Injectable Hydrogels of Stimuli-Responsive Elastin and Calmodulin-Based Triblock Copolypeptides for Controlled Drug Release

Abstract

A variety of block copolypeptides with stimuli responsiveness have been of growing interest for dynamic self-assembly. Here, multistimuli-responsive triblock copolypeptides composed of thermosensitive elastin-based polypeptides (EBP) and ligand-responsive calmodulin (CalM) were genetically engineered, over-expressed, and nonchromatographically purified by inverse transition cycling. Diluted EBP-CalM-EBP (ECE) triblock copolypeptides under physiological conditions self-assembled into vesicles at the nanoscale by temperature-triggered aggregation of the EBP block with lower critical solution temperature behaviors. Furthermore, concentrated ECE triblock copolypeptides under identical conditions exhibited thermally induced gelation, resulting in physically crosslinked hydrogels. They showed controlled rheological and mechanical properties depending on the conformational change of the CalM middle block induced by binding either Ca2+ or Ca2+ and trifluoperazines (TFPs) as ligands. In addition, both Ca2+-free and Ca2+-bound ECE triblock copolypeptide hydrogels exhibited biocompatibility, while those bound to both Ca2+ and TFPs showed severe cytotoxicity because of controlled TFP release of the CalM blocks. The ECE triblock hydrogels with stimuli responsiveness would be useful as injectable drug delivery depots for biomedical applications.

Injectable organo-hydrogels influenced by click chemistry as a paramount stratagem in the conveyor belt of pharmaceutical revolution

“…Depending on the conformational change of the CalM middle block brought about by binding either Ca 2+ or Ca 2+ and trifluoperazines (TFPs) as ligands, they displayed controlled rheological and mechanical properties. 65 Turabee et al aimed at delivering protein molecules without deteriorating their inherent bioactivity through fabrication of a temperature- and pH-responsive bioresorbable injectable hydrogel system composed of polypeptide block copolymers, namely polyethylene glycol (PEG), temperature-responsive poly(γ-benzyl- l -glutamate) (PBLG), and pH-responsive oligo(sulfamethazine) (OSM), for the sustained delivery of proteins in vivo . The anionic properties of the scaffold allowed for the efficient loading of the model protein-lysozyme and when injected into the Sprague-Dawley rats subcutaneously, it was observed that the lysozyme loaded block copolymer formed a viscoelastic gel in vivo which could deliver the model protein in a sustained manner for at least a week.…”

Peptide–polyurea hybrids: a platform for tunable, thermally-stable, and injectable hydrogels

“…37,38 We conclude that intramolecular α-helices are the driving force for gelation in PZLY-PEG PPU hydrogels via self-assembly of lysine segments, which has been observed in amphiphilic polylysine copolymer systems. 39–41 Simultaneously, cooperative intramolecular bonding of α-helical domains can provide significant mechanical reinforcement, which has been shown to dictate properties, such as moduli and recovery of hybrid networks. 27,42,43…”

Vascular Endothelial Growth Factor Receptor 1 Targeting Fusion Polypeptides with Stimuli-Responsiveness for Anti-angiogenesis

“…The modified pET-21a (+) (mpET-21a) vector with an adaptor DNA sequence was prepared, as previously reported. , The adaptor of mpET-21a has the identical cleavage site of BseRI and AcuI and leaves two different sticky ends of XbaI and BamHI for seamless ligation of EBP n and anti-Flt1-EBP n encoded genes. EBP n with the pentapeptide unit of Val-Pro-Ala-X aa -Gly (VPAXG), where X at the fourth residue is an amino acid except Pro, is composed of n integer repeats of the six pentapeptides with the X composition of Ala:Gly:Ile of 1:4:1.…”

“…They were screened to determine a length of EBP1 gene by agarose gel electrophoresis with double digestion of XbaI and BamHI and confirmed by DNA sequencing. When a series of genes of EBP n ( n = 1–6) were constructed, EBP12 and EBP24 were prepared by serial multimerization of EBP6 gene by modified recursive directional ligation (RDL), as reported. ,, Likewise, to construct various genes encoding anti-Flt1-EBP n ( n = 3, 6, 12, 24), 90 pmol annealed anti-Flt1 peptide-encoded dsDNA with both BseRI and AcuI cohesive ends was incubated with 30 pmol BseRI-restricted EBP n -encoding mpET-21a vector using 1 U T4 DNA ligase for ligation. Both gene and its length of anti-Flt1 peptide and EBP n were confirmed with XbaI and BamHI digestion by agarose gel electrophoresis and DNA sequencing.…”

“…To non-chromatographically purify EBP n or anti-Flt1-EBP n , the E. coli cells were separated by centrifugation at 4500 rpm for 10 min at 4 °C and LCST-based ITC was executed, as reported. ,, In brief, the cells resuspended in 30 mL of PBS were lysed on ice by ultra-sonication with intervals of 10 s on and 20 s off for 5 min (VC-505, Sonic and Materials, Danbury, CT, USA), followed by centrifugation at 13 000 rpm for 15 min at 4 °C. When the insoluble cell debris was precipitated, the supernatant with soluble EBP n or anti-Flt1-EBP n was mixed with polyethyleneimine (0.5% w/v), and the mixture solution was centrifuged under identical conditions to precipitate the nucleic acids.…”