Efficient Dual siRNA and Drug Delivery Using Engineered Lipoproteoplexes

An engineered supercharged coiled-coil protein (CSP) and the cationic transfection reagent Lipofectamine 2000 are combined to form a lipoproteoplex for the purpose of dual delivery of siRNA and doxorubicin. CSP, bearing an external positive charge and axial hydrophobic pore, demonstrates the ability to condense siRNA and encapsulate the small-molecule chemotherapeutic, doxorubicin. The lipoproteoplex demonstrates improved doxorubicin loading relative to Lipofectamine 2000. Furthermore, it induces effective transfection of GAPDH (60% knockdown) in MCF-7 breast cancer cells with efficiencies comparing favorably to Lipofectamine 2000. When the lipoproteoplex is loaded with doxorubicin, the improved doxorubicin loading (∼40 μg Dox/mg CSP) results in a substantial decrease in MCF-7 cell viability

Protein Engineered Triblock Polymers Composed of Two SADs: Enhanced Mechanical Properties and Binding Abilities

Recombinant methods have been used to engineer artificial protein triblock polymers composed of two <i>different</i> self-assembling domains (SADs) bearing one elastin (E) flanked by two cartilage oligomeric matrix protein coiled-coil (C) domains to generate CEC. To understand how the two C domains improve small molecule recognition and the mechanical integrity of CEC, we have constructed C_L44AEC_L44A, which bears an impaired C_L44A domain that is unstructured as a negative control. The CEC triblock polymer demonstrates increased small molecule binding and ideal elastic behavior for hydrogel formation. The negative control C_L44AEC_L44A does not exhibit binding to small molecule and is inelastic at lower temperatures, affirming the favorable role of C domain and its helical conformation. While both CEC and C_L44AEC_L44A assemble into micelles, CEC is more densely packed with C domains on the surface enabling the development of networks leading to hydrogel formation. Such protein engineered triblock copolymers capable of forming robust hydrogels hold tremendous promise for biomedical applications in drug delivery and tissue engineering