Drug Delivery Matrix Containing Native Protein Precipitates Suspended in a Poloxamer Gel

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Abstract
Sustained delivery systems can achieve more constant blood levels of protein therapeutics than those obtained with bolus doses, leading to improved drug efficacy and fewer adverse side effects. Several different polymeric delivery systems have been studied, including poloxamers, which are unique because they can be prepared in aqueous buffers that are compatible with proteins. Poloxamers are nontoxic block copolymers of poly(ethylene oxide) and poly(propylene oxide). Certain poloxamers exhibit reversible thermal gelation. Thus, a solution of protein and poloxamer prepared at low temperatures and injected extravascularly will form a gel as it warms to body temperature. Subsequently, the protein is released slowly from the gel. To date, however, poloxamer gel delivery systems have been limited to relatively low protein concentrations (i.e., < or = 0.4 mg/mL) that produce a completely soluble protein and an optically clear gel. Much higher concentrations of other protein drugs might be needed to obtain an efficacious sustained dose. In the current in vitro study we found that a poloxamer 407 (22% wt/wt) matrix could be prepared containing tens of milligrams/mililiter of the model proteins alpha-chymotrypsin and lactate dehydrogenase. Under these conditions the protein forms a homogeneous suspension. Warming through the poloxamer 407 transition temperature (ca. 18 degrees C) results in a gel that retains a homogeneous distribution of protein precipitates for several days at 37 degrees C. Infrared spectroscopy documented that the precipitated proteins in the suspension have native secondary structure. Furthermore, the fully active protein can be recovered completely when the gel is dissolved in excess buffer. Finally, at the higher protein concentrations used to form the suspensions in poloxamer 407, protein stability during incubation at 37 degrees C was greatly improved over that seen at lower protein concentrations.