Design and characterization of bioengineered recombinant protein-based particles DM4R2 for chemotherapeutic agent and protein drug delivery

Spider silks are attractive materials due to their impressive material properties and their ability to self-assemble into a variety of morphological biomaterials for use as carriers for drug delivery. Previously, the M4R2 silk particles, self-assembled from recombinant spider silk protein, have demonstrated efficient loading capacity for positively charged antitumor polypeptides. However, the colloidal stability of these silk particles and their affinity for drugs still require improvement. In this study, we designed and produced a new bioengineered spider silk protein (DM4R2) by incorporating an aspartic acid residue. The DM4R2 particles exhibited a more negative charge and greater stability compared to M4R2 particles. Meanwhile, the DM4R2 particles showed no cytotoxicity and possessed a similar secondary structure composition to that of M4R2 particles. Although MTX and lysozyme were efficiently loaded onto both types of particles with up to 90% efficiency through strong electrostatic interactions, a maximum of nearly 50% MTX loading onto DM4R2 particles (loading capacity) was observed, compared to only ~ 35% MTX loading onto M4R2 particles. Furthermore, a significantly higher release efficiency of drugs from DM4R2 particles at pH 4.5 was observed, while at pH 5.5, M4R2 particles showed greater release, consistent with their respective isoelectric points and electrostatic binding affinities. Overall, the reasonable loading capacity and high loading efficiency, along with the desired release efficiency, make DM4R2 particles the promising carriers for delivering small-molecule chemotherapeutic drugs and protein drugs.