Poly(ethylene glycol)-modified proteins: implications for poly(lactide-co-glycolide)-based microsphere delivery

Abstract

The reduced injection frequency and more nearly constant serum concentrations afforded by sustained release devices have been exploited for the chronic delivery of several therapeutic peptides via poly(lactide-co-glycolide) (PLG) microspheres. The clinical success of these formulations has motivated the exploration of similar depot systems for chronic protein delivery; however, this application has not been fully realized in practice. Problems with the delivery of unmodified proteins in PLG depot systems include high initial "burst" release and irreversible adsorption of protein to the biodegradable polymer. Further, protein activity may be lost due to the damaging effects of protein-interface and protein-surface interactions that occur during both microsphere formation and release. Several techniques are discussed in this review that may improve the performance of PLG depot delivery systems for proteins. One promising approach is the covalent attachment of poly(ethylene glycol) (PEG) to the protein prior to encapsulation in the PLG microspheres. The combination of the extended circulation time of PEGylated proteins and the shielding and potential stabilizing effects of the attached PEG may lead to improved release kinetics from PLG microsphere system and more complete release of the active conjugate.

Fig. 1

Simulated serum hGH level versus time profiles showing differences between subcutaneous injections of hGH (plus signs), PEG-modified hGH (crosses), and hGH encapsulated in a PLG depot device (dots). A possible profile for PEG-hGH encapsulated in a PLG depot device (stars) is also presented for illustrative purposes. The profile for an ideal zero-order release is also shown for comparison (solid line). Profiles were generated using a single compartmental kinetic model, with the following kinetic expression: d[hGH]/dt = −(d(hGH)_dose/dt)/v − k _c[hGH], where [hGH], (hGH)_dose, v, and k _c represent the serum hGH concentration at time t, the hGH dose mass at time t, the serum volume, and the clearance rate, respectively. Assumptions made were that the body (serum) is a single compartment, a subcutaneous dose form supplies drug to the serum compartment by either a first- or zero-order process beginning at time zero, and the serum volume is 5 l; the serum concentration is assumed to be negligible at time t = 0. The model for ideal first-order release was −d(hGH)_dose/dt = k _r(hGH)_dose; that for zero-order release had the expression −d(hGH)_dose/dt = k _r. The serum half-lives specified were 2.5 h and 6 days for hGH and PEG-hGH species, respectively (3,5); the relationship between serum half-life and the clearance rate constant is t _1/2 = ln2/k _c. The release rate constants used were 1 h⁻¹, 1 h⁻¹, 0.0832 μg/h, and 0.0016 μg/h for hGH, PEG-hGH, hGH depot, and PEG-hGH depot, respectively. The rates of release were selected in order to achieve therapeutic serum concentrations of ∼0.1 μg/ml, which are scaled from clinical data (–5). Dose mass loads used were 0.6, 0.3, 30, and 30 μg for hGH, PEG-hGH, hGH in depot, and PEG-hGH in depot, respectively

Fig. 2

Chemical structure of poly(lactide-co-glycolide). Indices x and y refer to the relative amounts of lactide and glycolide units, respectively, in a specific PLG copolymer. These indices are incorporated in PLG nomenclature; e.g. an x = 85% lactide, y = 15% glycolide is indicated as 85:15 PLG. x and y can be manipulated to alter the degradation rate of the PLG. An increase in x results in a slower degradation rate

Fig. 3

a In vitro release profile of protein from PLG microspheres carrying native or PEGylated IFN (IFN-mPEG₂₀₀₀ and IFN-mPEG₅₀₀₀) as shown with closed circle, inverted closed triangle, and open circle, respectively [replotted from (15), p. 111, Copyright (2003), with permission from Elsevier]. For the IFN study, 50:50 PLG microspheres had an average size of 1.8, 1.2, and 1.5 μm for those containing IFN, IFN-mPEG₂₀₀₀, and IFN-mPEG₅₀₀₀, respectively (15). b RNase A (filled squares), mono-PEG-RNase A (open triangles), and di-PEG-RNase A (filled circles) release from 85:15 PLG microspheres [replotted from (16), p. 866, Copyright (2005), with permission from John Wiley & Sons, Inc.]. In the RNase A study, 85:15 PLG microspheres, with a broad distribution of diameters ranging from 5 to 50 μm, were used (16)

Fig. 4

Total internal reflection fluorescence-based adsorption kinetic profiles of a 0.685 μM lysozyme solution and b 0.289 μM monoPEG-lysozyme solution in pH 7.4, 5 mM triethanolamine buffer on 85:15 PLG. The average surface concentrations were 0.0146 ± 0.0022 and 0.0023 ± 0.0006 molecules/nm² for unmodified and PEGylated lysozyme, respectively