In vitro degradation and erosion behavior of commercial PLGAs used for controlled drug delivery.

Copolymers of lactic (or lactide) and glycolic (or glycolide) acids (PLGAs) are among the most commonly used materials in biomedical applications, such as parenteral controlled drug delivery, due to their biocompatibility, predictable degradation rate, and ease of processing. Besides manufacturing variables of drug delivery vehicles, changes in PLGA raw material properties can affect product behavior. Accordingly, an in-depth understanding of polymer-related "critical quality attributes" can improve selection and predictability of PLGA performance. Here, we selected 19 different PLGAs from five manufacturers to form drug-free films, submillimeter implants, and microspheres and evaluated differences in their water uptake, degradation, and erosion during in vitro incubation as a function of L/G ratio, polymerization method, molecular weight, end-capping, and geometry. Uncapped PLGA 50/50 films from different manufacturers with similar molecular weights and higher glycolic unit blockiness and/or block length values showed faster initial degradation rates. Geometrically, larger implants of 75/25, uncapped PLGA showed higher water uptake and faster degradation rates in the first week compared to microspheres of the same polymers, likely due to enhanced effects of acid-catalyzed degradation from PLGA acidic byproducts unable to escape as efficiently from larger geometries. Manufacturer differences such as increased residual monomer appeared to increase water uptake and degradation in uncapped 50/50 PLGA films and poly(lactide) implants. This dataset of different polymer manufacturers could be useful in selecting desired PLGAs for controlled release applications or comparing differences in behavior during product development, and these techniques to further compare differences in less reported properties such as sequence distribution may be useful for future analyses of PLGA performance in drug delivery.

Optimizing zinc-HisTag coordination remote loading of proteins in PLGA microspheres.

Metal-HisTag coordination remote loading (MHCRL) of proteins in PLGA microspheres was previously developed to provide a useful tool for discovery and preclinical development of controlled release protein formulations. Here we describe optimization of MHCRL, including (1) reducing thermal stress, (2) decreasing the complexity and duration of the procedure, (3) increasing loading capacity, (4) increasing the penetration depth of protein, and (5) improving the release profile. Directly encapsulating ZnCO3as a Zn2+source for HisTag coordination, rather than remotely loading Zn2+, increased the Zn content ∼6-fold. Microspheres with directly encapsulated ZnCO3more deeply encapsulated green fluorescent protein and more efficiently encapsulated human serum albumin at protein loading solutions concentrations ≥100 µg/mL than remotely loaded Zn2+microspheres. Tributyl acetylcitrate plasticized microspheres in terms of decreasingTg, but led to a decrease in protein encapsulation efficiency. As such, the plasticizer was not deemed useful. The loading/healing cycles were reduced in time and temperature from 48 h/42 h at 43 °C to 2 h/6h at 37 °C while maintaining strong encapsulation efficiency, resulting in significantly improved protein stability. Immunoreactive protein was slowly released for months following a modest burst release. The improved microspheres and shorter, low-temperature encapsulation could be a valuable asset to drug discovery scientists interested in controlled release of delicate and/or costly biologic candidates.

Metal-HisTag coordination for remote loading of very small quantities of biomacromolecules into PLGA microspheres.

Challenges to discovery and preclinical development of long-acting release systems for protein therapeutics include protein instability, use of organic solvents during encapsulation, specialized equipment and personnel, and high costs of proteins. We sought to overcome these issues by combining remote-loading self-healing encapsulation with binding HisTag protein to transition metal ions. Porous, drug-free self-healing microspheres of copolymers of lactic and glycolic acids with high molecular weight dextran sulfate and immobilized divalent transition metal (M2+) ions were placed in the presence of proteins with or without HisTags to bind the protein in the pores of the polymer before healing the surface pores with modest temperature. Using human serum albumin, insulin-like growth factor 1, and granulocyte-macrophage colony-stimulating factor (GM-CSF), encapsulated efficiencies of immunoreactive protein relative to nonencapsulation protein solutions increased from ~41%, ~23%, and ~9%, respectively, without Zn2+ and HisTags to ~100%, ~83%, and ~75% with Zn2+ and HisTags. These three proteins were continuously released in immunoreactive form over seven to ten weeks to 73%-100% complete release, and GM-CSF showed bioactivity >95% relative to immunoreactive protein throughout the release interval. Increased encapsulation efficiencies were also found with other divalent transition metals ions (Co2+, Cu2+, Ni2+, and Zn2+), but not with Ca2+. Ethylenediaminetetraacetic acid was found to interfere with this process, reverting encapsulation efficiency back to Zn2+-free levels. These results indicate that M2+-immobilized self-healing microspheres can be prepared for simple and efficient encapsulation by simple mixing in aqueous solutions. These formulations provide slow and continuous release of immunoreactive proteins of diverse types by using a amount of protein (e.g., <10 µg), which may be highly useful in the discovery and early preclinical development phase of new protein active pharmaceutical ingredients, allowing for improved translation to further development of potent proteins for local delivery.