ВЫСОКОМОЛЕКУЛЯРНЫЕ СОЕДИНЕНИЯ, Серия А, 2009, том 51, № 6, с. 1033-1042
УДК 541(64+49):532.72
BLOCK IONOMER COMPLEX MICELLES WITH CROSS-LINKED CORES FOR DRUG DELIVERY1
© 2009 г. Jong Oh Kim, Nataliya V. Nukolova, Hardeep S. Oberoi, Alexander V. Kabanov, and Tatiana K. Bronich
Department of Pharmaceutical Sciences and Center for Drug Delivery and Nanomedicine, College of Pharmacy, University of Nebraska Medical Center, 985830 Nebraska Medical Center, Omaha, NE 68198-5830, USA E-mail: akabanovr@unmc.edu; tbronich@unmc.edu
Abstract—Soft polymeric nanomaterials were synthesized by template-assisted method involving condensation of the poly(ethylene oxide)-b-polycarboxylate anions by metal ions into core-shell block ionomer complex micelles followed by chemical cross-linking of the polyion chains in the micelle cores. The resulting materials represent nanogels and are capable of swelling in a pH-dependent manner. The structural determinants that guide the self-assembly of the initial micelle templates and the swelling behavior of the cross-linked micelles include the block ionomer structure, the chemical nature of metal ions, the structure of the cross-links and the degree of cross-linking. The application of these materials for loading and release of a drug, cisplatin, is evaluated. These cross-linked block ionomer micelles have promise for delivery of pharmaceutical agents.
INTRODUCTION
Nanoscale polymeric particles are emerging as novel drug delivery systems in biomedical applications. In particular, self-assembled block copolymer micelles have been utilized in pharmaceutics for development of novel therapeutic [1] and diagnostic modalities [2]. Advantages of the polymer micelles include their small size, long circulation in the bloodstream, ability to circumvent renal excretion and extravasation at sites of enhanced vascular permeability. Recently nanofabrication of polymer micelles was significantly advanced by employing charge driven self-assembly of block copolymers containing ionic and nonionic blocks ("block ionomers"). The idea of using the block ionomers for design of new nanocom-posite materials was stimulated by discussions that took place in 1994 between Viktor Kabanov (Moscow State University), Adi Eisenberg (McGill University), and the one of the authors of this paper, Alexander Ka-banov. This was inspired by the merge of the knowledge and expertise in structural organization and solution behavior of interpolyectrolyte complexes [3], self-assembly phenomena in the solutions of ionic block copolymers [4], and the potential application of such materials for drug and gene delivery [1]. During the last fourteen years diverse materials were synthesized by reacting the block ionomers with oppositely charged molecules such as synthetic linear polyelec-trolytes [5, 6] or block ionomers [7, 8], surfac-tants/lipids [9-15], proteins [16-18], or DNA [19,
1This work was supported by the grants from U.S.A. National Institute of Health CA116590 (T.B.), National Science Foundation DMR-0513699 (A.V.K.) and (T.B.) and Department of Defense USAMRMC 06108004 (A.VK.).
20]. The driving force for such binding is the release of low molecular mass counterions, originally associated with the components of the complexes, into the external media, which is accompanied by a substantial entropy gain. These complexes belong to the special classes of nanostructured materials combining the properties of cooperative polyelectrolyte complexes [3] and amphiphilic block copolymers [21, 22]. These materials are called "block ionomer complexes (BIC)" [5] or "polyion complex micelles" [7]. BIC containing nonionic water-soluble block, such as poly(ethylene oxide), PEO, form stable aqueous dispersions of ~10 to 100 nm diameter particles with core-shell architecture even upon complete neutral-
Fig. 1. BIC compositions and morphologies comprise a broad and series of complexes for incorporating non-natural and natural charged molecules including: (a) synthetic polyions, (b) ionic surfactants, (c) metal ions, (d) DNA, (e) RNA, (f) proteins/protein complexes, (g) low molecular mass peptides and (h) pharmaceutical drugs.
Table 1. Physicochemical characteristics of block copolymers
Block copolymer0 Mw x 10-3 MJM„ Weight fraction of PEO blocks (fEo)a
PEO(170)-£-PMA(180) 23.0 1.45 0.33
PEO(125)-£-PMA(180) 21.0 1.16 0.26
PEO(114)-£-PMA(81) 12.0 1.15 0.42
PEO(114)-£-PAA(93) 11.7 1.20 0.43
PEO(80)-£-PAA(104) 11.0 1.25 0.32
a The average number of monomer units in PEO and polyacid blocks and content of PEO blocks in block copolymers were calculated using the average molecular weights provided by manufacturer.
ization of charges (Figure 1). The core can comprise polyion/polyion (a), polyion/surfactant (b), or poly-ion/metal (c) complexes. The core of such BIC can also incorporate a variety of compounds or even particles including biologically active molecules through a combination of electrostatic, hydrophobic, and hydrogen bonding interactions. Such materials are uniquely suited for the delivery of biomacromolecules [23-26], as well as ionic and non-ionic drugs [27—31]. Ionic block lengths, charge density, and ionic strength of the solution affect the formation of stable BIC and, therefore, control the amount of the drug that can be incorporated within the micelles. BIC display transitions induced by changes in pH, salt concentration, chemical nature of low molecular mass counterions as well as temperature, and can be fine-tuned to respond to environmental changes occurring in a very wide range of conditions that could realize during delivery of biological and imaging agents.
Unique features of BIC are also relevant for their use as nanoreactors in the diverse fields of medical and biological engineering. Recently, we proposed to use BIC as micellar templates to synthesize novel polymer micelles with cross-linked ionic cores [32, 33]. Indeed, the cores of the BIC formed between PEO-&-polymethacrylate anions (PEO-^-PMA) and divalent metal cations (Fig. 1c) were utilized as nanoreactors for cross-linking reactions. Resulting particles were entirely hydrophilic nanospheres, which combine several key structural features that make these systems very beneficial for effective drug delivery. These are: a cross-linked ionic core; a hydrophilic PEO shell; and nanoscale size. They are, in essence, nanoscale single molecules that are stable upon dilution and can withstand environmental challenges such as changes in pH, ionic strength, solvent composition, and shear forces without structural deterioration. These favorable characteristics of the polymer micelles with cross-linked ionic cores motivated our ongoing efforts to elucidate their potential as efficient carriers for the delivery of anticancer drugs. Here we would like to report on the further progress in the use of BIC as templates
for preparation of cross-linked polymeric micelles. Specifically, the objective of this work is to ascertain the structural determinants that guide the self-assembly of BIC templates and physicochemical characteristics of the resulting cross-linked micelles. Indeed, it is important to understand whether the properties of such particles can be easily tuned by varying the composition of the "soft" core (i.e., structure of block ionomer, cross-linker and number of cross-links) and/or by altering the length of PEO chains in the outer corona of the micelles. This, in turn, may affect drug loading efficiency, swelling, interaction and attachment to surfaces, diffusion and drug release properties.
EXPERIMENTAL
Block copolymers of PEO and methacrylic acid or acrylic acid (further designated as PEO-^-PMA and PEO-&-PAA) were purchased from Polymer Source Inc., Canada. The sodium salts of these block copolymers were used for preparation of BIC. The list of block copolymers used in this work and their molecular characteristics are presented in Table 1. Diblock copolymer samples are denoted as PEO(x)-^-PMA(or PAA)(y), where x and y represent the degree of polymerization of the PEO segment and PMA/PAA segment, respectively. For example, PEO(170)-&-PMA(180) represents a diblock copolymer containing 170 ethylene oxide repeat units and 180 sodium methacrylate units. The concentration of carboxylate groups in the copolymer samples was estimated by potentiometric titration. Calcium chloride, barium chloride and gadolinium chloride, various cross-linker molecules (1,2-ethylenediamine, 1,5-diaminopentane, N-(3-aminopropyl-1,3-propanediamine, cystamine di-hydrochloride, 2,2'-(ethylenedioxy)bis(ethylamine)), cw-dichlorodiamminoplatinum (II) (cisplatin), and other chemicals were purchased from Sigma—Aldrich (St Louis, MO) and were used as received.
Turbidity measurements. The turbidity measurements were carried out at 420 nm using a Perkin-Elm-er Lambda 25 UV/VIS spectrophotometer after equilibration of the system for 3 min, which was proven to be sufficient for equilibration. The data are reported as (100 - 7)/100, where T is transmittance (%).
General procedure for the synthesis of cross-linked micelles. Cross-linked micelles were prepared by the previously described method with a slight modification [32]. In brief, PEO-^-polyacid/MeB+ complexes were prepared by mixing an aqueous solution of corresponding of PEO-^-polyacid with a solution of MeClB at a molar ratio of [MeB+]/[COO-] = 0.3-1.3. The 1-(3 - dimethylaminopropyl) - 3 - ethylcarbodiimide hy-drochloride (EDC) was added into solution of PEO-¿-polyacid/MeB+ complexes to create an active-ester intermediate with carboxylic groups of polyacid segments followed by addition of the solution of cross-linker. The extent of degree of cross-linking was con-
Polyanion-metal complex core Cross-linked ionic core
PEO-A-polyanion
Polymer micelle peo shell
Fig. 2. Scheme for the synthesis of polymer micelles with cross-linked cores.
trolled by the ratio of the amine functional groups to carboxylic acid groups. The reaction mixture
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