ELASTIN-LIKE RECOMBINAMERS FOR MULTI-MODAL DRUG DELIVERY SYSTEMS

Elastin is a protein with a key role in most of mammalian tissues and it is widely expressed in the extracellular matrix present over myocardium, cartilage and skin. Its elastogenic activity relies on the main cellular components of the tissue network, for instance, endothelial cells, fibroblasts, chondrocytes, and keratinocytes [1]. Nevertheless, the human elastin is naturally synthesized in early age, leading to a drawback based on the low availability, due to the stop in the natural synthetic mechanism with ageing. A clever strategy to overcome such an issue is based on the development of genetically-engineered elastin-mimicking peptides fabrication, so-called elastin-like recombinamers (ELRs), thus balancing the low availability of natural elastin and tuning the biomaterial structuring and behaviour. Relevant advances in the field can derive from the investigation of the morphological, mechanical, in-vitro and delivery-related properties of ELRs-based systems, fabricated in the form of either hydrogel or microspheres. Different scaffold constructs are studied herein, i.e., microspheres, hydrogel and microsphere integrated hydrogel in order to assess their delivery suitability and thoroughly understand the hierarchical complex structuring of the elastin-like recombinamer self- assembly mechanisms. We used two ELRs (1-HRGD6-cyclooctyne, 2-REDV-N3) modified with the two different reactive groups needed to form hydrogels via a click reaction and functionalized with two different bioactive sequences RGD and REDV that would promote cell adhesion. In this study the most stable and optimal concentration ratio of ELRs based hollow spheres exhibited no reduction in cellular metabolic activity. The sacrificial template-based method allowed us to engineer hollow spheres with a first layer of the ELRs HRGD6-component followed by a second layer of the ELRs REDV-component, by means of copper free click-chemistry reaction. The ELRs hollow spheres-tethered ELRs hydrogel was prepared by adding the pre-fabricated ELRs hollow spheres. The hydrogel construct was characterized by rheology, NMR, and Synchrotron Radiation SAXS (SRSAXS). Hollow spheres were characterized by TEM, SEM, DLS and FT-IR. Drug upload and release were assessed by means of ELISA, confocal microscopy and all constructs were successfully tested for cell metabolic activity, revealing no cytotoxicity. ELR-based hollow microspheres were fabricated and successfully entrapped into an ELR- hydrogel matrix. Release studies have been performed, determining the ELRs platform suitability as drug delivery system.