ALEJANDRO SOSNIK
5.2 Polymeric Micelles
Polymeric micelles are nano-sized (usually o100 nm) structures generated by the spontaneous self-assembly of amphiphilic block copolymers above a given concentration known as critical micellar concentration (CMC).19Hydrophobic blocks associate to form an inner core capable of solubilizing lipophilic drugs, while the hydrophilic ones form a corona that comes into direct contact with the external medium, stabilizing the system.20The corona also constitutes the interface between the drug reservoir and the release medium, and depending on its properties (e.g. microfluidity) and on the drug/corona interaction, the drug release could be facilitated or hampered. In general, when the hydrophilic block is longer than the hydrophobic one, micelles are spherical, while copolymers
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with longer hydrophobic blocks can give place to micelles of different morphology (e.g. rods and lamellae) or to polymeric vesicles (polymersomes).21 Depending on the molecular arrangement, reverse polymeric micelles with hydrophobic corona and hydrophilic core can be also produced in non-aqueous media.22However, only the former are relevant for drug-delivery purposes. The molecular properties of polymeric micelles can be tailored to adjust the size of the core and the nature and strength of core-drug interactions. In addition, the corona can be decorated with specific ligands to facilitate active drug targeting by means of the selective uptake mediated by specific receptors.
Polymeric micelles can be administered by different routes such as oral23,24and ocular,25,26though parenteral is the most extensively investigated one.19Approved conventional surfactants (e.g. polyethoxylated castor oil or polysorbate 80) form regular micelles in water and they are profusely employed for drug solubili- zation.27Nevertheless, these pharmaceutical excipients are not deprived of toxic effects. Moreover, when regular micelles undergo dilution to a final concentration below the CMC, they disassemble instantaneously and the drug is released into the medium. Conversely, polymeric micelles are safer for parenteral administration and more stable under dilution, providing more prolonged circulation times.
In addition, cores are larger resulting in greater encapsulation capacity.19
5.2.1 Micellar Encapsulation
The capacity of polymeric micelles to encapsulate a drug can be expressed by (i) the micelle–water partition coefficient defined as the ratio between the drug concentration inside the micelle and in the aqueous medium,28,29(ii) the number of moles solubilized per gram of hydrophobic block, and (iii) the molar solubilization ratio (MSR) that is the molar ratio between the drug and the copolymer.
Even though some solubilization capacity can be observed at copolymer concentrations below the CMC,30the most substantial solubilization is expected above this point owing to the ability of drug molecules to accommodate within the core. Paterson et al.30 proposed two simple equations to describe the solubilization process below the CMC (Equation 5.1) and above it (Equation 5.2):
IfCsoCMC
Sapparent
S ¼1þKunimerCs ð5:1Þ
IfCs4CMC Sapparent
S ¼1þKunimerCMCþKmicelle ðCsCMCÞ ð5:2Þ where Sapparent is the aqueous solubility of the drug measured in the micellar system, S is the molar intrinsic solubility in pure water, CS is the copolymer concentration in water,KunimerandKmicelleare equilibrium constants describing the solute-unimer (oCMC) and solute-micelle (4CMC) interactions.
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The incorporation capacity of drug molecules by a certain copolymer depends on its molecular weight and its hydrophilic-lipophilic balance (HLB).
In general, for similar molecular weights, the lower the HLB is, the greater the encapsulation capacity. Concomitantly, copolymers displaying similar HLB and greater molecular weight tend to display greater encapsulation capacity than those with smaller molecular weight. Moreover, encapsulation is also governed by drug features such as molecular volume or lipophilicity. In other words, the performance of specific amphiphiles needs to be assessed for each single molecule. The ideal solubility of a drug is governed by the intensity of the solute–solute interactions; the stronger these forces, the higher the melting temperature, Tm. Solubilization depends on the generation of strong solute–core interactions (e.g. hydrophobic forces) that overcome the solute–solute ones. Thus, drugs displaying low Tm are encapsulated more effectively than those with a greater one. For example, Chiappettaet al.31,32 investigated the solubilization of triclosan (289.5 g/mol; Tm¼55–571C) and triclocarban (315.6 g/mol;Tm¼2551C) in a variety of branched poly(ethylene oxide)-poly(propylene oxide) (PEO-PPO) polymeric micelles. These two anti- bacterial agents display similar molecular structure and molecular weight though very different Tm. The physical stability of the drug-loaded micelles was strongly dependent on the Tm of the encapsulated drug, being high for triclosan and very low for triclocarban. It is also interesting to note that even though the drug–core interaction plays a fundamental role in the encapsulation process, in some cases the interaction of certain hydrophilic functional groups of the drug molecule with the corona can contribute to improve the solubilization performance, as demonstrated with triclosan under different pH conditions.31
5.2.2 Preparation Methods
According to (i) the physico-chemical properties of the copolymer and more specifically those of the hydrophobic block and (ii) the properties of the drug, different techniques can be employed to produce drug-loaded polymeric micelles.21,33 The direct method may comprise the solubilization of the copolymer to obtain polymeric micelles and the subsequent solubilization of the drug that initially remains in suspension and it is gradually incorporated into the micelles until its complete dissolution. When the encapsulation capacity of a certain copolymer with respect to a drug is being assessed, a drug excess is added and the system is allowed to reach the equilibrium for 48–72 h at a constant temperature. Then, the drug excess is removed by filtration and the drug payload quantified. This method is the preferred one because it prevents the use of organic solvents and the implementation of additional operations to remove them. It is commonly used with copolymers of intermediate hydro- phobicity that are water soluble and for the encapsulation of drugs with low to intermediate molecular weight. In contrast, when the copolymer is highly hydrophobic and it does not solubilize conveniently in water, both copolymer and drug are primarily solubilized in a water-miscible organic solvent
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(e.g. dimethylformamide or acetone) and poured into water. Then, polymeric micelles are formed upon the elimination of the organic solvent by evaporation or dialysis. The main disadvantage of dialysis is that part of the encapsulated drug can be lost in the dialysis medium, thus leading to lower drug payloads.
Alternatively, the drug and the copolymer can be dissolved in a water- immiscible solvent, which is subsequently evaporated. Only then, the film is reconstituted with water to form the drug-loaded micelles.21A main constraint of this approach is that highly hydrophobic copolymers can be used in relatively low concentrations between 1 and 2%. Also, the presence of organic solvent residues needs to be quantified to ensure that the remaining concen- trations are below the maximum established limits. Other procedures can be applied to fit the properties of the copolymer and the drug. It is worth mentioning though that changes in the technique may result in systems with different drug payloads, micellar size and size distribution and physico- chemical stability.34,35
5.2.3 Physical Stability
One of the main drawbacks of polymeric micelles is that they tend to dis- assemble upon administration and dilution in the biological environment.
Disassembled polymeric micelles cannot maintain the encapsulated molecule within the core, and the drug is released into the medium where it can undergo nucleation, crystallization and precipitation. Even though the system is thermodynamically instable below the CMC and will finally disassemble, the kinetics of the process depends on the properties of the copolymer (e.g.
molecular weight, HLB, core amorphousness or semi-crystallinity and cohesion).36,37In general, the stability of relatively hydrophilic copolymers is jeopardized to a greater extent than that of more hydrophobic ones because the gap between the CMC and the final concentration upon dilution is greater.
Thus, several works improved the stability of the systems by increasing the hydrophobicity of the amphiphiles.37 However, the analysis is not so straightforward because the intrinsic properties of the drug may also condition the overall physical stability of the system; drugs displaying stronger solute–solute interactions and higher Tm tend to precipitate faster than those with lower Tm.31,32 Also, encapsulated drugs may favor or disfavor the aggregation of the amphiphile itself and greater drug–core cohesion may result in higher physical stability when compared to the drug-free micelle.38 For example, Chiappetta et al.39 showed that the antiretroviral efavirenz (EFV) promotes the self-aggregation of pristine and N-methylated branched PEO- PPO block copolymers (poloxamines). In this regard, it is crucial to monitor the long-term physical stability of the drug-loaded micelles upon dilution in media that are relevant to the clinical use;e.g. gastric-mimicking medium.
Two main approaches have been developed to stabilize physically polymeric micelles and to prevent their disassembly upon dilution: (i) core cross-linking, and (ii) corona cross-linking.40 Both pathways demand the chemical modifi- cation of the amphiphile with reactive functional groups and show pros and Temperature- and pH-sensitive Polymeric Micelles for Drug Encapsulation 119
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cons. The physical41 or chemical42–45 cross-linking of the core conserves the functionality of the terminal groups exposed on the micellar surface and enables the conjugation of ligands that are useful in active drug targeting. On the other hand, the drug loading capacity and the release rate can be reduced owing to a more densely packed core. Conversely, to cross-link the corona, terminal groups (e.g. hydroxyl) are modified and reacted with coupling bifunctional molecules or by free radical polymerization. Depending on the cross-linking density, the corona displays variable permeability, this parameter affecting drug encapsulation and release.46–49This is an interesting feature that can be exploited to develop micelles that display rate-controlling coronas and to fine-tune the release kinetics from the drug reservoir. It is worth remarking that to prevent the chemical modification of the drug, the stabilization stage is often carried out before the drug loading. Thus, the chemical modification of the micelle can alter the encapsulation capacity with respect to the pristine copolymer. Moreover, irreversibly cross-linked systems might display problems related to a more limited clearance from the body and they need to be engineered appropriately to ensure biocompatibility and to prevent accumu- lation.37In this context, a number of researchers developed covalently cross- linked polymeric micelles that disassemble in vivo by different biological pathways and that are eliminated more effectively.50–53
Regardless of the fact that stabilized micelles extend the circulation time under dilution, this phenomenon does not necessarily result in an improved therapeutic index of the drug. Thus, the appropriate balance between effective drug encapsulation and stabilization and drug release needs to be attained.54 Otherwise a strong core–drug interaction will curtail the gradual release of free drug with an appropriate kinetics. Amphiphiles bearing a semi-crystalline core such as those with hydrophobic blocks made of poly(e-caprolactone) (PCL) usually withstand better the dilution phenomena than those with amorphous cores (e.g. poly(propylene glycol)). However, they are not physically stable in suspension and tend to fuse and cluster into larger aggregates that finally precipitate.55To conserve these micelles in the long term, they usually need to undergo freeze-drying, a process that needs to be conducted in the presence of lyo-/cryoprotectants.56–59Regardless of the stabilization strategy, the physical stability and the properties of each drug-loaded system need to be compre- hensively assessed to understand their behaviorin vivo. Some pathologies are characterized by the localized increase of the temperature or the decrease of the pH. In this context, nanocarriers can be tailored to release the drug locally upon a change in the microenvironment.