On the importance of Polyurethane and Polyurea Nanosystems for Future Drug Delivery
During the last decade, polyurethanes and polyureas (PUUa) have emerged as promising alternatives to classical polyacrylate-, polyester- and polyamino acid-based drug delivery nanosystems. Not only are polyurethanes and polyureas biocompatible and biodegradable, but they also facilitate the manufacture of nanoparticles of one order superior in size in quantitative yields. The versatile chemistry reduce the amount of organic solvents required and allows the straightforward multifunctionalization of polymer of these compounds helps to precursors with the desired targeting molecule at each stage of the process.
To highlight the common issues encountered in current drug delivery systems and the state of the art of polyurethane and polyurea polymers that self-assemble in a stratified manner by hydrophobic interactions. Finally, we discuss the importance of taking a holistic view when applying polymer nanotechnologies, in order to enhance their efficiency during preclinical and clinical studies.
PUUa nanoparticles emerge as suitable platforms to be manufactured in a cost-effective manner at industrial scale and following environmentally friendly synthetic methods. Furthermore, they allow the controlled delivery of a wide range of drugs and can be rapidly adapted to many clinical requirements by means of FDA-approved precursors. Additionally, the ease with which PUUa nanoparticles are biodegraded ensures control over temporal aspects of drug delivery compared to other nanosystems. These advantages make PUUa NPs attractive drug delivery vehicles as long as adequate safety and ethical guidelines for new NP formulations are developed.
1. Introduction to polymer nanosystems
The use of nanoparticulate systems in biomedicine has evolved from a mere promise of future benefits to a reality. As an evidence of this, a search for the term “nanoparticle” in Pubmed reveals that 90,000 out of a total of 140,000 articles have been published since 2012. Equally relevant is the impetus given by the European Commission under the Horizon 2020 research and innovation framework, which is providing nearly €80 billion of funding over 7 years (2014 to 2020), in addition to private investment in this field.
The market is the indisputable judge of today’s progress and in this regard nanomedicine is no exception. It is clear that there is a need to advance towards the development of novel therapeutic solutions with reliable scalability and under good manufacturing practices. Large pharmaceutical companies have made significant investments in buying IP rights of small companies in order to be in a better position to respond to current and future demands. Most efforts have been channeled into nanosystems with a high level of biocompatibility, biodegradability and industrialization feasibility. Understandably, as generally occurs with cutting-edge technologies, regulatory issues limit the application of nanosystems or slow them down. Therefore, the most clinically advanced polymeric NPs currently available are made of biodegradable and biocompatible polyaminoacid and polyester polymers, such as PLA, PGA, PLGA, and PCL1. Regulatory bodies have approved these systems as therapeutically safe, leading to a wider application of polymeric NPs. They exhibit excellent biocompatibility and biodegradability, good encapsulation yields and acceptable tunability2. However, in advanced development stages, these aspects can be significantly affected due to complex bio-nano interactions and scale-up processes, resulting into premature biodegradation3,4, poor encapsulation stability5 and limited polymers tunability6. Therefore, nanomedicine demands industrially viable polymers fulfilling these requirements using more environmentally friendly methodologies and a realistic manufacturing cost.
1.1. Biodegradability of polymer nanosystems
In cancer therapy, biodegradation of the nanosystem is essential to achieve an effective release of the enclosed drug to the desired tissue. Likewise, the degradation of the polymeric residues that previously configured the shell is equally important to guarantee a safe drug delivery. However, a biodegradation occurring too prematurely leads to a non-effective controlled release. This fact is observed when physical and chemical interactions between the nanosystem and external biomolecules are sufficient to cause a significant destructuration of the polymeric shell that ends with leakage. For instance, the most widely used polyaminoacid polymers, such as PLGA self-assembled structures, are too rapidly hydrolyzed by esterases, leading to an early release before reaching the target4,7. Such effect can be avoided if the polymers enclosing the drug are forming a crosslinked network.
1.2. Encapsulation stability
To improve encapsulation yields and stability, liposomal structures made of amphiphilic polymers are used, but they do not fulfill in vitro and in vivo requirements, since they present a diffuse and too labile configuration. This might be the result of low nanostructure control at the emulsification step, which leads to a random aggregation, irregular structuration and eventually causes the walls to leak8,9. These kind of amphiphilic structures facilitate the superficial entrapment of hydrophobic cargos instead of a successful encapsulation in the core; thus, a natural interaction and extraction of the drug by external molecules (phospholipids, cholesterol, lipophilic proteins, etc.) is boosted5,10,11. We believe that to avoid such undesired effects, polymers should be tailor-designed, promoting the generation of hermetic multi-walled shells that prevent the drug release before reaching the desired target. Another issue that should be tackled in common DDS is the use of external surfactants. When surfactants are used to decrease the size of the nanodispersion, they also interfere with the interface where the polymerization takes place. This creates some diffusion channels, decreasing the global robustness and promoting Ostwald ripening effects12. These phenomena need to be compensated with other external additives. Such excess of free surfactants has proved to be a primordial cause of later toxicities13,14, e.g. by the immune system, and therefore leading to nanosystem failure.
1.3. Polymer nanosystems tunability
The capacity of a polymer to convey multiple functionalities, tunability, is directly related to its intrinsic reactivity. Ideally, a polymer should exhibit high reactivity in intermediate stages to allow its bioactivation before final end-capping/crosslinking steps. For instance, fluorescent tags, cell targeting compounds and labile moieties should be easily linked to the main polymer backbone via covalent bonds to achieve multifunctionality. Nevertheless, a polymer should also be modulable in terms of structure in order to create not only linear or crosslinked structures but also to generate more sophisticated morphologies, such as multilayers or dendrimers. To achieve such a broad range of structures the use of a versatile chemistry offering multiple possibilities is essential. In this regard, the desired morphology will just depend on the adequate selection of certain starting monomers.15,16
In this context, polyurethane-polyurea polymers (PUUa) will play a crucial role in the near future due to their chemical versatility, scalability and biocompatibility17. During the last years, PUUa with a broad range of chemical properties and good biocompatibility have been developed for tissue engineering and drug delivery purposes18–34. They have proved to exhibit outstandingly flexible physicochemical properties, enabling the introduction of specific block copolymers into the polymer backbone. These developments have situated PUUa among the biomaterials of choice in various clinical applications35.
2. Self-stratified nanostructures of polyurethane-polyurea polymers by hydrophobic interactions
Although stratification in macromolecular structures has been accepted for a long time36–38, its importance has not been recognized until the oil-in-water interfacial properties allowed to predict structure-property relationships39,40. The understanding of the molecule-molecule interactions at the interface led to the development of a new generation of multifunctional polymeric materials and particularly, colloidal nanomaterials. Hydrophobic interactions are common and crucial in and between biological systems, in fact they are essential to life (e.g. protein folding processes, phospholipids self-assembling in cell membranes, DNA tertiary structure assembly, etc.). Those interactions can also be found in colloids and are starting to be rationally applied to design everyday materials with technological applications41,42. Recent advances in the development of stimuli-sensitive colloidal materials with stratifying capacities will be discussed in the following section. Special emphasis is given to the importance of hydrophobic interactions to control the nanostructure and properties of nanosystems. Hydrophobic forces are capable of forming compact but also flexible structures to allow adaptability in cellular and sub-cellular membranes of live organisms. Although Nature uses hydrophobic forces to create compartments, the connectors between them are not trivial. It has been demonstrated that the self-assembly of amphiphilic molecules, and more specifically polymers, is under thermodynamic control. Thus, self-stratified structures simply result from the tendency of each molecule to reach its lowest chemical potential, or in other words, the highest thermodynamic stability. Some in-house experiments point out that there is a parallelism between what occurs in vesicle formation, e.g. phospholipid vesicles, and in nanostructuration of amphiphilic polymers. In the former, the polar heads tend to segregate while the hydrophobic tails drive subsequent nanostructuration whereas in the latter, the hydrophilic and hydrophobic dangling chains self-orientate according to the external environment, causing the same effect. This would entail that above a critical micelle concentration (c.m.c), the opposing thermodynamic preferences presented by the two tails of an amphiphilic polymer are mostly accommodated by self-association to form a peculiar aggregate. Such entity would contain the hydrocarbon chains oriented towards a hydrophobic core, avoiding at all cost contact with water and the hydrophilic pendant chains pointing outwards43. Interestingly, the longest the hydrophobic chain, the lowest the c.m.c becomes, being approximately 1 M for C6 carbon chains and down to 10-9 M for C36 biological phospholipids37. The size and shape of amphiphilic nanostructures is determined both by geometric and thermodynamic factors. One of the geometric elements is the surface/volume ratio: as the number of hydrophobic chains in the core (volume) increases, the surface area per chain decreases44. The second but equally important geometric factor is that the core cannot be smaller than two hydrophobic chains coming from opposite sides of the shell. However, that would generate very small structures with large spaces between the polar heads, promoting contact between hydrophobic tails and water, which would result into destabilization and coalescence of nanoassemblies. In order to solve that thermodynamic issue, longer polar groups, such as PEG-based chains are used in the amphiphilic polymers, providing flexibility and adopting certain configurations such as brushes or mushrooms45. Hydrophilic chains self-accommodate at the oil-in-water interface (polyethylene oxide polymers are partially amphiphilic) and cause a decrease in hydrophobic tail-water interactions45. In this regard, it has been observed that a certain polyethylene glycol monomethyl ether molecule containing two very close alcohols (YMER™ N120) produces a highly packed molecular structure with an improved barrier effect. This prevents diffusion of entrapped cargos and makes encapsulations more stable per se. To create self-stratified polymer nanostructures through hydrophobic effects, both hydrophilic and hydrophobic segments of an amphiphilic polymer need to be specifically designed in order to increase thermodynamic and geometric stability. With the aim of reducing the energy barrier and thus achieving the formation of stratified polymer structures geometrical factors have to be considered. For instance, the length of hydrophobic and hydrophilic segments and their position in the polymer backbone (in the main chain or sideways) have to be accurately addressed. Specifically, the hydrophilic PEG and YMER™ N120 monomers, both 1000 Mw approximately, are hydrophilic and have almost equivalent reactivity (Figure 1a and 1b), but when polymerized they show distinct morphologies in water.
Figure 1. a), b) Ideal chemical structures of YMER™ N120 and PEG 1000, c) Adopted morphology of polymerized YMER™ N120 towards the aqueous phase of an oil-in-water emulsion, d) Representation of the adopted morphology of polymerized PEG 1000 towards the aqueous phase of an oil-in-water emulsion.
Recent data supports that when these hydrophilic diols are covalently attached to the backbone of an amphiphilic polymer, they self-orientate toward the aqueous phase differently. When emulsified, YMER™ N120 decreases the required energy to be solvated and that enhances the formation of colloidal systems thanks to the adoption of a two-headed serpent structure (Figure 1c). In contrast, PEG 1000, which contains two hydroxyl groups separated by the whole polyethoxylenated segment, leads to a morphological constriction of the hydrophilic chain. In addition, the resulting twisted hydrophilic polyethoxylenated chains would be repelled from each other. Therefore, this rearrangement would be less thermodynamically favored and the colloidal systems formed less stable44 (Figure 1d).
A similar behavior is observed in the hydrophobic segments of an amphiphilic polymer. Albericio’s group recently published that polymer nanoparticles exhibiting hydrophobic side-chains showed high encapsulation stability in bioresponsive shells with diameters below-50 nm 46. This phenomenon becomes more relevant when the formation of self-stratified nanostructures is sought. Energetic constraints between hydrophobically stratified polymers will be less important when long hydrophobic pendant chains are responsible for such stratification. As explained before, hydrophobic pendant chains will facilitate interactions of an amphiphilic polymer with a hydrophobic one through superposition of hydrophobic chains (Figure 2a). Non-pendant hydrophobic chains will be geometrically restricted to self-orientate towards the oily phase and thermodynamically unstable due to their energetically disfavored proximity to hydrophilic water molecules, which induces NP aggregation in water (Figure 2b).
Figure 2. Ideal structures of amphiphilic prepolymers at the oil-in-water interface. a) Hydrophobic pendant chains easily self-accomodate towards the oily phase (nanoparticle core). Hyfob and Amphil hydrophobic chains stabilize by hydrophobic effects the stratified nanostructure of a nanoparticle polymeric shell. b) Hydrophobic non-pendant chains are more restricted to self-allocate toward the oily phase as they need to constrict the whole amphiphilic prepolymer to adopt a pendant structure. Hyfob and Amphil hydrophobic chains remain close to the aqueous environment, in contact with water molecules, which is thermodynamically unfavored and destabilizes the oil-in-water interface.
Hydrophobic interactions are usually strong enough to remain stable in water or in ionic aqueous media. However, when external amphiphilic molecules come into play, some structural rearrangements might occur in order to reach thermodynamic stability again. Accordingly, amphiphilic and hydrophobic moieties have proved to be able to disrupt hydrophobically stratified entities both in in vitro and in vivo experiments10,11,44. In drug delivery systems mediated by polymer nanoparticles, the stability of a nanovehicle is intended to remain unchanged during blood circulation until reaching the desired site. On the contrary, the cargo would be unspecifically released and thus ineffective. In this regard, the chemical crosslinking would be an interesting approach to avoid destabilization of stratified nanostructures by external factors that would jeopardize the cell-targeted delivery of the encapsulated cargo46,47. For instance, crosslinking of isocyanate-reactive stratified nanostructures with highly reactive polyamines would “freeze” previously formed configurations, increase the stability against amphiphilic and hydrophobic molecules (e.g. phospholipids, cholesterol, proteins, etc.) and eliminate toxicological issues related to free isocyanate functional groups48–50 (Figure 3).
Figure 3. Ideal structures of reactive PUUa prepolymers at the oil-in-water interface a) before and b) after crosslinking via urea bonds with polyamines.
3. Holistic considerations on therapy with polymer nanoparticles
There is an erroneous tendency to believe that nanomedicines are able to, for instance, kill any type of tumor or eradicate bacterial infections located in any body tissue. Unfortunately, this vision can lead to misinterpret nanotechnologies as a "one-for-all" remedy and even turn them into another bubble51. Like any other discipline the release of drugs in a controlled manner using polymer nanotechnology is a field involving multiple factors. This holistic view applied to nanomedicine aims to take into account different elements: the engineering of new high-throughput synthetic methodologies, the route of administration, the molecular composition of the nanosystem and the coordination with human temporal cycles. Finally, the proper adjustment of the safety measures and ethical regulations to reduce the hurdles in the translation of smart polymeric NPs from preclinical studies to clinical practice.
There is a growing demand for novel starting products to prepare polymeric nanomaterials in high purities, with minimal purification steps and following green chemistry methodologies. The commercial availability of precursors and the straightforward scale-up of the final product are essential for the translation of lab nanosystems to commonly use of nanomedicines.
It would be desirable to select the route of administration for a specific drug delivery system in a rational basis. Intravenous administration is sometimes overestimated and sometimes alternative routes can provide a more efficient delivery to the desired tissue. In addition, it should be noted that other factors such as potential interactions occurring between the NP and the surroundings of the target tissue can influence the delivery rate, targeting efficiency and even cytotoxicity. Consequently, pharmacokinetic studies for a specific drug delivery system should be performed taking into account the release profile of the drug under in vivo conditions. Specific and efficient molecular targeting will be successful only when all the factors affecting it are thoroughly analyzed.
Among all the factors affecting targeting efficiency, the most noticeable one is the unspecific NP uptake observed in biodistribution experiments. There are various parameters that are significant to the NP fate: morphology, size, shape and surface properties. They can be properly adjusted and modulated by varying the physicochemical properties of the system, such as crosslinking degree, HLB value and amphiphilic or amphoteric behavior. The protein corona formed around the NP shell upon incubation in serum proteins has been demonstrated to cause undesired receptor-mediated uptake and clear distortion of the specific interactions occurring between the receptor and the targeting molecule. Therefore, several efforts have been invested in coating NPs with stealth polymers to minimize physisorption of circulating proteins that promote sequestration by the RES52. Specific active targeting is also hampered by the fact that the molecular target is sometimes not uniquely overexpressed in the tissue, leading to a specific targeting but not only in diseased cells. A promising approach would be the local delivery of the nanosystem.
A long circulation is essential to achieve drug accumulation in the diseased tissue. Highly functionalized shells can often cause increased immunogenicity and reduced circulation times, which should be appropriately addressed by modulating the amount of ligand to achieve minimal immuno-responses and maximal cell specificity. Finally, the drug needs to reach the intended intracellular target. For instance, in case a NP is engineered to release its cargo upon interaction with cytosolic glutathione, the ability of the NP to escape the endosome-lysosome should be proved (small enough NPs, changing amphiphilic surface, functionalization with a cell-penetrating peptide to enter the cytosol, etc.).
In the last fifteen years, chronopharmacological aspects of drug delivery systems have been addressed to increase patient compliance and drug efficacy. In the rapidly evolving scenario of nanomedicines, drug delivery systems with sustained release have become obsolete while modern smart self-controlled DDS have emerged. Chronopharmaceutics serve to generate knowledge ranging from the biology of disease to the development of models for the design and evaluation of DDS. In this regard, modeling approaches are available in a broad range of fields: cardiovascular diseases, cancer chemotherapy, rheumatoid arthritis and diabetes, among others. For instance, it has been proved that the plasmatic levels of certain proteins involved in the pain suffered by arthritic patients are subject to a circadian rhythm. Thus, drugs currently used to treat this pain show statistically significant differences in toxicity and efficacy depending on the time of the day they are administered.
Finally, in order to maximize the social and therapeutic benefits of nanomedicine and reduce potential health hazards, regulatory systems should guarantee the appropriate infrastructures for the development of ethical and safe nanotechnologies. Moreover, special emphasis should be put on dissemination activities to properly inform the society about potential applications of nanomedicine, its advantages and disadvantages as an alternative to classical medicine.
In summary, poylurethane and polyurea NPs emerge as suitable platforms to be manufactured in a cost-effective manner at industrial level and following environmentally friendly synthetic methods. Furthermore, they allow the controlled delivery of a wide range of drugs and can be rapidly adaptable to many clinical requirements by using FDA-approved precursors. Additionally, the ease with which the PUUa NPs can be biodegraded ensures the control over temporal aspects of drug delivery compared to other nanosystems. These advantages make PUUa NPs attractive drug delivery vehicles but only if adequate safety and ethical guidelines for new NP formulations are developed.
1 N. Kamaly, Z. Xiao, P. M. Valencia, A. F. Radovic-Moreno and O. C. Farokhzad, Chem. Soc. Rev., 2012, 41, 2971–3010.
2 R. A. Jain, Biomaterials, 2000, 21, 2475–2490.
3 G. Kumar, N. Shafiq and S. Malhotra, Crit. Rev. Ther. Drug Carr. Syst., 2012, 29, 149–182.
4 G. Crotts and T. G. Park, J. Microencapsul., 1998, 15, 699–713.
5 P. Xu, E. Gullotti, L. Tong, C. B. Highley, D. R. Errabelli, T. Hasan, J.-X. Cheng, D. S. Kohane and Y. Yeo, Mol. Pharm., 2009, 6, 190–201.
6 I. Bala, S. Hariharan and M. R. Kumar, Crit. Rev. Ther. Drug Carrier Syst., 2004, 21, 387–422.
7 R. Jalil and J. R. Nixon, J. Microencapsul., 1990, 7, 297–325.
8 N. Mody, R. K. Tekade, N. K. Mehra, P. Chopdey and N. K. Jain, AAPS PharmSciTech, 2014, 15, 388–399.
9 R. Gref, A. Domb, P. Quellec, T. Blunk, R. H. Müller, J. M. Verbavatz and R. Langer, Adv. Drug Deliv. Rev., 2012, 64, 316–326.
10 H. Chen, S. Kim, L. Li, S. Wang, K. Park and J.-X. Cheng, Proc. Natl. Acad. Sci. , 2008, 105, 6596–6601.
11 P. Zou, H. Chen, H. J. Paholak and D. Sun, Mol. Pharm., 2013, 10, 4185–94.
12 L. Ratke and P. W. Voorhees, Growth and Coarsening : Ostwald Ripening in Material Processing, Springer Berlin Heidelberg, 2002.
13 J. M. Asua, Prog. Polym. Sci., 2014, 39, 1797–1826.
14 K. Knop, R. Hoogenboom, D. Fischer and U. S. Schubert, Angew. Chem. Int. Ed. Engl., 2010, 49, 6288–308.
15 H.-W. Engels, H.-G. Pirkl, R. Albers, R. W. Albach, J. Krause, A. Hoffmann, H. Casselmann and J. Dormish, Angew. Chem. Int. Ed. Engl., 2013, 52, 9422–41.
16 D. K. Chattopadhyay and K. V. S. N. Raju, Prog. Polym. Sci., 2007, 32, 352–418.
17 E. Delebecq, J.-P. Pascault, B. Boutevin and F. Ganachaud, Chem. Rev., 2013, 113, 80–118.
18 M. Ding, L. Zhou, X. Fu, H. Tan, J. Li and Q. Fu, Soft Matter, 2010, 6, 2087.
19 M. Ding, J. Li, X. Fu, J. Zhou, H. Tan, Q. Gu and Q. Fu, Biomacromolecules, 2009, 10, 2857–65.
20 X. Jiang, J. Li, M. Ding, H. Tan, Q. Ling, Y. Zhong and Q. Fu, Eur. Polym. J., 2007, 43, 1838–1846.
21 X. Jiang, F. Yu, Z. Wang, J. Li, H. Tan, M. Ding and Q. Fu, J. Biomater. Sci. Polym. Ed., 2010, 21, 1637–52.
22 C. Yang, X. Ni and J. Li, J. Mater. Chem., 2009, 19, 3755.
23 J. Zhang, M. Wu, J. Yang, Q. Wu and Z. Jin, Colloids Surfaces A Physicochem. Eng. Asp., 2009, 337, 200–204.
24 G. Morral-Ruíz, P. Melgar-Lesmes, C. Solans and M. J. García-Celma, J. Control. Release, 2013, 171, 163–71.
25 G. Morral-Ruíz, P. Melgar-Lesmes, A. López-Vicente, C. Solans and M. J. García-Celma, Nano Res., 2015, 8, 1729–1745.
26 F.-Y. Hsieh, H.-H. Lin and S.-H. Hsu, Biomaterials, 2015, 71, 48–57.
27 S. Sartori, V. Chiono, C. Tonda-Turo, C. Mattu and C. Gianluca, J. Mater. Chem. B, 2014, 2, 5128.
28 K. Landfester, Angew. Chemie Int. Ed., 2009, 48, 4488–4507.
29 J. Zhang, Z. Sun, H. Zhu, Q. Guo, C. He, A. Xia, H. Mo, X. Huang and J. Shen, J. Mater. Chem. B, 2016, 4, 1116–1121.
30 Y. Yao, H. Xu, C. Liu, Y. Guan, D. Xu, J. Zhang, Y. Su, L. Zhao and J. Luo, RSC Adv., 2016, 6, 9082–9089.
31 V. H. Shargh, H. Hondermarck and M. Liang, Nanomedicine (Lond)., 2016, 11, 63–79.
32 E. Ajorlou, A. Y. Khosroushahi and H. Yeganeh, Pharm. Res., 2016, 33, 1426–39.
33 C. Mattu, A. Silvestri, T. R. Wang, M. Boffito, E. Ranzato, C. Cassino, G. Ciofani and G. Ciardelli, Polym. Int., 2016, 65, 770–779.
34 G. Morral-Ruíz, P. Melgar-Lesmes, C. Solans and M. J. García-Celma, Advances in Polyurethane Biomaterials, Elsevier Science, 2016.
35 B. Brooks, A. Brooks and D. Grainger, in Biomaterials Associated Infection SE - 13, eds. T. F. Moriarty, S. A. J. Zaat and H. J. Busscher, Springer New York, 2013, pp. 307–354.
36 H. J. Hamilton, Zygon�, 1977, 12, 289–335.
37 C. Tanford, Science, 1978, 200, 1012–1018.
38 N. Muller, Acc. Chem. Res., 1990, 23, 23–28.
39 O. Félix, Z. Zheng, F. Cousin and G. Decher, Comptes Rendus Chim., 2009, 12, 225–234.
40 D. Chandler, Nature, 2005, 437, 640–647.
41 M. W. Urban, Curr. Opin. Colloid Interface Sci., 2014, 19, 66–75.
42 M. W. Urban, Prog. Polym. Sci., 2009, 34, 679–687.
43 L. Maibaum, A. R. Dinner and D. Chandler, 2004, 1–13.
44 S. C. Owen, D. P. Y. Chan and M. S. Shoichet, Nano Today, 2012, 7, 53–65.
45 O. Garbuzenko, Y. Barenholz and A. Priev, Chem. Phys. Lipids, 2005, 135, 117–29.
46 P. Rocas, Y. Fernández, S. Schwartz, I. Abasolo, J. Rocas and F. Albericio, J. Mater. Chem. B, 2015, 3, 7604–7613.
47 C. Cuscó, J. Garcia, E. Nicolás, P. Rocas and J. Rocas, Polym. Chem., 2016, 7, 6457–6466.
48 Ecopol Tech, S.L., WO2014114838A3, 2014.
49 M. Talelli, M. Barz, C. J. F. Rijcken, F. Kiessling, W. E. Hennink and T. Lammers, Nano Today, 2015, 10, 93–117.
50 S. Shu, X. Zhang, Z. Wu, Z. Wang and C. Li, Biomaterials, 2010, 31, 6039–6049.
51 H. Möhwald and P. S. Weiss, ACS Nano, 2015, 9, 9427–9428.
52 B. Kang, P. Okwieka, S. Schöttler, S. Winzen, J. Langhanki, K. Mohr, T. Opatz, V. Mailänder, K. Landfester and F. R. Wurm, Angew. Chemie Int. Ed., 2015, n/a-n/a.