ReviewPost screenPharmacokinetics, metabolism and distribution of PEGs and PEGylated proteins: quo vadis?
Introduction
Polyethylene glycols (PEGs) are synthetic water-soluble polymers of varying chain lengths, structures and molecular weights. The PEGylation technology has been widely used to modify (bio)pharmaceuticals chemically to decrease their in vivo clearance and also potentially to ameliorate the immunogenicity of highly immunogenic proteins such as enzymes 1, 2. A decrease in clearance of the PEGylated moiety is observed when one PEG chain with a molecular weight (MW) of 30 kDa or a branched PEG chain with two 20 kDa chains or several chains of 5 kDa 3, 4, 5 are attached to the peptide or protein. The animal and human pharmacokinetics (PK) of a PEGylated peptide or protein are usually well characterized during the development process before submission to the health authorities [6]. By contrast, information about the PK and distribution of the higher MW PEG part is usually sparse. Although there is no evidence for PEG-related clinical safety issues with PEGs and PEGylated proteins 7, 8, 9, there have been reports of increased vacuolation in tissues from animals that have been administered with PEGylated proteins 10, 11. Repeated parenteral administration of PEGylated proteins to animals has been associated with cellular vacuolation in macrophages and/or histiocytes in various organs and in other cells such as renal tubular cells and ependymal cells of the choroid plexus 9, 11, 12 (http://www.ema.europa.eu/ema/index.jsp?curl=pages/medicines/human/medicines/001037/human_med_001294.jsp&mid=WC0b01ac058001d125; http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2012/11/WC500135123.pdf; http://www.ema.europa.eu/docs/en_GB/document_library/EPAR_-_Summary_for_the_public/human/001037/WC500069733.pdf; http://www.ema.europa.eu/docs/en_GB/document_library/EPAR_-_Summary_for_the_public/human/000409/WC500054622.pdf; http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2012/11/WC500135123.pdf).
To make an informed risk:benefit assessment of PEGylated proteins in the clinic, information about the PK, metabolism and biodistribution of PEG and PEGylated proteins is important, including knowledge about the cellular uptake mechanisms for these products. This will also help to establish a relationship to the functional effects, especially when administered chronically and/or in a pediatric population. The present article reviews the existing information related to the PK, metabolism and biodistribution of PEG and PEGylated proteins. We also highlight some of the discrepancies and gaps in our knowledge in this area. Together with what is known or not known so far about the PK and biodistribution of PEGylated proteins, we attempt to provide an understanding of the mechanism(s) of vacuolization in tissues after administration of PEGylated proteins.
The PK, metabolism and biodistribution are reviewed and discussed in separate sections for PEG alone and PEGylated proteins. It is important to note that the PK profiles of the conjugated protein, the naked protein (with and without linker) and the PEG generated from the PEGylated protein (hereafter referred to as ‘metabolic’ PEG) are similar and sometimes overlap, and cannot be clearly delineated when considering the fate of PEGylated proteins.
Section snippets
PK and metabolism of PEGs
High MW PEGs (PEGs > 30 kDa) are usually mixtures of PEG with a range of randomly varying chain lengths. Linear and branched PEGs, the latter containing two PEG molecules attached to a central core, have been used for PEGylation of proteins 1, 13. More recently, monomolecular PEG has also been used to modify proteins [14].
Low MW PEGs (e.g. PEG300 up to 65%, PEG3350 up to 6%) are frequently used as pharmaceutical excipients, including in intravenous formulations. An oral administration of 17 g
PK and metabolism of PEGylated proteins
PEGylated proteins are usually absorbed at a slower rate after subcutaneous administration than nonPEGylated proteins, with a time to maximum concentration (Tmax) in the range of 1–3 days and an absolute bioavailability of 50–60% in the case of PEGylated erythropoietin (http://www.accessdata.fda.gov/drugsatfda_docs/nda/2007/125164TOC.cfm) but a Tmax of 2–7 days and a bioavailability of 80% in the case of certolizumab pegol (Cimzia®) (//www.ema.europa.eu/ema/index.jsp%3Fcurl=pages/medicines/human/medicines/001037/human_med_001294.jsp%26mid=WC0b01ac058001d125
Biodistribution of PEGs
There are limited published studies on the tissue or cellular distribution of PEG following systemic administration. Moreover, the material and the methodologies used to study tissue distribution were different, making it difficult to draw conclusions across studies (see below: Bioanalytics of PEGs and PEGylated proteins). On the basis of our review of the literature to date, either the PEGylated peptide or protein 18, 23, 24, 25 and/or the PEG alone 1, 3, 12, 17 were administered to study the
Biodistribution of administered PEG
Longley et al. [17] used a linear 40 kDa [14C]-PEG-Ala with a 14C radiolabel that was incorporated into the ether backbone of PEG. The radioactivity was widely distributed to most tissues following administration of 1250 mg/kg PEG–Ala, and concentrations decreased over time. The maximum concentration was observed in the kidney [9.5% injected dose (ID)/gram tissue]. The lung, heart and liver concentrations ranged from 2.6 to 1.5% ID/g (in descending order), whereas concentrations in the
Biodistribution of metabolic PEG
The distribution of metabolic PEG following administration of PEGylated peptides or proteins has been reported in a few studies 18, 23, 24, 25. Wang et al. [23] studied the distribution of 40 kDa 14C-PEGylated adnectin (20 kDa PEG × 2) in mice. At 2 hours the radioactivity was high in highly perfused organs such as lung, liver, heart and kidney. By 53 hours maximum radioactivity was observed in the liver, followed by the kidney. Concentrations in other tissues such as brain and muscle were very
Biodistribution of PEGylated proteins
The biodistribution of PEGylated proteins appears to be largely driven by the properties of the PEG and the protein, although the dominant mechanism for any given PEGylated molecule is not yet clear. Stork et al. [27] reported increased uptake of 125I-scDb-Aʹ-40 kDa PEG [a single chain diabody that binds to carcinoembryonic antigen (CEA) and CD3 and is conjugated to 40 kDa PEG] compared with 125I-scDb in CEA-positive tumors present in mouse xenograft models. After 4 hours, distribution of 125
Bioanalytics of PEG and PEGylated proteins
The bioanalysis of PEG is challenging because it has no chromophore and radiolabeling is usually confined to the penultimate structure, which is more subject to metabolism. Therefore, measurements of PEG usually refer to the total of PEG-like structures without confirmation that the chain length is maintained. High MW PEGs are recalcitrant to mass spectrometry analysis owing to their polydispersed nature and the difficulty of ionization. Therefore, in the past, the metabolic fate of PEG
Discussion
The PK of PEGylated proteins is initially driven by the two major parts of the molecule, the protein itself and its conjugated PEG part. When the PEG is cleaved from the molecule resulting in the formation of metabolic PEG the biodistribution and PK are then governed by PEG-related mechanisms.
The earlier belief that the (hydrated) PEG at a MW of about 30 kDa is large enough to no longer be efficiently filtered by renal glomeruli [5] might need to be modified based on more-recent data 17, 22. An
Concluding remarks
The safety and efficacy of PEG and PEG proteins is demonstrated with the approval of more than ten drugs and devices [33]. Although there is no evidence for PEG-related safety issues with PEGylated proteins in the clinic, questions relating to the disposition of PEG are now being raised more frequently by health authorities, particularly for PEGylated proteins used chronically and/or in the pediatric population (//www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2012/11/WC500135123.pdf
Acknowledgements
The authors would like to thank colleagues from Genentech, Bayer and Roche for valuable comments during review of this manuscript. The authors would also like to thank Allison Bruce from Genentech for drawing the figures and Anshin BioSolutions for their help in editing the manuscript.
References (34)
- et al.
PEGylation, successful approach to drug delivery
Drug Discov. Today
(2005) Distribution and tissue uptake of poly(ethylene glycol) with different molecular weights after intravenous administration to mice
J. Pharm. Sci.
(1994)Prolongation of the serum half-life period of superoxide dismutase by poly (ethylene glycol) modification
J. Control. Release
(1997)- et al.
Pharmacokinetic and biodistribution properties of poly(ethylene glycol)–protein conjugates
Adv. Drug Deliv. Rev.
(2003) Safety assessment on polyethylene glycols (PEGs) and their derivatives as used in cosmetic products
Toxicology
(2005)Short communication: renal tubular vacuolation in animals treated with polyethylene-glycol-conjugated proteins
Toxicol. Sci.
(1998)Biodistribution and excretion of radiolabeled 40 kDa PEG following intravenous administration in mice
J. Pharm. Sci.
(2013)Investigation of the distribution and elimination of the PEG component of certolizumab pegol in rats
J. Crohn's Colitis. Suppl.
(2008)Administration, distribution, metabolism and elimination of polymer therapeutics
J. Control. Release
(2012)Biodistribution of a bispecific single-chain diabody and its half-life extended derivatives
J. Biol. Chem.
(2009)
Current challenges and opportunities in nonclinical safety testing of biologics
Drug Discov. Today
New challenges and opportunities in nonclinical safety testingof biologics
Regulatory Toxicology and Pharmacology
Pegloticase: in treatment-refractory chronic gout
Drugs
PEGylated therapeutic proteins for haemophilia treatment. A review for haemophilia caregivers
Haemophilia
PEGylated proteins: evaluation of their safety in the absence of definitive metabolism studies
Drug Metab. Dispos.
PEG and PEG conjugate toxicity: towards an understanding of the toxicity of PEG and its relevance to PEGylated biological
Cited by (120)
Ultrasound robotics for precision therapy
2024, Advanced Drug Delivery ReviewsBiodegradable trimethyl chitosan nanofiber mats by electrospinning as bioabsorbable dressings for wound closure and healing
2023, International Journal of Biological MacromoleculesPoly(2-oxazoline)-derived star-shaped polymers as potential materials for biomedical applications: A review
2023, European Polymer JournalFactors affecting peptide and protein absorption, metabolism, and excretion
2023, Peptide and Protein Drug Delivery Using PolysaccharidesRecent advances in degradable synthetic polymers for biomedical applications - Beyond polyesters
2022, Progress in Polymer ScienceToxicity of high-molecular-weight polyethylene glycols in Sprague Dawley rats
2022, Toxicology LettersCitation Excerpt :Little is known about the tissue distribution and toxicity of the higher molecular weight PEGs. High-molecular-weight PEGs show slow renal clearance and consequently have a greater potential to accumulate within cells (Baumann et al., 2014; Caliceti and Veronese, 2003). The accumulation of PEGs in cells has been linked to cellular vacuolation in five of nine approved PEGylated biologicals (Ivens et al., 2015).