Sustained release of antibiotics from injectable and thermally responsive polypeptide depots

Abstract

Biodegradable polymeric scaffolds are of interest for delivering antibiotics to local sites of infection in orthopaedic applications, such as bone and diarthrodial joints. The objective of this study was to develop a biodegradable scaffold with ease of drug loading in aqueous solution, while providing for drug depot delivery via syringe injection. Elastin-like polypeptides (ELPs) were used for this application, biopolymers of repeating pentapeptide sequences that were thermally triggered to undergo in situ depot formation at body temperature. ELPs were modified to enable loading with the antibiotics, cefazolin, and vancomycin, followed by induction of the phase transition in vitro. Cefazolin and vancomycin concentrations were monitored, as well as bioactivity of the released antibiotics, to test an ability of the ELP depot to provide for prolonged release of bioactive drugs. Further tests of formulation viscosity were conducted to test suitability as an injectable drug carrier. Results demonstrate sustained release of therapeutic concentrations of bioactive antibiotics by the ELP, with first-order time constants for drug release of approximately 25 h for cefazolin and approximately 500 h for vancomycin. These findings illustrate that an injectable, in situ forming ELP depot can provide for sustained release of antibiotics with an effect that varies across antibiotic formulation. ELPs have important advantages for drug delivery, as they are known to be biocompatible, biodegradable, and elicit no known immune response. These benefits suggest distinct advantages over currently used carriers for antibiotic drug delivery in orthopedic applications.

Figure 1

Release of cefazolin from ELP constructs prepared at 150mg/ml ELP (Group I samples). Data presented as averages and standard deviations for n=5 per group.

Figure 2

Release of vancomycin from ELP constructs prepared at 150mg/ml ELP (Group I samples). Data presented as averages and standard deviations for n=5 per group.

Figure 3

Release of vancomycin from 5 mg drug-loaded ELP constructs prepared at 150mg/ml ELP (Group I samples) and 225 mg/ml ELP. Data presented as averages and standard deviations for n=5 per group.

Figure 4

Representative agar plate assay showing method for determining the zone of inhibition (ZOI) following antibiotic placement upon substrates cultured with B. subtillis. Plate shown on left illustrates ZOI for calibrating solutions of vancomycin, along with ELP control samples cultured for periods of 1 to 28 days (italic labels). The white ring (with arrow) illustrates a typical measurement of diameter for ZOI calculation. Panels on right illustrate B. subtillis cultures after exposure to supernatant from vancomycin-loaded ELP constructs after 7 to 28 days of culture. Comparison of white ring sizes demonstrates that activity of supernatants was equivalent to 0.05-0.1 mg/ml vancomycin.

Figure 5

Dynamic viscosity (|η*|) measures for a representative ELP construct, before and after preparation by THPP crosslinking (150 mg/ml). A frequency dependence was observed for the crosslinked constructs, but not the uncrosslinked ELP.