In the past decade, stimuli-responsive hydrogels are increasingly studied as biomaterials for tissue engineering and regenerative medicine purposes. Smart hydrogels can not only replicate the physicochemical properties of the extracellular matrix but also mimic dynamic processes that are crucial for the regulation of cell behavior. Dynamic changes can be influenced by the hydrogel itself (isotropic vs anisotropic) or guided by applying localized triggers. The resulting swelling-shrinking, shape-morphing, as well as patterns have been shown to influence cell function in a spatiotemporally controlled manner. Furthermore, the use of stimuli-responsive hydrogels as bioinks in 4D bioprinting is very promising as they allow the biofabrication of complex microstructures. This perspective discusses recent cutting-edge advances as well as current challenges in the field of smart biomaterials for tissue engineering. Additionally, emerging trends and potential future directions are addressed.

Polymeric micelles constitute an important class of nanomaterials that are highly attractive for pharmaceutical applications. The hydrophobic core of the micelles can be loaded with poorly water soluble drugs, while the shell of the micelles provides colloidal stability in vitro and in vivo. In recent years, covalent cross-linking of micelles is attracting increasing attention, because in vitro data show that it can prevent the self-assembled micelles from dissociation, can modulate drug release and may provide tools for triggered release. This paper reviews the methods used to cross-link either the core or the shell of the micelles, and focuses on drug delivery applications of cross-linked micelles. Whereas non-cross-linked micelles generally do not improve pharmacokinetics of encapsulated drugs when compared to common drug formulations, recent in vivo data show that cross-linking provides dramatic improvements in both pharmacokinetics and biodistribution of the micelles, and that drugs can fully benefit from that when they are covalently linked to the micelles. Although the field is still in its infancy, the latest results promise a bright future of cross-linked micelles for drug delivery and/or diagnostic applications.

Micelles composed of block copolymers of poly(ethylene glycol)-b-poly(N-2-benzoyloxypropyl methacrylamide) (mPEG-b-p(HPMA-Bz)) have shown great promise as drug delivery carriers due to their excellent stability and high loading capacity. In the present study, parameters influencing micelle size were investigated to tailor sizes in the range of 25 to 100 nm. Micelles were prepared by a nanoprecipitation method and their size was modulated by the block copolymer properties such as molecular weight, their hydrophilic to hydrophobic ratio, homopolymer content, as well as formulation and processing parameters. It was shown that the micelles have a core-shell structure using a combination of dynamic light scattering and transmission electron microscopy analysis. By varying the degree of polymerization of the hydrophobic block (NB) between 68 and 10, at a fixed hydrophilic block mPEG5K (NA=114), it was shown that the hydrophobic core of the micelle was collapsed following the power law of (NB×Nagg)1/3. Further, the calculated brush height was similar for all the micelles examined (10 nm), indicating that crew-cut micelles were made. Both addition of homopolymer and preparation of micelles at lower concentrations or lower rates of addition of the organic solvent to the aqueous phase increased the size of micelles due to partitioning of the hydrophobic homopolymer chains to the core of the micelles and lower nucleation rates, respectively. Furthermore, it was shown that by using different solvents, the size of the micelles substantially changed. The use of acetone, acetonitrile, ethanol, THF and dioxane, resulted in micelles in the size range from 45 to 60 nm after removal of the organic solvents. The use of DMF and DMSO led to markedly larger sizes of 75 and 180 nm respectively. In conclusion, the results show that by modulating polymer properties and processing conditions, micelles with tailorable sizes can be obtained.

In cancer treatment, polymeric nanoparticles (NPs) can serve as a vehicle for the delivery of cytotoxic proteins that have intracellular targets but that lack well-defined mechanisms for cellular internalization, such as saporin. In this work we have prepared PEGylated poly(lactic acid-co-glycolic acid-co-hydroxymethyl glycolic acid) (PLGHMGA) NPs for the selective delivery of saporin in the cytosol of HER2 positive cancer cells. This selective uptake was achieved by decorating the surface of the NPs with the 11A4 nanobody that is specific for the HER2 receptor. Confocal microscopy observations showed rapid and extensive uptake of the targeted NPs (11A4-NPs) by HER2 positive cells (SkBr3), but not by HER2 negative cells (MDA-MB-231). This selective uptake was blocked upon pre-incubation of the cells with an excess of nanobody. Non-targeted NPs (Cys-NPs) were not taken up by either type of cells. Importantly, a dose-dependent cytotoxic effect was only observed on SkBr3 cells when these were treated with saporin-loaded 11A4-NPs in combination with photochemical internalization (PCI), a technique that uses a photosensitizer and local light exposure to facilitate endosomal escape of entrapped nanocarriers and biomolecules. The combined use of saporin-loaded 11A4-NPs and PCI strongly inhibited cell proliferation and decreased cell viability through induction of apoptosis. Also the cytotoxic effect could be reduced by an excess of nanobody, reinforcing the selectivity of this system. These results suggest that the combination of the targeting nanobody on the NPs with PCI are effective means to achieve selective uptake and cytotoxicity of saporin-loaded NPs.

To avoid poly(ethylene glycol)-related issues of nanomedicines such as accelerated blood clearance, fully N-2-hydroxypropyl methacrylamide (HPMAm)-based polymeric micelles decorated with biotin for drug delivery were designed. To this end, a biotin-functionalized chain transfer agent (CTA), 4-cyano-4-[(dodecylsulfanylthiocarbonyl)-sulfanyl]pentanoic acid (biotin-CDTPA), was synthesized for reversible addition-fragmentation chain-transfer (RAFT) polymerization. Amphiphilic poly(N-2-hydroxypropyl methacrylamide)-block-poly(N-2-benzoyloxypropyl methacrylamide) (p(HPMAm)-b-p(HPMAm-Bz)) with molecular weights ranging from 8 to 24 kDa were synthesized using CDTPA or biotin-CDTPA as CTA and 2,2'-azobis(2-methylpropionitrile) as initiator. The copolymers self-assembled in aqueous media into micelles with sizes of 40-90 nm which positively correlated to the chain length of the hydrophobic block in the polymers, whereas the critical micelle concentrations decreased with increasing hydrophobic block length. The polymer with a molecular weight of 22.1 kDa was used to prepare paclitaxel-loaded micelles which had sizes between 61 and 70 nm, and a maximum loading capacity of around 10 wt%. A549 lung cancer cells overexpressing the biotin receptor, internalized the biotin-decorated micelles more efficiently than non-targeted micelles, while very low internalization of both types of micelles by HEK293 human embryonic kidney cells lacking the biotin receptor was observed. As a consequence, the paclitaxel-loaded micelles with biotin decoration exhibited stronger cytotoxicity in A549 cells than non-targeted micelles. Overall, a synthetic pathway to obtain actively targeted poly(ethylene glycol)-free micelles fully based on a poly(HPMAm) backbone was established. These polymeric micelles are promising systems for the delivery of hydrophobic anticancer drugs.