle group (eq. 20 mg/kg); E. healthy rats; F. healthy rats injected with the blank micelles.