Polyethylene Glycol (PEG) is a general term for ethylene glycol polymers with relative molecular weights of 200-8000 and above, which have different physicochemical properties depending on their molecular weights and structures. It has excellent lubricant, moisturizing, dispersing and adhesive properties, and can be used as antistatic agent and softener, etc. It is widely used in cosmetics, pharmaceutical, chemical fiber, rubber, plastic, paper, paint, electroplating, pesticide, metal processing and food processing industries.

What Are PEG Linkers?

Through a chemical synthesis reaction, PEG linkers can be made by precisely introducing functionalized groups with strong reactivity at specific molecular ends of PEG.  Since a variety of groups can be introduced, PEG linkers are widely used in biological and medical research due to their solubility in water, excellent biocompatibility, and non-immunogenicity. 

There are two classes of PEG linkers, monodispersed and polydispersed PEG linkers. The molecular weight of polydisperse PEG is uncertain and distributed in a certain range. Often polydispersity index (PDI) is used to determine how narrow the molecular weight distribution is. On the country, Monodisperse PEG refers to polyethylene glycol (PEG) with a precise, discrete molecular weight (ie, with a fixed molecular weight rather than a range).

Application of PEG Linkers

A long-acting drug is a traditional application of PEGylation, which has been clinically applied for more than 30 years and gradually become the mainstream solution for long-acting drugs. As the research of PEGylation progresses, its application fields are widening, besides peptide and protein drugs, it is also applied to small molecule drugs, ADC drug linker, gene-drug (siRNA, mRNA) delivery and hydrogel-based devices, etc.

1. PEG in Long-Acting Drugs (Mainly Protein & Peptide Drugs)

In the 1970s, Professor Frank Davis of Rutgers University modified bovine serum albumin with PEG in order to reduce the immunogenicity of recombinant proteins, prolong their metabolism in vivo and enhance protein activity. Since then, this technology has been widely used in many fields of biomedicine. In 1981, Professor Davis founded Enzon Pharmaceuticals, and in 1990, the FDA approved Adagen® (pegademase bovine) , the world's first PEGylated drug for severe combined immunodeficiency disease. Since then, PEGylated ADA, IFN, CPT, rhG-CSF, L-asparagi-nase, and many other products have been introduced. To date, there are more than 30 FDA or EU-approved PEGylated drugs on the market worldwide, with a market size of more than $10 billion. Below, the tabel shows the FDA approved PEGylated drugs up to 2022.

The principle of long-acting PEGylated protein and peptide drugs is to reduce renal clearance and reduce the rate of enzymatic degradation. Protein and peptide drugs are composed of amino acids, which are often rapidly excreted through the kidneys or inactivated by proteolytic enzymes after their action in the body. As a result, their efficacy is short and can cause large changes in the blood concentration of drugs within a short period of time. In addition, these drugs may be recognized by the body's immune system as foreign substances in vitro, causing further adverse reactions.

Linking with specific PEG derivatives can increase the relative molecular weight of drugs, making them less likely to be degraded and filtered out of the body by the kidneys, and prolonging the duration of effective drug concentration maintained in the body. In addition, the long-chain PEG derivatives can wrap the drug to avoid rapid enzymatic degradation or recognition by the immune system, so that the drug can be released slowly in the body, stabilizing the blood concentration and reducing the frequency of drug administration. 

Advantages of PEGylated proetin & peptide, image source: biocompare.com

2. PEG in Small Molecule Drugs

PEGylation can address certain deficiencies in the physicochemical properties and pharmacokinetics of small molecule drugs.

First, the problem of water solubility of drugs can be solved. Many small molecules with drug activity are often insoluble in water and have high toxic side effects, making them difficult to be injected into human body. Since PEG derivatives have good water solubility, the drugs formed by coupling PEG with small molecules can also be dissolved in water quickly, so that they can be used as injections and absorbed by the human body.

Secondly, the half-life of the drug can be increased. Due to the increased relative molecular weight of PEGylated small molecule drug, it can avoid being rapidly filtered out of the body by the kidneys. As a result, the effective drug concentration can be maintained in the human body for a longer period of time with a single injection, increasing the half-life of the drug and reducing the frequency of drug administration for patients.

In 2014, AstraZeneca received FDA approval for its PEGylated derivative of naloxone (Movantik), making it the world's first approved PEGylated small molecule drug for use in adult patients with chronic non-cancer pain and opioid-induced constipation.

Movantik, shown above, is a pegylated derivative of α-naloxol.

In addition, PEGylation has a significant toxicity-reducing effect on antitumor small molecule drugs. The most studied PEGylated small molecules are relatively simple antitumor drugs, mainly paclitaxel and camptothecin, which have generally high toxicity but require relatively high doses to achieve therapeutic efficacy. 

3. PEG in Drug Delivery

PEGylated drug delivery platforms are the frontier application area of PEG in the pharmaceutical field. PEGylation technology can also be used in delivery systems such as siRNA, mRNA, and pDNA, and the future development of related technologies will significantly drive the development of PEGylated drug delivery platforms.

➢ LNP (Lipid Nanoparticle) is the most mainstream non-viral vector for the delivery of mRNA to target cells. It is composed of ionizable lipids that are positively charged at low pH and neutralize the negative charge of mRNA. In addition, LNP includes neutral lipids and cholesterol, which self-assemble into a core lipid structure with a surface layer that mimics the cell membrane. The LNP binds phospholipids conjugated to PEG to increase the hydrophilicity of the LNP surface and provide stability to the mRNA carrier.

The principle of RNA delivery by LNPs is currently not completely clarified. However, it is generally believed that the lipophilicity of LNPs allows the particles to bind to the cell membrane of the body and be taken up by endocytosis. Upon entry into the cell, the mRNA escapes from the initially encapsulated vesicles and is released into the cytoplasm to express the target protein. LNPs can also be expelled from the cell through reverse exocytosis, which is of particular concern for mRNA delivery via LNPs. The target organ of the mRNA-LNPs complex is the liver. After entering the body, the agent binds to apolipoprotein E, and is then taken up by liver cells through receptor-mediated endocytosis, where it acts.

On the other hand, LNPs not only deliver mRNA to human cells, but also act as immune adjuvants to enhance the immune response. These microscopic oil-like droplets, approximately 0.1 μm in diameter, surround and protect the fragile mRNA during vaccine production, transport and final injection into the body. According to the public data, PEG has never been used in any other vaccine worldwide before, and the use of PEG in mRNA vaccines last year was a new application of PEG in the pharmaceutical field.

Ingredients of BioNtech and Moderna mRNA Vaccines, image source: Elsevier

➢ LNP can also be used for siRNA drug delivery. siRNA faces a series of barriers including rapid renal clearance, internalization by phagocytic cells, aggregation with serum proteins, and easy enzymatic degradation by endogenous nucleases. PEGylated polymer nanoparticles can increase the efficiency of gene penetration, increase the concentration of gene drugs in cells, increase gene binding ability, and improve endosomal escape efficiency, which can effectively deliver siRNA into target cells and promote endosomal escape. Onpattro, the first RNAi-based therapeutic to receive FDA approval, is enabled by a lipid nanoparticle (LNP) system that facilitates siRNA delivery into the cytoplasm of target cells (hepatocytes). 

PEGylation for RNA Drug Delivery System, image source: Biomaterials

Biopharma PEG is a dynamic science company dedicated to PEG derivatives. We can supply the PEG products used in COVID-19 mRNA vaccines, such as mPEG-N,N-Ditetradecylacetamide, ALC-0159, mPEG2000-DMG, mPEG2000-DSPE, etc.

4. PEG Linker in Antibody Drug Conjugate (PEG8/12)

ADC (antibody-drug conjugate) drugs are composed of monoclonal antibodies, linkers and active drugs. A suitable linker helps maintain the stability between the antibody and the drug and helps the antibody to selectively deliver the drug to tumor cells and accurately release the drug. PEG is one of the most widely used linkers in targeted therapy. PEG linker has the characteristics of high utilization rate, high targeting ability, and pH regulating properties, etc. Through the selection of multiple functional groups, PEG linkers can be combined with different antibodies and drugs to form different linkers, such as pH-sensitive linkers, disulfide bond linkers, β-glucuronide linkers, etc. PEG binds to a drug target through a chemical linker to achieve specific site-specific linkage, degradation and sustained release. 

Among the approved ADCs, two of them, Trodelvy and Zynlonta, were developed with PEG moiety as part of their linker technology to improve solubility and stability in vivo. 

PEG linker for Zynlonta

PEG linker for Trodelvy

5. PEG Linker in PROTACs

Proteolysis targeting chimera (PROTAC) is a technique developed based on the ubiquitin-protein enzymatic system to induce target protein degradation by small molecule compounds. PROTAC consists of three components: a ligand that binds to an E3 ligase to guide protein degradation, a ligand that binds to a target protein to guide the targeting of small molecules, and a linker that is responsible for the binding of the two ligands.

Each unit of PROTAC affects its protein degradation properties, and in particular, the linker topology plays a critical role in both biological and physicochemical properties. The linker topology affecting the properties of PROTAC is based on three main points: length, rigidity and binding site.

(1) The length of the linker needs to be optimized. If the linker is too short, spatial conflict will occur, which will disrupt the ternary complex and reduce the degradation ability of PROTAC. If the linker is too long, it will increase the relative motility of the two ends of PROTAC and decrease the binding constant of E3 ligase to the target protein, thus reducing the stability of the ternary complex. In addition, the molecular weight will be increased and its cellular permeability may be reduced.

(2) Second, increasing the rigidity of the linker is an effective way to improve the pharmacokinetic properties and oral bioavailability of PROTAC. Appropriate rigidity of the linker can also limit the bioactive conformation of PROTAC, which may improve the degradation activity of the target protein.

(3) Finally, differences in the attachment sites of the target protein ligands and E3 ligase ligands had significant effects on the binary binding affinity, ternary complex conformation, and physicochemical and pharmacokinetic properties of PROTAC.

The article Current strategies for the design of PROTAC linkers: a critical review, published by Maple et al. on October 30, 2020, found that by far the most common motifs in PROTAC linker structures are PEG and alkyl chains of different lengths, accounting for approximately 54% and 31%, respectively.

6. PEG in Medical Devices

Multi-arm PEG derivatives is widely used as a medical device material for wound adhesion, hemostasis, anti-leakage and anti-adhesion in various human surgical procedures for its soluble, biocompatible, non-toxic and low immunogenic properties. At the same time, PEG can also be used as a raw material for implanted human medical devices, replacing the existing widely used plant-derived, animal-derived and human-derived materials. PEG is expected to become a new generation material in biodegradable medical devices after chitosan and sodium hyaluronate.

Tissue isolation for radiation therapy is a cutting-edge application area for PEG hydrogel products. When treating cancer patients with radiation therapy, the goal is to destroy cancer cells while avoiding damage to surrounding healthy tissue. Tissue isolation is achieved by injecting PEG hydrogels between tissues, which effectively reduces the radiation dose reaching other healthy tissues.

Marketed Polyethylene Glycol Medical Devices

Biopharma PEG is A Leading PEG Supplier

Biopharma PEG, a worldwide leader of PEG linkers,  offers a wide array of  5,000 PEG derivatives, covering monodispersed and polydispersed structures with molecular weights ranging from 200 to 4w. These compounds feature great aqueous solubility, a smart choice of PEG length, and a broad selection of functional groups to choose from. Contact us at sales@biochempeg.com for more details.

Article Original Source: https://us.huatengsci.com/news/show/1526.html

References:
[1] Lawrence, Billings, et al., "Conjugation Strategy Strongly Impacts the Conformational Stability of a PEG-Protein Conjugate", ACS chemical biology, 11, 17, (2016): 1805-1809, https://pubs.acs.org/doi/abs/10.1021/acschembio.6b00349
[2] Kalyanram, Puri, et al., "Understanding the Stealth Properties of PEGylated lipids: A Mini-Review", International Journal of Lipids, 1, 2, (2020): xxx-xxx,https://openaccesspub.org/article/1432/ijl-20-3457.pdf
[3] Tolcher, "Antibody drug conjugates: lessons from 20 years of clinical experience", Annals of Oncology, 27, 12, (2016): 2168-2172,https://pubmed.ncbi.nlm.nih.gov/27733376/
​[4] Buschmann, et. al., "Nanomaterial Delivery Systems for mRNA Vaccines", Vaccines, 9, 1, (2021): 65, https://doi.org/10.3390/vaccines9010065

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