Therapeutic protein-polymer conjugates: advancing beyond PEGylation.

Protein-polymer conjugates are widely used as therapeutics. All Food and Drug Administration (FDA)-approved protein conjugates are covalently linked to poly(ethylene glycol) (PEG). These PEGylated drugs have longer half-lives in the bloodstream, leading to less frequent dosing, which is a significant advantage for patients. However, there are some potential drawbacks to PEG that are driving the development of alternatives. Polymers that display enhanced pharmacokinetic properties along with additional advantages such as improved stability or degradability will be important to advance the field of protein therapeutics. This perspective presents a summary of protein-PEG conjugates for therapeutic use and alternative technologies in various stages of development as well as suggestions for future directions. Established methods of producing protein-PEG conjugates and new approaches utilizing controlled radical polymerization are also covered.

[1]  N. Vorsa,et al.  Dendrimer versus linear conjugate: Influence of polymeric architecture on the delivery and anticancer effect of paclitaxel. , 2006, Bioconjugate chemistry.

[2]  James R. Baker,et al.  Protein Modification, Bioconjugation, and Disulfide Bridging Using Bromomaleimides , 2010, Journal of the American Chemical Society.

[3]  Lei Tao,et al.  Branched polymer-protein conjugates made from mid-chain-functional P(HPMA). , 2009, Biomacromolecules.

[4]  G. Gregoriadis,et al.  Polysialylated insulin: synthesis, characterization and biological activity in vivo. , 2003, Biochimica et biophysica acta.

[5]  R. Shorr,et al.  Physiological effect of polyethylene glycol conjugation on stroma-free bovine hemoglobin in the conscious dog after partial exchange transfusion. , 2008, Artificial organs.

[6]  U. Schubert,et al.  Poly(ethylene glycol) in drug delivery: pros and cons as well as potential alternatives. , 2010, Angewandte Chemie.

[7]  Carmen Scholz,et al.  Tailored polymer architectures for pharmaceutical and biomedical applications , 2013 .

[8]  J. Leroux,et al.  Sustained gastrointestinal activity of dendronized polymer-enzyme conjugates. , 2013, Nature chemistry.

[9]  R. Shorr,et al.  Effect of polyethylene glycol conjugated bovine hemoglobin in both top-load and exchange transfusion rat models. , 2008, Artificial organs.

[10]  Krzysztof Matyjaszewski,et al.  Controlled/"living" radical polymerization. atom transfer radical polymerization in the presence of transition-metal complexes , 1995 .

[11]  F. Veronese,et al.  Polyoxazoline: chemistry, properties, and applications in drug delivery. , 2011, Bioconjugate chemistry.

[12]  S. Evans,et al.  Site-directed conjugation of "clicked" glycopolymers to form glycoprotein mimics: binding to mammalian lectin and induction of immunological function. , 2007, Journal of the American Chemical Society.

[13]  H. Maeda,et al.  Macromolecular Therapeutics , 2003, Clinical pharmacokinetics.

[14]  D. Gigmes,et al.  Degradable and comb-like PEG-based copolymers by nitroxide-mediated radical ring-opening polymerization. , 2013, Biomacromolecules.

[15]  R. Müller,et al.  Particle size, surface hydrophobicity and interaction with serum of parenteral fat emulsions and model drug carriers as parameters related to RES uptake. , 1992, Clinical nutrition.

[16]  J. Choi,et al.  Poly(ethylene oxide sulfide): new poly(ethylene glycol) derivatives degradable in reductive conditions. , 2005, Biomacromolecules.

[17]  Neel S. Joshi,et al.  N-terminal protein modification through a biomimetic transamination reaction. , 2006, Angewandte Chemie.

[18]  J. Kizhakkedathu,et al.  Influence of architecture of high molecular weight linear and branched polyglycerols on their biocompatibility and biodistribution. , 2012, Biomaterials.

[19]  Francesco M Veronese,et al.  PEGylation, successful approach to drug delivery. , 2005, Drug discovery today.

[20]  David R. Liu,et al.  Enzyme-Free Translation of DNA into Sequence-Defined Synthetic Polymers Structurally Unrelated to Nucleic Acids , 2013, Nature chemistry.

[21]  K. Ulbrich,et al.  Polymeric drugs based on conjugates of synthetic and natural macromolecules. I. Synthesis and physico-chemical characterisation. , 2000, Journal of controlled release : official journal of the Controlled Release Society.

[22]  T. Allen,et al.  Evaluation of blood clearance rates and biodistribution of poly(2-oxazoline)-grafted liposomes. , 1996, Journal of pharmaceutical sciences.

[23]  J. Adamson,et al.  Design and Chemical Synthesis of a Homogeneous Polymer-Modified Erythropoiesis Protein , 2003, Science.

[24]  J. Singer,et al.  Biological and clinical characterization of paclitaxel poliglumex (PPX, CT-2103), a macromolecular polymer–drug conjugate , 2006, International journal of nanomedicine.

[25]  Lei Tao,et al.  Differences in cytotoxicity of poly(PEGA)s synthesized by reversible addition-fragmentation chain transfer polymerization. , 2009, Chemical communications.

[26]  H. Maynard,et al.  In situ preparation of protein-"smart" polymer conjugates with retention of bioactivity. , 2005, Journal of the American Chemical Society.

[27]  R. Duncan The dawning era of polymer therapeutics , 2003, Nature Reviews Drug Discovery.

[28]  L. Kiessling,et al.  Recognition Specificity of Neoglycopolymers Prepared by Ring-Opening Metathesis Polymerization , 1996 .

[29]  Luiz A Canalle,et al.  Polypeptide-polymer bioconjugates. , 2010, Chemical Society reviews.

[30]  M. Essler,et al.  Synthesis, biodistribution and excretion of radiolabeled poly(2-alkyl-2-oxazoline)s. , 2007, Journal of controlled release : official journal of the Controlled Release Society.

[31]  D. Brooks,et al.  In vivo biological evaluation of high molecular weight hyperbranched polyglycerols. , 2007, Biomaterials.

[32]  N. Adkinson,et al.  Hypersensitivity to Polyethylene Glycols , 2013, Journal of clinical pharmacology.

[33]  En-Wei Lin,et al.  Trehalose glycopolymers as excipients for protein stabilization. , 2013, Biomacromolecules.

[34]  J. B. Jones,et al.  Glycodendriproteins: a synthetic glycoprotein mimic enzyme with branched sugar-display potently inhibits bacterial aggregation. , 2004, Journal of the American Chemical Society.

[35]  M. Textor,et al.  Poly-2-methyl-2-oxazoline: a peptide-like polymer for protein-repellent surfaces. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[36]  M. Morbidelli,et al.  Process for protein PEGylation. , 2014, Journal of controlled release : official journal of the Controlled Release Society.

[37]  A. N. Boyce,et al.  The effects of naphthaleneacetic acid and gibberellic acid in prolonging bract longevity and delaying discoloration of bougainvillea spectabilis , 2009 .

[38]  F. Davis,et al.  Alteration of immunological properties of bovine serum albumin by covalent attachment of polyethylene glycol. , 1977, The Journal of biological chemistry.

[39]  Huub Schellekens,et al.  The Immunogenicity of Polyethylene Glycol: Facts and Fiction , 2013, Pharmaceutical Research.

[40]  David J Brayden,et al.  Site-specific N-terminus conjugation of poly(mPEG1100) methacrylates to salmon calcitonin: synthesis and preliminary biological evaluation , 2009 .

[41]  J. Loo,et al.  A Heparin-Mimicking Polymer Conjugate Stabilizes Basic Fibroblast Growth Factor (bFGF) , 2013, Nature chemistry.

[42]  R. Shorr,et al.  Transitional vacuole formation following a bolus infusion of PEG-hemoglobin in the rat. , 1996, Artificial cells, blood substitutes, and immobilization biotechnology.

[43]  R. Duncan,et al.  The effect of dextrin-rhEGF on the healing of full-thickness, excisional wounds in the (db/db) diabetic mouse. , 2011, Journal of controlled release : official journal of the Controlled Release Society.

[44]  N. Smorodinsky,et al.  A conjugate between a purified timothy allergen and poly(N-vinylpyrrolidone) suppresses the specific IgE response in mice , 1981 .

[45]  R. Shorr,et al.  The impact of polyethylene glycol conjugation on bovine hemoglobin's circulatory half-life and renal effects in a rabbit top-loaded transfusion model. , 2008, Artificial organs.

[46]  Lei Tao,et al.  Design and synthesis of N-maleimido-functionalized hydrophilic polymers via copper-mediated living radical polymerization: a suitable alternative to PEGylation chemistry. , 2005, Journal of the American Chemical Society.

[47]  Daniela C Dieterich,et al.  Selective identification of newly synthesized proteins in mammalian cells using bioorthogonal noncanonical amino acid tagging (BONCAT). , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[48]  C. Barner‐Kowollik,et al.  Well-defined protein-polymer conjugates via in situ RAFT polymerization. , 2007, Journal of the American Chemical Society.

[49]  Beat Ernst,et al.  Drug discovery today. , 2003, Current topics in medicinal chemistry.

[50]  N. Lameire,et al.  What’s new in the controversy on the renal/tissue toxicity of starch solutions? , 2014, Intensive Care Medicine.

[51]  P. Stayton,et al.  pH-responsive polymer–antigen vaccine bioconjugates , 2011 .

[52]  F. Veronese,et al.  Selective conjugation of poly(2-ethyl 2-oxazoline) to granulocyte colony stimulating factor. , 2012, Journal of controlled release : official journal of the Controlled Release Society.

[53]  J. Chiefari,et al.  Living free-radical polymerization by reversible addition - Fragmentation chain transfer: The RAFT process , 1998 .

[54]  Shaoyi Jiang,et al.  Poly(zwitterionic)protein conjugates offer increased stability without sacrificing binding affinity or bioactivity. , 2011, Nature chemistry.

[55]  P. Caliceti,et al.  Branched and Linear Poly(Ethylene Glycol): Influence of the Polymer Structure on Enzymological, Pharmacokinetic, and Immunological Properties of Protein Conjugates , 1997 .

[56]  M. Finn,et al.  Virus-glycopolymer conjugates by copper(I) catalysis of atom transfer radical polymerization and azide-alkyne cycloaddition. , 2005, Chemical communications.

[57]  G. Mantovani,et al.  Living Radical Polymerization as a Tool for the Synthesis of Polymer‐Protein/Peptide Bioconjugates , 2007 .

[58]  Holger Frey,et al.  Beyond poly(ethylene glycol): linear polyglycerol as a multifunctional polyether for biomedical and pharmaceutical applications. , 2014, Biomacromolecules.

[59]  L. Illum,et al.  Human serum albumin as a probe for surface conditioning (opsonization) of block copolymer-coated microspheres. , 1992, Biomaterials.

[60]  Nicholas A Peppas,et al.  Opsonization, biodistribution, and pharmacokinetics of polymeric nanoparticles. , 2006, International journal of pharmaceutics.

[61]  H. Frey,et al.  Universal concept for the implementation of a single cleavable unit at tunable position in functional poly(ethylene glycol)s. , 2013, Biomacromolecules.

[62]  K. Matyjaszewski,et al.  Synthesis of uniform protein-polymer conjugates. , 2005, Biomacromolecules.

[63]  H. Maynard,et al.  Synthesis of protein-polymer conjugates. , 2007, Organic & biomolecular chemistry.

[64]  E. Åkerblom,et al.  Polyethylene glycol reactive antibodies in man: titer distribution in allergic patients treated with monomethoxy polyethylene glycol modified allergens or placebo, and in healthy blood donors. , 1984, International archives of allergy and applied immunology.

[65]  R. Duncan Polymer therapeutics as nanomedicines: new perspectives. , 2011, Current opinion in biotechnology.

[66]  P. Caliceti,et al.  In situ growth of side-chain PEG polymers from functionalized human growth hormone-a new technique for preparation of enhanced protein-polymer conjugates. , 2010, Bioconjugate chemistry.

[67]  K. Ulbrich,et al.  Poly[N-(2-Hydroypropyl)Methacrylamide] Conjugates of Bovine Seminal Ribonuclease. Synthesis, Physicochemical, and Preliminary Biological Evaluation , 2000 .

[68]  A. Baas,et al.  FDA-approved poly(ethylene glycol)–protein conjugate drugs , 2011 .

[69]  H. Maynard,et al.  Streptavidin as a macroinitiator for polymerization: in situ protein-polymer conjugate formation. , 2005, Journal of the American Chemical Society.

[70]  R. Offord,et al.  Site-specific attachment of functionalized poly(ethylene glycol) to the amino terminus of proteins. , 1996, Bioconjugate chemistry.

[71]  S. Brocchini,et al.  Site-specific PEGylation of native disulfide bonds in therapeutic proteins , 2006, Nature chemical biology.

[72]  T. Kogan The Synthesis of Substituted Methoxy-Poly(Ethyleneglycol) Derivatives Suitable for Selective Protein Modification , 1992 .

[73]  P. Caliceti,et al.  Immunological properties of uricase conjugated to neutral soluble polymers. , 2001, Bioconjugate chemistry.

[74]  B. G. Davis,et al.  Allyl sulfides are privileged substrates in aqueous cross-metathesis: application to site-selective protein modification. , 2008, Journal of the American Chemical Society.

[75]  David J Brayden,et al.  PK/PD modelling of comb-shaped PEGylated salmon calcitonin conjugates of differing molecular weights. , 2011, Journal of controlled release : official journal of the Controlled Release Society.

[76]  Harm-Anton Klok,et al.  Peptide/protein-polymer conjugates: synthetic strategies and design concepts. , 2008, Chemical communications.

[77]  G. Shopp,et al.  Short communication: renal tubular vacuolation in animals treated with polyethylene-glycol-conjugated proteins. , 1998, Toxicological sciences : an official journal of the Society of Toxicology.

[78]  H. Maynard,et al.  Cysteine-reactive polymers synthesized by atom transfer radical polymerization for conjugation to proteins. , 2004, Journal of the American Chemical Society.

[79]  Y. Sohn,et al.  Thermogelling poly(ethylene oxide-b-propylene oxide-b-ethylene oxide) disulfide multiblock copolymer as a thiol-sensitive degradable polymer. , 2006, Biomacromolecules.

[80]  K. Matyjaszewski,et al.  Synthesis, characterization, and in vitro cell culture viability of degradable poly(N-isopropylacrylamide-co-5,6-benzo-2-methylene-1,3-dioxepane)-based polymers and crosslinked gels. , 2008, Journal of biomedical materials research. Part A.

[81]  A. Sehon,et al.  Suppression of IgE antibodies to the (4‐hydroxy‐3‐iodo‐5‐nitrophenyl)acetyl (NIP) group and induction of NIP‐specific suppressor cells with NIP‐poly‐N‐vinylpyrrolidone conjugates , 1981, European journal of immunology.

[82]  B. Klumperman,et al.  Polymer-protein conjugates from omega-aldehyde endfunctional poly(N-vinylpyrrolidone) synthesised via xanthate-mediated living radical polymerisation. , 2008, Chemical communications.

[83]  B. Sumerlin,et al.  Temperature-regulated activity of responsive polymer-protein conjugates prepared by grafting-from via RAFT polymerization. , 2008, Journal of the American Chemical Society.

[84]  M. Vicent,et al.  Do HPMA copolymer conjugates have a future as clinically useful nanomedicines? A critical overview of current status and future opportunities. , 2010, Advanced drug delivery reviews.

[85]  F. Veronese,et al.  PEGylated Protein Drugs: Basic Science and Clinical Applications , 2009 .

[86]  S. Brocchini,et al.  Poly(2-methacryloyloxyethyl phosphorylcholine) for protein conjugation. , 2008, Bioconjugate chemistry.

[87]  D. Fischer,et al.  Surface-modified biodegradable albumin nano- and microspheres. II: effect of surface charges on in vitro phagocytosis and biodistribution in rats. , 1998, European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V.

[88]  D. Brems,et al.  Characterization and Stability of N-terminally PEGylated rhG-CSF , 1996, Pharmaceutical Research.

[89]  Andrew G. Spencer,et al.  Synthetic glycoprotein mimics inhibit L-selectin-mediated rolling and promote L-selectin shedding. , 2004, Chemistry & biology.

[90]  H. Klok,et al.  Degradable Polymer Brushes Prepared via Surface-Initiated Controlled Radical Polymerization , 2009 .

[91]  K. Matyjaszewski,et al.  Polymer science : a comprehensive reference , 2012 .

[92]  H. Maynard,et al.  Protein-polymer conjugates: synthetic approaches by controlled radical polymerizations and interesting applications. , 2010, Current opinion in chemical biology.

[93]  Lei Tao,et al.  Synthesis and bioactivity of poly(HPMA)-lysozyme conjugates: the use of novel thiazolidine-2-thione coupling chemistry. , 2009, Organic & biomolecular chemistry.

[94]  A. Chilkoti,et al.  In situ growth of a PEG-like polymer from the C terminus of an intein fusion protein improves pharmacokinetics and tumor accumulation , 2010, Proceedings of the National Academy of Sciences.

[95]  S. Caddick,et al.  Polymeric dibromomaleimides as extremely efficient disulfide bridging bioconjugation and pegylation agents. , 2012, Journal of the American Chemical Society.

[96]  Tom P. Carberry,et al.  "Bio"-macromolecules: polymer-protein conjugates as emerging scaffolds for therapeutics. , 2014, Macromolecular rapid communications.

[97]  T. Hey,et al.  Half‐Life Extension through HESylation® , 2012 .

[98]  Ray Yin,et al.  A double antigen bridging immunogenicity ELISA for the detection of antibodies to polyethylene glycol polymers. , 2011, Journal of pharmacological and toxicological methods.

[99]  C. Barner‐Kowollik,et al.  Direct Synthesis of Pyridyl Disulfide-Terminated Polymers by RAFT Polymerization , 2007 .

[100]  Richard Hoogenboom,et al.  Poly(2-oxazoline)s: a polymer class with numerous potential applications. , 2009, Angewandte Chemie.

[101]  Hiroshi Maeda,et al.  A lipophilic derivative of neocarzinostatin. A polymer conjugation of an antitumor protein antibiotic. , 2009, International journal of peptide and protein research.

[102]  Lei Tao,et al.  Protein release from biodegradable polyHPMA-lysozyme conjugates resulting in bioactivity enhancement. , 2011, Chemistry, an Asian journal.

[103]  E. Harth,et al.  Controlled branching of polyglycidol and formation of protein-glycidol bioconjugates via a graft-from approach with "PEG-like" arms. , 2013, Chemical communications.

[104]  K. Ulbrich,et al.  Conjugates of Semitelechelic Poly[N-(2-Hydroxypropyl)Methacrylamide] with Enzymes for Protein Delivery , 1999 .

[105]  Eric D. Pressly,et al.  Poly[(ethylene oxide)-co-(methylene ethylene oxide)]: A hydrolytically-degradable poly(ethylene oxide) platform. , 2012, ACS macro letters.

[106]  H. Frey,et al.  Squaric acid mediated synthesis and biological activity of a library of linear and hyperbranched poly(glycerol)-protein conjugates. , 2012, Biomacromolecules.

[107]  M. Sawamoto,et al.  Polymerization of Methyl Methacrylate with the Carbon Tetrachloride/Dichlorotris- (triphenylphosphine)ruthenium(II)/Methylaluminum Bis(2,6-di-tert-butylphenoxide) Initiating System: Possibility of Living Radical Polymerization , 1995 .

[108]  H. Maynard,et al.  Trehalose glycopolymers for stabilization of protein conjugates to environmental stressors. , 2012, Journal of the American Chemical Society.

[109]  A. Turberfield,et al.  Multistep DNA-templated reactions for the synthesis of functional sequence controlled oligomers. , 2010, Angewandte Chemie.

[110]  F. Davis,et al.  Effect of covalent attachment of polyethylene glycol on immunogenicity and circulating life of bovine liver catalase. , 1977, The Journal of biological chemistry.

[111]  C. Barner‐Kowollik,et al.  In situ formation of protein-polymer conjugates through reversible addition fragmentation chain transfer polymerization. , 2007, Angewandte Chemie.

[112]  F. Veronese,et al.  Preparation of characterization of poly(ethylene glycol) vinyl sulfone. , 1996, Bioconjugate chemistry.

[113]  E. Toone,et al.  In situ growth of a stoichiometric PEG-like conjugate at a protein's N-terminus with significantly improved pharmacokinetics , 2009, Proceedings of the National Academy of Sciences.