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Cite this: Photochem. Photobiol. Sci., 2012, 11, 1285
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β-Cyclodextrin polymer nanoparticles as carriers for doxorubicin and artemisinin: a spectroscopic and photophysical study† Resmi Anand,a Francesco Manoli,a Ilse Manet,a Samia Daoud-Mahammed,b Valentina Agostoni,b Ruxandra Grefb and Sandra Monti*a
Published on 11 May 2012. Downloaded on 16/09/2014 06:30:20.
Received 20th January 2012, Accepted 2nd April 2012 DOI: 10.1039/c2pp25014d
The association of doxorubicin (DOX) and artemisinin (ART) to a β-CyD-epichlorohydrin crosslinked polymer ( pβ-CyD), organized in nanoparticles of ca. 15 nm size, was investigated in neutral aqueous medium by circular dichroism (CD), UV-vis absorption and fluorescence. The stability constants and the absolute CD spectra of the drug complexes were determined by global analysis of multiwavelength data from spectroscopic titrations. The polymer pβ-CyD proved able to disrupt the DOX dimer when the latter is the predominant form of DOX in solution. The spectroscopic and photophysical properties of the complexes evidenced an alcohol-like environment for ART and an improved inherent emission ability for DOX in the nanoparticle frame.
1.
Introduction
Cyclodextrins (CyDs) are water soluble, biocompatible cyclic oligosaccharides, made of α-D-glucopyranose units joined by α(1–4) linkages. They have received attention as potential carriers/solubilizers for drugs due to their hydrophobic cavity, which is able to host lipophilic guests.1 Various CyD-based nanoassemblies, loading drugs via non covalent interactions2 or labile covalent bonds,3 have been synthesized and proposed as delivery platforms in preclinical studies. In this framework, β-CyD polymers ( pβ-CyD) spontaneously forming nanoparticles in aqueous solutions, have recently been developed.4,5 They dramatically enhance the apparent solubility of several guests, compared to natural CyDs, validating the significant interest they have received in the drug delivery field. In this paper, we present an investigation of an epichlorohydrin crosslinked β-CyD polymer of high molecular weight6 as a carrier for two important drugs (Scheme 1), both with poor water solubility: doxorubicin (DOX) and artemisinin (ART). The first molecule is a powerful anticancer agent, usually employed in the treatment of leukaemia and various solid tumors, which is experiencing serious problems due to multidrug resistance phenomena observed for several cancer cell lines.7 The second is a sesquiterpene endoperoxide lactone, parent term of the trioxane drug family, the most effective class of drugs against multidrug resistant forms of Plasmodium falciparum nowadays used to treat malaria.8,9 For ART derivatives, an anticancer activity has also been reported.10 a
Istituto per la Sintesi Organica e la Fotoreattività, ISOF-CNR, via P. Gobetti 101, 40129 Bologna, Italy. E-mail:
[email protected]; Tel: +390516399813 b Université Paris-Sud, UMR CNRS 8612, 92296 Chatenay Malabry, France † Electronic supplementary information (ESI) available: See DOI: 10.1039/c2pp25014d
Scheme 1
(A) Doxorubicin (DOX), (B) artemisin (ART).
CyD derivatives as potential carriers for DOX have been explored since the 1990s. They have revealed an ability to improve the activity of DOX on both sensitive and multidrugresistant cancer cell lines,11 and to promote drug release to brain tissues.12 In addition, saccharide-conjugated CyDs13 and various CyD-containing nanoassemblies14 have been shown to possess great potential as delivery media. One of the goals of using CyDs is to avoid DOX self-association in aqueous medium. This tendency, which has been clearly evidenced by UV-vis absorption,15 circular dichroism (CD)16 and NMR,17 is clearly detrimental for the drug pharmacological action, because the monomer plays a major antitumoral role whereas the dimer has no efficacy by itself.18 Of the native CyDs, only γ-CyD forms inclusion complexes of significant stability with DOX.19,20 Recently, we have shown that γ-CyD cannot disrupt the drug dimer when it is the predominant form in solution.21 CyDs have also proved to be potentially useful for ART delivery.22 Scarce solubility in aqueous medium resulting in poor drug absorption upon oral administration, is indeed a serious drawback for the use of ART. This problem has been faced with the drug functionalization (e.g. substitution of the carbonyl group with a more hydrophylic group like in artemether and arteether),8 but an
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Photochem. Photobiol. Sci., 2012, 11, 1285–1292 | 1285
Published on 11 May 2012. Downloaded on 16/09/2014 06:30:20.
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additional strategy presently being pursued is the development of suitable nanocarriers, able to contrast the fast drug clearance from the body.23 In this context, we have investigated the interaction of ART with β-CyD using CD and molecular modelling to show that the peroxidic group is well outside the cavity, thus the carrier does not interfere with the pharmacological action.24,25 For the evaluation of pβ-CyD as a delivery platform for DOX and ART we have investigated the incorporation of the drugs within the polymer nanoparticles, performing accurate CD and UV-vis absorption titrations and fluorescence measurements. We have evidenced the association of DOX and ART to the β-CyD units of the polymer network and determined apparent binding constants by global analysis of multiwavelength CD data. Despite the apparent low stability of the complexes, both drugs are incorporated in the pβ-CyD-epichlorohydrin crosslinked polymer—present in water as tiny nanoparticles—where they experience a hydrophobic environment. For bound DOX, the CD spectrum revealed the dimer is disrupted in the entrapment process. These observations allow us to envisage a potential application of the β-CyD-epichlorohydrin crosslinked polymer for the delivery of DOX and ART.
2.
Experimental
2.1 Materials
Doxorubicin (DOX, Adriamycin), from ALEXIS Biochemicals, artemisinin (ART) from Aldrich and β-CyD (Serva) were used as received. Water was purified by passage through a Millipore MilliQ system. 0.01 M Phosphate or 0.01 M Tris buffer at pH 7.4 was used. DOX was easily dissolved at a concentration of 2 × 10−4 M in these media. The polymer pβ-CyD was prepared by crosslinking β-CyD with epichlorohydrin according to a procedure already described.6 The β-CyD content of the polymer was 77% w/w as determined by NMR investigations, and the average molecular weight determined by aqueous gel permeation chromatography was 2.1 × 105 g mol−1 (see SI-1†). This material is highly soluble in water (more than 100 mg mL−1) where it exists in the form of nanoparticles of
