(M1130-09-49) Controlled Release of Proteins with Hydralese^® Formulations for Long-Acting Implants

Purpose: Controlled release of protein therapeutics offers unique advantages like reduced frequency of drug administration which minimizes side effects. This is attributed to the sustained release of the protein while still providing high drug loading to keep it in the optimal therapeutic range. However, developing delivery systems that ensure controlled release while maintaining stability and intended activity remains a considerable challenge. Our group has developed the Hydralese™ platform based on a family of poly(glycerol ester) urethanes (PGEUs), which exhibits a range of mechanical properties and release profiles. Poly(glycerol sebacate) urethane (PGSU) is a PGEU that has been shown to be efficient in release of small molecules. However, drug assay quantification and release of large molecules (e.g., proteins, mAbs, nucleic acids) from PGSU is a challenge because of the hydrophobicity and small mesh size of the cross-linked network. A new generation of PGEUs were developed by our group that provide increased control over the physicochemical characteristics of the polymer such as monomer composition and molecular weight, enabling better release characteristics for large molecules. Lysozyme, a 15 kDa enzyme, was chosen as a model active pharmaceutical ingredient (API) and the optimum release profile as well as drug loading were determined from various PGEU formulations.

Methods: Lysozyme (chicken egg white), purchased from Thermofisher, was milled prior to the formulation to obtain a uniform particle size. Lysozyme-loaded PGEU cylindrical implants were synthesized from poly glycerol esters (PGEs) cross-linked with a diisocyanate cross-linker in the presence of a tin catalyst; implants were manufactured by reaction injection molding. The implants were cured at room temperature before demolding. Three distinct PGEU cylindrical implant (40 mm in length and 3 mm in diameter) formulations with 40% drug loading (w/w) were manufactured. Total drug assay was performed by extracting lysozyme into dimethyl sulfoxide (DMSO). In vitro release (IVR) studies were performed in 10 mM PBS at pH 7.4 in an incubator shaker at 37°C. IVR samples were collected on Day 1 and weekly thereafter. Full media exchanges were performed at every time point, and the lysozyme release was quantified using high performance liquid chromatography (HPLC).

Results: Ideal protein delivery formulations should be able to provide controlled release for an extended period of time and maintain the necessary stability of the formulated protein. To explore the capability of our group’s Hydralese platform for protein release, we tested different urethane-based polymers for the release of lysozyme; the details of one formulation are presented below. SEM imaging was used to examine the implants post-curing. Lateral and cross-sectional images of the formulation revealed no obvious defects in the implants (Figure 1). Total drug content within the implants was quantified using extraction in DMSO, resulting in 100% extraction of lysozyme within 24 hours as measured by HPLC (data not shown). After extraction, three rods from the formulation were selected for an in vitro release study to evaluate the release profile of lysozyme (Figure 2). A linear release rate was observed for the first 9 weeks followed by gradual reduction for the next 9 weeks. A cumulative release of 92.5 mg (61.75 %), 97.02 mg (68.47%), and 81.87 mg (55.28%) were observed for rods R2, R6, and R10, respectively.

Conclusion: Even as effective biotherapeutics, the full clinical potential of proteins has not yet been realized due to various challenges that still must be addressed (e.g., poor stability and activity, short half-lives, low permeability, and the need for frequent administration). Technologies that enable the long-term controlled release of proteins alleviate many of these concerns. Polymer-based formulations, with their diverse array of physicochemical characteristics, provide a unique opportunity to address some of these hurdles. PGEUs provide a novel platform technology for delivering proteins, like lysozyme. Various PGEUs with differences in molecular weight and chemical composition were tested for their ability to release lysozyme and achieved a sustained release for up to 9 weeks followed by continued release for another 9 weeks from one particular formulation. Our Hydralese platform provides the basis to study the effects of the polymer size, composition, and degradation rates that play a significant role in the release of biological APIs to improve how we tackle the unique challenges of protein drug delivery.

Figure 1: Lateral and cross-sectional image of a 40% lysozyme-loaded PGEU implant.

Figure 2: IVR release of lysozyme from three rods for a particular PGEU formulation.