Modification of biopharmaceutical molecules by covalent conjugation of polyethylene glycol (PEG) molecules is known to enhance pharmacologic and pharmaceutical properties of proteins and other large ...molecules and has been used successfully in 12 approved drugs. Both linear and branched-chain PEG reagents with molecular sizes of up to 40 kDa have been used with a variety of different PEG derivatives with different linker chemistries. This review describes the properties of PEG itself, the history and evolution of PEGylation chemistry, and provides examples of PEGylated drugs with an established medical history. A trend toward the use of complex PEG architectures and larger PEG polymers, but with very pure and well-characterized PEG reagents is described. Nonclinical toxicology findings related to PEG in approved PEGylated biopharmaceuticals are summarized. The effect attributed to the PEG part of the molecules as observed in 5 of the 12 marketed products was cellular vacuolation seen microscopically mainly in phagocytic cells which is likely related to their biological function to absorb and remove particles and macromolecules from blood and tissues. Experience with marketed PEGylated products indicates that adverse effects in toxicology studies are usually related to the active part of the drug but not to the PEG moiety.
Polyethylene glycol modification (PEGylation) can enhance the pharmacokinetic properties of therapeutic proteins by the attachment of polyethylene glycol (PEG) to the surface of a protein to shield ...the protein surface from proteolytic degradation and limit aggregation. However, current PEGylation strategies often reduce biological activity, potentially as a result of steric hindrance of PEG. Overall, there are no structure‐based guidelines for selection of conjugate sites that retain optimal biological activity with improved pharmacokinetic properties. In this study, site‐specific PEGylation based on the FGF2‐FGFR1‐heparin complex structure is performed. The effects of the conjugate sites on protein function are investigated by measuring the receptor/heparin binding affinities of the modified proteins and performing assays to measure cell‐based bio‐activity and in vivo stability. Comprehensive analysis of these data demonstrates that PEGylation of FGF2 that avoids the binding sites for fibroblast growth factor receptor 1 (FGFR1) and heparin provides optimal pharmacokinetic enhancement with minimal losses to biological activity. Animal experiments demonstrate that PEGylated FGF2 exhibits greater efficacy in protecting against traumatic brain injury‐induced brain damage and neurological functions than the non‐modified FGF2. This rational structure‐based PEGylation strategy for protein modification is expected to have a major impact in the area of protein‐based therapeutics.
Current polyethylene glycol modification (PEGylation) strategies often reduce biological activity of recombinant protein due to the steric hindrance of polyethylene glycol. Site‐specific PEGylation is performed based on the structure of FGF2‐FGFR1‐heparin complex. Through in‐depth analysis, a rational structure‐based guideline is established for the selection of optimal conjugate sites, which is expected to promote the development of protein‐based therapeutics.
Site-Specific PEGylation of Therapeutic Proteins Dozier, Jonathan K; Distefano, Mark D
International Journal of Molecular Sciences,
10/2015, Letnik:
16, Številka:
10
Journal Article, Book Review
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Odprti dostop
The use of proteins as therapeutics has a long history and is becoming ever more common in modern medicine. While the number of protein-based drugs is growing every year, significant problems still ...remain with their use. Among these problems are rapid degradation and excretion from patients, thus requiring frequent dosing, which in turn increases the chances for an immunological response as well as increasing the cost of therapy. One of the main strategies to alleviate these problems is to link a polyethylene glycol (PEG) group to the protein of interest. This process, called PEGylation, has grown dramatically in recent years resulting in several approved drugs. Installing a single PEG chain at a defined site in a protein is challenging. Recently, there is has been considerable research into various methods for the site-specific PEGylation of proteins. This review seeks to summarize that work and provide background and context for how site-specific PEGylation is performed. After introducing the topic of site-specific PEGylation, recent developments using chemical methods are described. That is followed by a more extensive discussion of bioorthogonal reactions and enzymatic labeling.
The paper discusses general problems in using PEG for conjugation to high or low molecular weight molecules. Methods of binding PEG to different functional groups in macromolecules is reported ...together with their eventual limitations. Problems encountered in conjugation, such as the evaluation of the number of PEG chains bound, the localisation of the site of conjugation in polypeptides and the procedure to direct PEGylation to the desired site in the molecule are discussed. Finally, the paper reports on more specific methods regarding reversible PEGylation, cross-linking reagents with PEG arms, PEG for enzyme solubilization in organic solvent and new polymers as alternative to PEG.
Proteins and peptides have played a pivotal role in revolutionizing disease treatment over the last century. Despite their commercial success, protein therapeutics can be eliminated or inactivated in ...the body via excretion or other metabolic pathways. Polymeric materials have been used to stabilize these biomolecules in the presence of external stressors as excipients, conjugates, and in nanomaterial formulations. Numerous advantages arise from the combination of therapeutic agents with polymeric carriers, including improved stability, solubility, prolonged blood circulation, and reduced immunogenicity. PEGylation, the covalent conjugation of poly(ethylene glycol) to a biomolecule of interest, is a common technique that has been employed in 31 FDA-approved therapeutic protein formulations to date. Although PEGylation has been widely adopted, there have been numerous advancements in the protein stabilization field using a variety of polymers including, but not limited to, poly(oxazolines), polypeptides, zwitterionic polymers, and polysaccharides with additional beneficial properties such as biocompatibility and biodegradability. Polymeric carriers can also protect lyophilized protein-peptide products from the stresses of supercooling, ice crystallization, sublimation, and desorption. This review discusses recent progress on the design principles of polymeric tools for biomolecule stabilization and delivery, with a focus on conjugates and nanomaterials. The clinical status of these materials and current challenges impeding the clinical translation are presented. In addition, various future possibilities for polymeric-protein therapies are also highlighted. Finally, the current computational landscape that harnesses the tools of machine learning combined with experimental validation to design polymeric systems tailored for biomolecule stability are discussed.
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Poly(ethylene glycol) (PEG) is widely used as a gold standard in bioconjugation and nanomedicine to prolong blood circulation time and improve drug efficacy. The conjugation of PEG to proteins, ...peptides, oligonucleotides (DNA, small interfering RNA (siRNA), microRNA (miRNA)) and nanoparticles is a well-established technique known as PEGylation, with PEGylated products have been using in clinics for the last few decades. However, it is increasingly recognized that treating patients with PEGylated drugs can lead to the formation of antibodies that specifically recognize and bind to PEG (i.e., anti-PEG antibodies). Anti-PEG antibodies are also found in patients who have never been treated with PEGylated drugs but have consumed products containing PEG. Consequently, treating patients who have acquired anti-PEG antibodies with PEGylated drugs results in accelerated blood clearance, low drug efficacy, hypersensitivity, and, in some cases, life-threatening side effects. In this succinct review, we collate recent literature to draw the attention of polymer chemists to the issue of PEG immunogenicity in drug delivery and bioconjugation, thereby highlighting the importance of developing alternative polymers to replace PEG. Several promising yet imperfect alternatives to PEG are also discussed. To achieve asatisfactory alternative, further joint efforts of polymer chemists and scientists in related fields are urgently needed to design, synthesize and evaluate new alternatives to PEG.
Targeted therapy approaches have become the core of modern translational science and as an intriguing field, it is the solution of the conventional drug delivery problems that were once unanswered. ...Traditional methods of delivering drugs and therapeutics faced issues of solubility, sustained release, not enough amount getting through the diseased site, for e.g a tumor. Various formulations of liposomes, polymers, dendrimers, etc have succeeded and made their way for clinical trials trying to enhance the pharmacokinetic and biodistribution of the drug. Many stealth coatings that include hydrophilic polymers (PEG, chitosan, polyacrylamides, etc) can act as a covering around the nanoparticle that can shield the surface from aggregation, opsonization and evade immune system, thus considered in Generally Recognized as Safe (GRAS) category. Several other polymers such as poly-2-oxazoline, polyethylene oxide, PEG-based surfactant (polysorbate-80), and zwitterionic phospholipids have also been tested for their antifouling properties. However, the polymer coating approach requires labor-intensive procedures and conjugation chemistries that often fail in mice model. Besides, due to immunogenicity and allergic reactions evoked by the PEG-coated nanoparticles, there was an urge to find biomimicking materials that can prove better as shielding agents which paved the way for cell membrane coated nanoparticles (CMCNPs) to come into the limelight. CMCNPs consist of a nanoparticle inner core covered by cell membrane that can be implicated in targeted drug delivery approaches, photothermal therapy, diagnosis or imaging making it a powerful theranostic tool. In this review, mode of preparation of CMCNPs, different sources of cell membranes (RBCs, WBCs, platelets, cancer cells, stem cells with some other unconventional sources) and nanoparticle cores that are employed have been thoroughly emphasized. In addition to this, advancements and limitations with respect to this newly emerging field have been focussed.
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Carbon nanostructures are considered distinctive multi-functional drug delivery platforms for cancer therapy. However, their potential biocompatibility is still a significant challenge and rational ...design of these carriers is a prospect in the context of efficient cancer therapy. Herein, we report on the synthesis of two bio-inspired carbon nanohorns (CNH) platforms with tumor targeting capabilities for the delivery of cisplatin. Oxidized CNH wrapped with either polyethylene glycol (PEG) and fusogenic lipids (PEG/DSPG CNH-CP) or glutamic acid-cisplatin complex (CNH-GA-CP), revealed nanosized dahlia-like structure approved by dynamic light scattering (DLS) and transmission electron microscopy (TEM) analysis. Further structure features analysis by using RAMAN, FTIR spectroscopy and TGA indicated CNH surface modification. Excitingly, the superior nanofeatures of the CNH platforms was due to the reduced hydrophobic interactions, and thus closely linked to the surface modifications. Both platforms preserved cisplatin toxicity in vitro on C26 cells. The PEGylated nanoplatform demonstrated elongated circulation time compared to the free drug, and showed a site-specific delivery to the tumor site, while reducing the accumulation within kidney. Following 3 i.v. doses at 3 mg/kg of cisplatin, PEGylated CNH significantly delayed tumor growth and improved the life span of C26-bearing BALB/c mice. These notable results were further approved by the histopathological data, demonstrating reduced degeneration, necrosis and inflammation in both kidney and liver tissues following PEGylated nanoplatform administration. In conclusion, our findings suggest that the systemic toxicity of cisplatin including nephrotoxicity can be resolved by delivering this chemotherapeutic through a rationally-fabricated surface-modified carbon nanoplatform to improve the cancer therapy outcome.
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•CNHs hydrophobic interactions reduce biocompatibility and bio-applications.•Surface modification with PEG and fusogenic lipids improved CNH nano-features.•PEGylated CNHs preserved cisplatin toxicity on C26 cells in vitro.•PEGylated CNHs showed long circulation time and reduced tumor mass in animals.•Kidney toxicity was reduced following treatment with PEGylated CNHs bearing cisplatin.
The search for novel anticancer agents to replace the current platinum-based treatments remains an ongoing process. Palladacycles have shown excellent promise as demonstrated by our previous work ...which yielded BTC2, a binuclear palladadycle with a non-ionisable polyethylene glycol (PEG) tether. Here, we explore the importance of the PEG-tether length on the anticancer activity of the binuclear palladacycles by comparing three analogous binuclear palladacycles, BTC2, BTC5 and BTC6, in the oestrogen receptor positive MCF7 and triple-negative MDA-MB-231 breast cancer cell lines. In addition, these are compared to another analogue with an ionisable morpholine tether, BTC7. Potent anticancer activity was revealed through cell viability studies (MTT assays) revealed that while BTC6 showed similar potent anticancer activity as BTC2, it was less toxic towards non-cancerous cell lines. Interestingly, BTC7 and BTCF were less potent than the PEGylated palladacycles but showed significantly improved selectivity towards the triple-negative breast cancer cells. Cell death analysis showed that BTC7 and BTCF significantly induced apoptosis in both the cancer cell lines while the PEGylated complexes induced both apoptosis and secondary necrosis. Furthermore, experimental and computational DNA binding studies indicated partial intercalation and groove binding as the modes of action for the PEGylated palladacycles. Similarly, experimental and computational BSA binding studies indicated and specific binding sites in BSA dependent on the nature of the tethers on the complexes.
The nature of the tether controls the activity, selectivity and biomolecular interactions of the palladacycle. Display omitted
•Developed a series of binuclear palladacycles with ionisable and non-ionisable tethers.•Potent anticancer activity observed via MTT assays.•BTC6 is an improvement on its precursor BTC2 – less toxic and more soluble.•BTC7 shows selectivity towards triple-negative breast cancer cell line MDA-MB-231•Experimental and computational DNA and BSA binding studies show that the nature of the tether determines the type and location of the interaction observed.
A layer of mucus covers the surface of all wet epithelia throughout the human body. Mucus is a hydrogel mainly composed of water, mucins (glycoproteins), DNA, proteins, lipids, and cell debris. This ...complex composition yields a tenacious viscoelastic hydrogel that lubricates and protects the exposed epithelia from external threats and enzymatic degradation. The natural protective role of mucus is nowadays acknowledged as a major barrier to be overcome in non-invasive drug delivery. The heterogeneity of mucus components offers a wide range of potential chemical interaction sites for macromolecules, while the mesh-like architecture given to mucus by the intermolecular cross-linking of mucin molecules results in a dense network that physically, and in a size-dependent manner, hinders the diffusion of nanoparticles through mucus. Consequently, drug diffusion, epithelial absorption, drug bioavailability, and ultimately therapeutic outcomes of mucosal drug delivery can be attenuated.
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