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PEGylation is a classic strategy to reduce denaturation and immunogenicity in therapeutic proteins, bioimaging, and drug delivery. However, this concept has not been applied to incorporate biogenic materials as functional components in optoelectronics. Herein, PEGylation is rationalized as an effective tool to stabilize fluorescent proteins in solid‐state photon manipulation down‐converters integrated into bio‐hybrid light‐emitting diodes. In short, the archetypal red‐emitting protein mCherry is nonspecifically PEGylated with methoxypolyethylene glycols (mPEG) chains of different lengths (mPEG‐350/‐750/‐2000). These derivatives hold the same photoluminescence figures‐of‐merit of mCherry, but an improved resistance against organic solvents, pH, temperature, and polymers (in solution/dry‐coatings) upon increasing the mPEG length. Indeed, the best‐performing mCherry‐mPEG‐2000 adduct leads to deep‐red devices with threefold enhanced color stability (>300 h) under harsh operation conditions (150 mW cm−2) over the prior art. Different device architectures along with spectroscopic, thermocycling, and electrochemical impedance spectroscopy studies of the prepared coatings aid in rationalizing that PEGylation successfully reduces i) nonreversible thermal denaturation, ii) aggregation in solid‐state, and iii) photo‐induced oxidation and H+‐transfer deactivation of the chromophore, since oxygen diffusivity and H+‐reorganization across the protein skeleton are slowed down by strong H+‐bonding/hydrophobic mCherry‐mPEG interactions. Hence, this simple supramolecular modification is of utmost relevance for future advances in protein‐based optoelectronics. [ABSTRACT FROM AUTHOR]

PEGylated PAMAM dendrimers (PEG‐PAMAM) are well‐characterized biomaterials with still unexplored applications as carriers of drugs acting via membrane receptors, such as octreotide. This work confirmed the safety and negligible internalization capacity of fourth‐generation 50%‐PEG‐PAMAM in HEK‐293 cells, to then assessed their supramolecular binding to octreotide through tryptophan quenching experiments and Gaussian‐accelerated molecular dynamics (GaMD) simulations. Tryptophan quenching showed that PEG‐PAMAM binds octreotide with a Kbind of 6 × 106 M−1 and a complex stoichiometry of 1:1.4, unlike native PAMAM. GaMD simulations revealed that octreotide binds at the outer PEG shell of PEG‐PAMAM, potentially hindering the drug from proteolytic degradation and enabling its release at a membrane level. Viability experiments on HeLa, PC‐12, and HEK‐293 cells incubated with increasing concentrations of octreotide in free drug solutions and equimolar mixtures with PEG‐PAMAM confirmed that the PEGylated dendrimer acts as an efficient supramolecular carrier for octreotide and enhances the antiproliferative effects of the drug. Our findings highlight a novel facet for PEG‐PAMAM dendrimers as macromolecular vehicles for peptide or non‐peptide drugs acting via membrane receptor sites. [ABSTRACT FROM AUTHOR]

This note aims to inspire through providing a personal view of the development and potential Drug Delivery Nanocarriers functionalized with polythyleneglycol (PEG). This polymer has been used extensively in Pharmaceutical Technology in a variety of compositions, including polyethylene oxide (PEO)-based surfactants. However, the concept of PEGylation, which started in the 70's, differs from the functionality of a surfactant, already discloses in the 50's. Here, we strictly adhere to the biological functionality of PEGylated nanocarriers intended to have a reduced interaction with proteins and, therefore, modify their biodistribution as well as facilitate their diffusion across mucus and other biological barriers. We analyze how this concept has evolved over the years and the benefit obtained so far in terms of marketed nanomedicines and provide the readers with a prospect view of the topic. [ABSTRACT FROM AUTHOR]

Recombinant human growth hormone (rhGH) and GH receptor antagonists (GHAs) are used clinically to treat a range of disorders associated with GH deficiency or hypersecretion, respectively. However, these biotherapeutics can be difficult and expensive to manufacture with multiple challenges from recombinant protein generation through to the development of long-acting formulations required to improve the circulating half-life of the drug. In this review, we summarize methodologies and approaches used for making and purifying recombinant GH and GHA proteins, and strategies to improve pharmacokinetic and pharmacodynamic properties, including PEGylation and fusion proteins. Therapeutics that are in clinical use or are currently under development are also discussed.

Liposomes being a promising colloidal system facilitates delivery of drugs with limited pharmacokinetic properties to achieve desirable clinical applications. However, development of a stable liposomal system is always challenging due to multiple complexities involved. Aqueous instability of liposomes and impact of various process and formulation parameters can lead to serious alteration of its therapeutic performance. In the proposed work, the authors aim to develop stable Ibrutinib-loaded liposomes using lyophilization and Quality-by-Design and assess their long-term stability. Ibrutinib-loaded liposomes were developed and optimized using Quality-by-Design technique and were further PEGylated and characterized for the same. Effect of cryoprotectants during lyophilization and other parameters are evaluated to obtain a robust formulation. The stability studies were conducted upto 6 months at various storage conditions to evaluate the effect of lyophilization. The impact of formulation, processing and lyophilization parameters on physicochemical properties of developed liposomal systems were evaluated and are critically discussed. Liquid dispersion exhibited a %degradation of 16–36% at 25 °C/60% RH which was reduced for less than 1% in lyophilized formulation for 6 months. Critical analysis and assessment of various parameters lead to identification of optimum conditions to manufacture this drug product and also opens way forward for further evaluation and translational possibilities. [ABSTRACT FROM AUTHOR]

Although the discovery of insulin 100 years ago revolutionized the treatment of diabetes, its therapeutic potential is compromised by its short half-life and narrow therapeutic index. Current long-acting insulin analogs, such as insulin-polymer conjugates, are mainly used to improve pharmacokinetics by reducing renal clearance. However, these conjugates are synthesized without sacrificing the bioactivity of insulin, thus retaining the narrow therapeutic index of native insulin, and exceeding the efficacious dose still leads to hypoglycemia. Here, we report a kind of di-PEGylated insulin that can simultaneously reduce renal clearance and receptor-mediated clearance. By impairing the binding affinity to the receptor and the activation of the receptor, di-PEGylated insulin not only further prolongs the half-life of insulin compared to classical mono-PEGylated insulin but most importantly, increases its maximum tolerated dose 10-fold. The target of long-term glycemic management in vivo has been achieved through improved pharmacokinetics and a high dose. This work represents an essential step towards long-acting insulin medication with superior safety in reducing hypoglycemic events.

Polyethylene glycol modification (PEGylation) is a widely used strategy to improve the physicochemical properties of various macromolecules, especially protein drugs. However, its application in enhancing the performance of enzymes for molecular biology remains underexplored. This study explored the PEGylation of Bst DNA polymerase, determining optimal modification reaction conditions. In comparison to the unmodified wild-type counterpart, the modified Bst DNA polymerase exhibited significantly improved activity, thermal stability, and inhibitor tolerance during loop-mediated isothermal amplification. When applied for the detection of Salmonella in crude samples, the modified enzyme demonstrated a notably accelerated reaction rate. Therefore, PEGylation emerges as a viable strategy for refining DNA polymerases, helping in the development of novel molecular diagnostic reagents. [ABSTRACT FROM AUTHOR]

Nanoparticle-based drug delivery systems have emerged as promising platforms for enhancing therapeutic efficacy while minimizing off-target effects. Among various strategies employed to optimize these systems, polyethylene glycol (PEG) modification, known as PEGylation—the covalent attachment of PEG to nanoparticles, has gained considerable attention for its ability to impart stealth properties to nanoparticles while also extending circulation time and improving biocompatibility. PEGylation extends to different drug delivery systems, in specific, nanoparticles for targeting cancer cells, where the concentration of drug in the cancer cells is improved by virtue of PEGylation. The primary challenge linked to PEGylation lies in its confirmation. Numerous research findings provide comprehensive insights into selecting PEG for various PEGylation methods. In this review, we have endeavored to consolidate the outcomes concerning the choice of PEG and diverse PEGylation techniques. [ABSTRACT FROM AUTHOR]

Small‐molecule ruthenium (Ru) complexes exhibit limitations in terms of nonspecific delivery, rapid metabolism, and low tumor accumulation. Their delivery can be improved through physical encapsulation into nanocarriers via hydrophobic forces, metallophilic interactions, or π–π stacking interactions. However, delivering Ru complexes for efficient therapy is substantially hindered by potential leakage of drugs, low drug‐loading capacity, or batch‐to‐batch variations. Moreover, current metalloprodrug‐based self‐delivery systems necessitate supramolecular interactions, which are unsuitable for Ru complexes because of their octahedral structures. Herein, two self‐assembled molecular Ru drugs, Ru‐3XOEG and Ru‐PEG, are reported. Ru‐3XOEG involves a three‐arm dendritic oligo(ethylene glycol) (OEG), while Ru‐PEG involves a linear poly(ethylene glycol) (PEG) chain. Furthermore, these drugs contain an anticancer Ru moiety and a planar pyrene moiety to render hydrophobic forces and π–π stacking supramolecular interactions. Ru‐3XOEG self‐assembles into large compound micelles. Ru‐PEG self‐assembles into vesicles. These Ru‐containing self‐delivered systems exhibit well‐defined structures, high Ru loading contents, and long circulation in blood without external nanocarriers. Red light irradiation induces the release of Ru‐H2O anticancer agents and the generation of 1O2 to inhibit tumor growth. The presented design of self‐assembled molecular Ru complexes opens avenues for the concept of self‐delivered metalloprodrugs. [ABSTRACT FROM AUTHOR]

Background: Polyoxyethylene hydrogenated castor oil (HCO ethoxylates) is a nonionic surfactant used as an excipient for ointments and injections in human and veterinary drugs. Several polyethylene glycol (PEG) derivatives can be obtained depending on the number of moles of ethylene oxide (EO). HCO ethoxylates have the potential to cause anaphylactoid reactions. There is little published information about these types of reactions in dogs. Objective: To determine the potential for HCO‐ethoxylate‐containing drugs to cause anaphylactoid reactions in dogs, employing intradermal testing (IDT) with various concentrations of HCO ethoxylates (HCO‐25, ‐40, ‐60 and ‐80). Animals: Four healthy male laboratory dogs. Materials and Methods: We performed IDT with drugs containing HCO ethoxylates and HCO ethoxylates alone to determine threshold concentrations. The IDT scores and threshold concentrations were compared. Analysis of skin biopsies from IDT sites was used to measure the percentage of degranulated mast cells. The effect of histamine at IDT sites was investigated by pre‐treatment with an antihistamine. Results: All HCO‐ethoxylate‐containing drugs caused a wheal‐and‐flare reaction. The threshold concentrations (0.001% and 0.00001%) of each HCO‐ethoxylate depended on the number of moles of EO (p < 0.05). Mast cell degranulation was enhanced by all HCO ethoxylates. The HCO‐60‐induced reaction was suppressed by an oral antihistamine. Conclusions and Clinical Relevance: The threshold concentration can serve as a consideration for developing safe new drug formulations and for clinical decision‐making around using drugs containing PEG derivatives. IDT is useful to predict the risk of adverse effects. Antihistamines could demonstrate a prophylactic effect. [ABSTRACT FROM AUTHOR]

In drug development, it is not uncommon that an active substance exhibits efficacy in vitro but lacks the ability to specifically reach its target in vivo. As a result, targeted drug delivery has become a primary focus in the pharmaceutical sciences. Since the approval of Doxil® in 1995, liposomes have emerged as a leading nanoparticle in targeted drug delivery. Their low immunogenicity, high versatility, and well-documented efficacy have led to their clinical use against a wide variety of diseases. That being said, every disease is accompanied by a unique set of physiological conditions, and each liposomal product must be formulated with this consideration. There are a multitude of different targeting techniques for liposomes that can be employed depending on the application. Passive techniques such as PEGylation or the enhanced permeation and retention effect can improve general pharmacokinetics, while active techniques such as conjugating targeting molecules to the liposome surface may bring even further specificity. This review aims to summarize the current strategies for targeted liposomes in the treatment of diseases. [ABSTRACT FROM AUTHOR]

Although the discovery of insulin 100 years ago revolutionized the treatment of diabetes, its therapeutic potential is compromised by its short half-life and narrow therapeutic index. Current long-acting insulin analogs, such as insulin-polymer conjugates, are mainly used to improve pharmacokinetics by reducing renal clearance. However, these conjugates are synthesized without sacrificing the bioactivity of insulin, thus retaining the narrow therapeutic index of native insulin, and exceeding the efficacious dose still leads to hypoglycemia. Here, we report a kind of di-PEGylated insulin that can simultaneously reduce renal clearance and receptor-mediated clearance. By impairing the binding affinity to the receptor and the activation of the receptor, di-PEGylated insulin not only further prolongs the half-life of insulin compared to classical mono-PEGylated insulin but most importantly, increases its maximum tolerated dose 10-fold. The target of long-term glycemic management in vivo has been achieved through improved pharmacokinetics and a high dose. This work represents an essential step towards long-acting insulin medication with superior safety in reducing hypoglycemic events. Distinct from conventional mono-PEGylated insulin, this work developed a kind of di-PEGylated insulin that could simultaneously reduce renal clearance and receptor-mediated clearance. [Display omitted] [ABSTRACT FROM AUTHOR]

Cobalt oxide nanoparticles (Co3O4-NPs) and cobalt sulfide nanoparticles (Co9S8-NPs) have shown extensive potential in electrochemical sensing, catalysis, specific drug targeting, and resonance imaging. However, Co9S8-NPs have been rarely explored for biomedical applications as compared to their oxide counterparts. Thus, in the current study, biocompatible PEGylated Co9S8-NPs and Co3O4-NPs are explored and compared for biological and photocatalytic properties. PEGylation of both the NPs is achieved using a simple chemical co-precipitation method followed by characterization using UV visible spectroscopy (UV-Vis), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), scanning electron microscopy (SEM), energy-dispersive X-ray analysis (EDX), and pH-responsive dispersion study. After thorough characterization, the NPs are evaluated and compared for various biological applications including antibacterial, antifungal, antileishmanial, antioxidant, and biocompatibility as well as photo-catalytic dye degradation studies. Both the NPs have shown excellent biological applications; however, Co9S8-NPs exhibit comparatively better antibacterial, antifungal, and antioxidant properties except antileishmanial potential where Co3O4-NPs show slight superiority. Furthermore, Co3O4-NPs indicate a higher degradation potential of methylene blue (MB) up to 46.93%. In comparison, Co9S8-NPs have a degradation ability of up to 42%, at 20 mg/ml within 3 h, which indicates considerable remediation potential of the NPs. Interestingly, both the NPs exhibit non-hemolytic behavior, thus demonstrating a compatible and bio-safe nature of both the NPs. [ABSTRACT FROM AUTHOR]

Herein, we have investigated and compared biocompatible calcium oxide nanoparticles (CaO-NPs) with different surface chemistries for biological properties. In the study, green synthesis is achieved using an aqueous extract of Potentilla bifurca while chemical synthesized polyethylene glycol (PEG) and cetyltrimethylammonium bromide (CTAB)-coated CaO-NPs are prepared using a facile coprecipitation approach. The nanoparticles are characterized using different techniques including, X-ray diffraction (XRD), Fourier-transformed infrared spectroscopy (FTIR), UV-visible spectroscopy (UV–Vis), scanning electron microscopy (SEM), and energy-dispersive X-ray analysis (EDX). Moreover, yield and pH-responsive dispersion studies are also carried out. After comprehensive characterization, the NPs are explored for potential biological properties such as antibacterial, antifungal, antileshmanial, antioxidant, and biocompatibility. Our study reveals that green synthesis results in CaO-NPs with high antioxidant and dispersion properties while chemical synthesis in particular CTAB-coated CaO-NPs exhibits remarkably high antibacterial, antifungal, and leishmancidal properties. For instance, the NPs resulted in 25 ± 1.4 mm and 16 ± 0.4 mm zone of inhibition (ZOI) against Bacillus subtilis and Aspergillus flavus,, respectively, while 81.10% and 79.34% inhibition against the promastigote and amastigote forms of the leishmania tropica. However, none of the NPs displayed hemolytic behavior, affirming the biocompatible nature of all the NPs. Our study thus concludes that surface characteristics play a vital role in defining the biological properties of CaO-NPs and the NPs could be tailored to harness the required biological properties. [ABSTRACT FROM AUTHOR]

Niosomes are arousing significant interest thanks to their low cost, high biocompatibility, and negligible toxicity. In this work, a supercritical CO2-assisted process was performed at 100 bar and 40 °C to produce niosomes at different Span 80/Tween 80 weight ratios. The formulation of cholesterol and 80:20 Span 80/Tween 80 was selected to encapsulate vancomycin, used as a model active compound, to perform a drug release rate comparison between PEGylated and non-PEGylated niosomes. In both cases, nanometric vesicles were obtained, i.e., 214 ± 59 nm and 254 ± 73 nm for non-PEGylated and PEGylated niosomes, respectively, that were characterized by a high drug encapsulation efficiency (95% for non-PEGylated and 98% for PEGylated niosomes). However, only PEGylated niosomes were able to prolong the vancomycin release time up to 20-fold with respect to untreated drug powder, resulting in a powerful strategy to control the drug release rate. [ABSTRACT FROM AUTHOR]

In this research work, we designed a smart biodegradable PEG-coated MnO2 nanoparticles conjugated with doxorubicin (PMnO2-Dox NPs) for dual chemo-photodynamic therapy and magnetic resonance imaging (MRI) application. This highly sensitive pH-responsive PMnO2-Dox NPs causes effective drug release under acidic pH environment. PMnO2-Dox NPs not only improves the efficacy of chemo-photodynamic therapy due to synergistic response as well as improved MRI imaging via boosting T1 MRI contrast. Manifold characterization techniques have been employed for materials investigation. Crystallography of PMnO2-Dox NPs were performed by using XRD analysis, which confirmed tetragonal crystal structure with an approximate crystallite size of 20–30 nm. Surface morphology was confirmed via SEM analysis, results indicated the spherical and asymmetric agglomerated nanocluster of PMnO2-Dox NPs. In in vitro bioassay, the anticancer activity of PMnO2-Dox NPs were tested against breast cancer (MCF-7) cell line using MTT test. Moreover, it can also inhibit the growth of primary and secondary cancers without light exposure. Results suggested that PMnO2-Dox NPs not only convenient for cancer treatment via combined chemo-photodynamic therapy but also address the way towards a comprehensive strategy for MRI application via bright contrast agent T1 after overcoming the problem of multidrug resistance (MDR) and synergistic response of therapeutic analysis.

Novel approaches are required to address the urgent need to develop lipid-based carriers of paclitaxel (PTX) and other hydrophobic drugs for cancer chemotherapy. Carriers based on cationic liposomes (CLs) with fluid (i.e., chain-melted) membranes (e.g., EndoTAG-1) have shown promise in preclinical and late-stage clinical studies. Recent work found that the addition of a cone-shaped poly(ethylene glycol)-lipid (PEG-lipid) to PTX-loaded CLs (CLsPTX) promotes a transition to sterically stabilized, higher-curvature (smaller) nanoparticles consisting of a mixture of PEGylated CLsPTX and PTX-containing fluid lipid nanodiscs (nanodiscsPTX). These CLsPTX and nanodiscsPTX show significantly improved uptake and cytotoxicity in cultured human cancer cells at PEG coverage in the brush regime (10 mol % PEG-lipid). Here, we studied the PTX loading, in vivo circulation half-life, and biodistribution of systemically administered CLsPTX and nanodiscsPTX and assessed their ability to induce apoptosis in triple-negative breast-cancer-bearing immunocompetent mice. We focused on fluid rather than solid lipid nanodiscs because of the significantly higher solubility of PTX in fluid membranes. At 5 and 10 mol % of a PEG-lipid (PEG5K-lipid, molecular weight of PEG 5000 g/mol), the mixture of PEGylated CLsPTX and nanodiscsPTX was able to incorporate up to 2.5 mol % PTX without crystallization for at least 20 h. Remarkably, compared to preparations containing 2 and 5 mol % PEG5K-lipid (with the PEG chains in the mushroom regime), the particles at 10 mol % (with PEG chains in the brush regime) showed significantly higher blood half-life, tumor penetration, and proapoptotic activity. Our study suggests that increasing the PEG coverage of CL-based drug nanoformulations can improve their pharmacokinetics and therapeutic efficacy.

Fabry disease, a rare X-linked genetic disorder, results from pathogenic variants in GLA, leading to deficient lysosomal α-galactosidase A enzyme activity and multiorgan manifestations. Since 2001, enzyme replacement therapy (ERT), using agalsidase alfa or agalsidase beta, has been the mainstay treatment, albeit with limitations such as rapid clearance and immunogenicity. Pegunigalsidase alfa, a novel PEGylated recombinant alpha-galactosidase, offers promise as an alternative. Produced in plant cells, pegunigalsidase alfa exhibits enhanced stability, prolonged half-life, and reduced immunogenicity due to pegylation. A phase 1/2 clinical trial demonstrated Gb3 clearance from renal capillary endothelial cells and its 48-month extension study revealed notable outcomes in renal function preservation. Three phase 3 clinical trials (BRIDGE, BRIGHT, and BALANCE) have shown favorable efficacy and safety profile, although caution is warranted in interpreting the results of BRIDGE and BRIGHT which lacked control groups. In BALANCE, the pivotal phase 3 trial comparing pegunigalsidase alfa with agalsidase beta, an intention-to-treat analysis of the eGFR decline over 2 years showed that the intergroup difference [95%confidence interval] in the median slope was -0.36 mL/min/ 1.73 m²/year [-2.44; 1.73]. The confidence interval had a lower limit above the prespecified value of -3 mL/min/1.73 m2/year and included zero. Despite challenges such as occasional hypersensitivity reactions and immunecomplex- mediated glomerulonephritis, pegunigalsidase alfa approval by the European Medicines Agency and the Food and Drug Administration represents a significant addition to Fabry disease therapeutic landscape providing an option for patients in whom enzyme replacement therapy with current formulations is poorly tolerated or poorly effective. [ABSTRACT FROM AUTHOR]

Antagonist peptides (ANTs) of vasoactive intestinal polypeptide receptors (VIP-Rs) are shown to enhance T cell activation and proliferation in vitro, as well as improving T cell-dependent anti-tumor response in acute myeloid leukemia (AML) murine models. However, peptide therapeutics often suffer from poor metabolic stability and exhibit a short half-life/fast elimination in vivo. In this study, we describe efforts to enhance the drug properties of ANTs via chemical modifications. The lead antagonist (ANT308) is derivatized with the following modifications: N-terminus acetylation, peptide stapling, and PEGylation. Acetylated ANT308 exhibits diminished T cell activation in vitro, indicating that N-terminus conservation is critical for antagonist activity. The replacement of residues 13 and 17 with cysteine to accommodate a chemical staple results in diminished survival using the modified peptide to treat mice with AML. However, the incorporation of the constraint increases survival and reduces tumor burden relative to its unstapled counterpart. Notably, PEGylation has a significant positive effect, with fewer doses of PEGylated ANT308 needed to achieve comparable overall survival and tumor burden in leukemic mice dosed with the parenteral ANT308 peptide, suggesting that polyethylene glycol (PEG) incorporation enhances longevity, and thus the antagonist activity of ANT308. [ABSTRACT FROM AUTHOR]

Foot-and-mouth disease (FMD) is a vesicular disease of cloven-hoofed animals with devastating economic implications. The current FMD vaccine, routinely used in enzootic countries, requires at least 7 days to induce protection. However, FMD vaccination is typically not recommended for use in non-enzootic areas, underscoring the need to develop new fast-acting therapies for FMD control during outbreaks. Interferons (IFNs) are among the immune system's first line of defense against viral infections. Bovine type III IFN delivered by a replication defective adenovirus (Ad) vector has effectively blocked FMD in cattle. However, the limited duration of protection--usually only 1-3 days post-treatment (dpt)--diminishes its utility as a field therapeutic. Here, we test whether polyethylene glycosylation (PEGylation) of recombinant bovine IFNλ3 (PEGboIFNλ3) can extend the duration of IFN-induced prevention of FMDV infection in both vaccinated and unvaccinated cattle. We treated groups of heifers with PEGboIFNλ3 alone or in combination with an adenovirus-based FMD O1Manisa vaccine (Adt-O1M) at either 3 or 5 days prior to challenge with homologous wild type FMDV. We found that pre-treatment with PEGboIFNλ3 was highly effective at preventing clinical FMD when administered at either time point, with or without co-administration of Adt-O1M vaccine. PEGboIFNλ3 protein was detectable systemically for >10 days and antiviral activity for 4 days following administration. Furthermore, in combination with Adt-O1M vaccine, we observed a strong induction of FMDV-specific IFNγ+ T cell response, demonstrating its adjuvanticity when co-administered with a vaccine. Our results demonstrate the promise of this modified IFN as a pre-exposure prophylactic therapy for use in emergency outbreak scenarios. [ABSTRACT FROM AUTHOR]

Thermostability is an important and desired feature of therapeutic proteins and is critical for the success or failure of protein drugs development. It can be increased by PEGylation—binding of poly(ethylene glycol) moieties—or glycosylation—post-translational modification to add glycans. Here, the thermostability and thermodynamic parameters of native, PEGylated, and glycosylated versions of the antileukemic enzyme crisantaspase were investigated. First-order kinetics was found to describe the irreversible deactivation process. Activation energy of the enzyme-catalyzed reaction (E*) was estimated for native, PEGylated, and glycosylated enzyme (10.2, 14.8, and 18.8 kJ mol−1 respectively). Half-life decreased progressively with increasing temperature, and longer half-life was observed for PEG-crisantaspase (87.74 min) at 50 °C compared to the native form (9.79 min). The activation energy of denaturation of PEG-crisantaspase (307.1 kJ mol−1) was higher than for crisantaspase (218.1 kJ mol−1) and Glyco-crisantaspase (120.0 kJ mol−1), which means that more energy is required to overcome the energy barrier of the unfolding process. According to our results, PEG-crisantaspase is more thermostable than its native form, while Glyco-crisantaspase is more thermosensitive. [ABSTRACT FROM AUTHOR]

With the rapid advancement of genetic and protein engineering, proteins and peptides have emerged as promising drug molecules for therapeutic applications. Consequently, there has been a growing interest in the field of chemical modification technology to address challenges associated with their clinical use, including rapid clearance from circulation, immunogenicity, physical and chemical instabilities (such as aggregation, adsorption, deamination, clipping, oxidation, etc.), and enzymatic degradation. Polyethylene glycol (PEG) modification offers an effective solution to these issues due to its favorable properties. This review presents recent progress in the development and application of PEGylated therapeutic proteins and peptides (TPPs). For this purpose, firstly, the physical and chemical properties as well as classification of PEG and its derivatives are described. Subsequently, a detailed summary is provided on the main sites of PEGylated TPPs and the factors that influence their PEGylation. Furthermore, notable instances of PEG-modified TPPs (including antimicrobial peptides (AMPs), interferon, asparaginase and antibodies) are highlighted. Finally, we propose the chemical modification of TPPs with PEG, followed by an analysis of the current development status and future prospects of PEGylated TPPs. This work provides a comprehensive literature review in this promising field while facilitating researchers in utilizing PEG polymers to modify TPPs for disease treatment. [ABSTRACT FROM AUTHOR]

Poly(ethylene glycol) (PEG) modification, also known as PEGylation, has been extensively used to improve the stability of nanoparticles for nanomedical applications. However, PEG exhibits antigenicity in some formulations, motivating researchers to explore alternative polymers. Herein, poly(vinyl ether) (PVE) derivatives are highlighted as promising alternatives to PEG because they form intermediate water molecules that suppress non‐specific protein adsorption and platelet adhesion to the material surface. We prepared a water‐soluble PVE derivative, poly(2‐methoxyethyl vinyl ether) (PMOVE), and utilized it as a surface modifier for gold nanoparticles (AuNPs) as model nanoparticles. PMOVE with a thiol terminus was synthesized and confirmed to form an intermediate water molecule using differential scanning calorimetry. Similar to the synthesis of PEGylated AuNPs (PEG‐AuNPs), PMOVE‐modified AuNPs (PMOVE‐AuNPs) were successfully fabricated with an appreciably high density of PMOVE palisades via a thiol‐gold coordination reaction. Similar to PEG‐AuNPs, PMOVE‐AuNPs showed reduced serum protein adsorption and prolonged blood circulation. Additionally, no significant cytotoxicity was observed after incubation of a murine macrophage cell line, RAW264.7, with PMOVE‐AuNPs. Our results indicate that the PMOVE modification increases the stealthiness of nanoparticles that is equivalent to that achieved by PEGylation. [ABSTRACT FROM AUTHOR]

• PEGylation of peptide 3 improved its stability in human serum and antiviral activities against all four DENV serotypes. • The PEGylated peptide 3 (PEG-P3) demonstrated all three modes of action against all four DENV serotypes: cell protection, direct virus inactivation, and inhibition during post-infection. • PEG-P3 was able to retain its antiviral activities up to 72 h incubation in human serum. Dengue presents a major public health concern in over 100 countries due to the absence of an effective vaccine and antiviral therapy against all four dengue virus (DENV) serotypes. Several antiviral peptides were previously reported to inhibit at least three or all four DENV serotypes. Chemical modifications such as d -amino acid substitutions, polyethylene glycol (PEG)ylation, and cyclization could be applied to peptides to improve their biological activities and stability in serum. The PEGylated peptide 3 (PEG-P3) was identified to be the most promising antiviral candidate as it demonstrated good inhibitory effects against all four DENV serotypes during the pre- and post-infection stages, Based on the RP-HPLC and LC/MS analysis, peptide 4 was identified to be more stable in human serum than peptide 3, with 78.9 % and 41.6 % of the peptides remaining after 72 h of incubation in human serum, respectively. Both peptides were also able to retain their antiviral activities against specific DENV serotypes after 72 h incubation in human serum. PEG-P3 was found to be more stable than the unmodified peptide 3 with 89.4 % of PEG-P3 remaining in the human serum after 72 h of incubation. PEG-P3 was able to retain its inhibitory effects against DENV-1 to 4 after 72 h of incubation in human serum. This study provided insights into the antiviral activities and stabilities of the unmodified and chemically modified peptides in human serum. [ABSTRACT FROM AUTHOR]

Polyethylene glycol (PEG) is a versatile polymer that is used in numerous pharmaceutical applications like the food industry, a wide range of disinfectants, cosmetics, and many commonly used household products. PEGylation is the term used to describe the covalent attachment of PEG molecules to nanocarriers, proteins and peptides, and it is used to prolong the circulation half-life of the PEGylated products. Consequently, PEGylation improves the efficacy of PEGylated therapeutics. However, after four decades of research and more than two decades of clinical applications, an unappealing side of PEGylation has emerged. PEG immunogenicity and antigenicity are remarkable challenges that confound the widespread clinical application of PEGylated therapeutics — even those under clinical trials — as anti-PEG antibodies (Abs) are commonly reported following the systemic administration of PEGylated therapeutics. Furthermore, pre-existing anti-PEG Abs have also been reported in healthy individuals who have never been treated with PEGylated therapeutics. The circulating anti-PEG Abs, both treatment-induced and pre-existing, selectively bind to PEG molecules of the administered PEGylated therapeutics inducing activation of the complement system, which results in remarkable clinical implications with varying severity. These include increased blood clearance of the administered PEGylated therapeutics through what is known as the accelerated blood clearance (ABC) phenomenon and initiation of serious adverse effects through complement activation-related pseudoallergic reactions (CARPA). Therefore, the US FDA industry guidelines have recommended the screening of anti-PEG Abs, in addition to Abs against PEGylated proteins, in the clinical trials of PEGylated protein therapeutics. In addition, strategies revoking the immunogenic response against PEGylated therapeutics without compromising their therapeutic efficacy are important for the further development of advanced PEGylated therapeutics and drug-delivery systems. [Display omitted] [ABSTRACT FROM AUTHOR]

Background: Although its immunomodulatory properties make thymopentin (TP5) appealing, its rapid metabolism and inactivation in the digestive system pose significant challenges for global scientists. PEGylated niosomal nanocarriers are hypothesized to improve the physicochemical stability of TP5, and to enhance its intestinal permeability for oral administration. Methods: TP5-loaded PEGylated niosomes were fabricated using the thin film hydration method. Co-cultured Caco-2 and HT29 cells with different ratios were screened as in vitro intestinal models. The cytotoxicity of TP5 and its formulations were evaluated using an MTT assay. The cellular uptake and transport studies were investigated in the absence or presence of variable inhibitors or enhancers, and their mechanisms were explored. Results and Discussion: All TP5 solutions and their niosomal formulations were nontoxic to Caco-2 and HT-29 cells. The uptake of TP5-PEG-niosomes by cells relied on active endocytosis, exhibiting dependence on time, energy, and concentration, which has the potential to significantly enhance its cellular uptake compared to TP5 in solution. Nevertheless, cellular transport rates were similar between TP5 in solution and its niosomal groups. The cellular transport of TP5 in solution was carried out mainly through MRP5 endocytosis and a passive pathway and effluxed by MRP5 transporters, while that of TP5-niosomes and TP5-PEG-niosomes was carried out through adsorptive- and clathrin-mediated endocytosis requiring energy. The permeability and transport rate was further enhanced when EDTA and sodium taurocholate were used as the penetration enhancers. Conclusions: This research has illustrated that PEG-niosomes were able to enhance the cellular uptake and maintain the cellular transport of TP5. This study also shows this formulation's potential to serve as an effective carrier for improving the oral delivery of peptides. [ABSTRACT FROM AUTHOR]

This article explores a manufacturing strategy for producing consistent and scalable PEGylated bioconjugates, which are proteins linked to polyethylene glycol (PEG) for improved therapeutic efficacy and safety. The study compares the use of a continuous-flow reaction system to the traditional batch-mode approach and finds that continuous-flow offers advantages in terms of selectivity and conversion rates. The study also analyzes the differences in solvent-accessible surface area (SASA) between two proteins, lysozyme and IL-6, and suggests that this may contribute to variations in selectivity. The authors conclude that continuous-flow chemistry has potential for protein modifications but further research is needed for optimization. [Extracted from the article]

In the present study, four different types of fourth-generation pegylated citric acid based dendrimers namely PE/TMP-MC-PEG-CA-PEG and PE/TMP-MS-PEG-CA-PEG were synthesized. The dendrimers were further conjugated with cefotaxime (CFTX) sodium drug. The structures of these dendrimers and drug-dendrimer conjugates were confirmed by using 1H-NMR, 13C-NMR, and FTIR spectroscopy. The confirmation of particle size, surface topography and structure of drug-dendrimer conjugates was done using Zeta sizer, AFM, and HR-TEM techniques. Cytotoxicity investigation revealed that the CFTX-encapsulated dendrimers are safe against normal epithelial cells and showed 20–22% anticancer activity against A549 Lung carcinoma cells; besides that, the synthesized dendrimers and drug-dendrimer conjugates also showed 15–28% and 2–3% hemolytic toxicity, respectively, against red blood cells (RBC). The antibacterial activity of all synthesized third and fourth generation drug-dendrimer conjugates were investigated on Gram-positive and Gram-negative bacteria. The results show that drug conjugate with third generation dendrimers depicted a greater effective zone of inhibition (ZOI) as compared to fourth generation drug-dendrimer conjugates against these pathogenic bacteria. [ABSTRACT FROM AUTHOR]

Polyethylene glycol (PEG) conjugation (PEGylation) is a well-established strategy to improve the pharmacokinetic and biocompatibility properties of a wide variety of nanomedicines and therapeutic peptides and proteins. This broad use makes PEG an attractive 'allround' candidate marker for the biodistribution of such PEGylated compounds. This paper presents the development of a novel strategy for PEG quantification in biological matrices. The methodology is based on sample hydrolysis which both decomposes the sample matrix and degrades PEGylated analytes to specific molecular fragments more suitable for detection by LC–MS/MS. Method versatility was demonstrated by applying it to a wide variety of PEGylated compounds, including polymeric poly(ethylbutyl cyanoacrylate) (PEBCA) nanoparticles, lipidic nanoparticles (Doxil®, LipImage 815™ and lipid nanoparticles for nucleic acid delivery) and the antibody Cimzia®. Method applicability was assessed by analyzing plasma and tissue samples from a comprehensive drug biodistribution study in rats, of both PEBCA and LipImage 815™ nanoparticles. The results demonstrated the method's utility for biodistribution studies on PEG. Importantly, by using the method described herein in tandem with quantification of nanoparticle payloads, we showed that this approach can provide detailed understanding of various critical aspects of the in vivo behavior of PEGylated nanomedicines, such as drug release and particle stability. Together, the presented results demonstrate the novel method as a robust, versatile and generic approach for biodistribution analysis of PEGylated therapeutics. [Display omitted] • PEGylation is widely used to improve pharmacological properties of medicines. • Our method uses detection of PEG to describe biodistribution of nanomedicines. • We demonstrate that PEG detection improves understanding of biodistribution. • The analytical concept is generic and can apply to non-endogenous PEG alternatives. [ABSTRACT FROM AUTHOR]

Purpose: The study utilized non-ionic polymer macrogol to transform the surface properties of the DepoFoam drug carrier system, developing “surface-remodeled DepoFoam (SR-DFO)” following quality by design (QbD) principles. The primary objectives were to prolong drug delivery, reduce sudden releases, and enhance the overall quality and stability of DepoFoam. The research hypotheses are centered on the capability of macrogol-based surface modification to create an optimized drug delivery system with improved stability, extended drug release, and enhanced pharmacokinetic properties. Methods: In this research, surface remodeling was achieved through a series of processes, including high-shear homogenizer-assisted double emulsification, PEGylation, and purification. The resulting SR-DFO formulations were comprehensively characterized for critical quality attributes. Optimization was conducted using the Box-Behnken design, resulting in significant enhancements in both quality and stability compared to conventional liposomes and unmodified DepoFoam. Results: Comprehensive product characterization validates anticipated quality parameters: entrapment efficiency (86.16 ± 0.44%), drug-loading capacity (25.28 ± 0.07%), vesicle size (40.47 ± 0.1 µm), polydispersity index (PDI) of 0.051 ± 0.03, lipocrit of 90.67 ± 0.26%, and zeta potential of − 31.25 ± 3.25 mV. Remarkably, macrogol-based SR-DFO consistently sustains drug release above 90% for 168 h, devoid of sudden spikes, and maintains stability at 4 °C for 180 days. Mathematical models confirm drug release mechanisms’ validity. Moreover, this study emphasizes the critical influence of key materials like macrogol, phospholipids, triglycerides, and process variables on shaping product quality. Conclusion: These findings highlight the inventive promise of macrogol-coated DFO in transforming drug delivery, quality, and stability. This research, driven by a well-formed hypothesis, meticulous execution, and precise data analysis, opens new horizons in polymer-based DepoFoam systems. First author: Jebastin Koilpillai, M.Pharm. [ABSTRACT FROM AUTHOR]

The growing prevalence of hyperuricemia necessitates the urgent development of more potent treatments. This study aimed to develop, optimize, and evaluate the safety and efficacy of porcine–human recombinant uricase (PHRU) both in vitro and in vivo. The study employed gene editing of PHRU through site-directed mutagenesis, with recombinant proteins expressed in vitro utilizing Escherichia coli. The polyethylene glycol (PEG) approach was employed to augment uricase stability and diminish immunogenicity. The pharmacokinetics and pharmacodynamics of PHRU were tested in vitro and in Sprague Dawley rats. Successful expression of the fusion protein in E. coli and the development of the PEGylated drug were achieved. In vitro experiments confirmed the efficacy of PEG-PHRU in degrading uric acid, with PEGylation not markedly affecting the biological activity of PHRU. Animal studies revealed that PEG-PHRU significantly lowered plasma uric acid levels and mitigated hyperuricemia-induced renal damage in rats. Both drug metabolism and pharmacokinetics exhibited favorable characteristics without observable adverse effects in experimental animals. This novel fusion protein shows the potential for ameliorating hyperuricemia and related renal complications, highlighting it as a promising drug candidate with substantial market applications.

[Display omitted] We herein report that filgrastim product Neupogen® and the filgrastim formulation buffer induced aggregate formation when mixed in vitro with human plasma, and formation of large membranous erythrocyte aggregates when mixed with human blood, similar to the aggregation induced by pegfilgrastim and by pegfilgrastim buffer [T. Arvinte, E. Poirier, N. Ersayin, G. Darpin, A. Cudd, J. Dowd, S. Brokx, Aggregation of human plasma and of human blood induced in vitro by pegfilgrastim originator formulation buffer and pegfilgrastim products, Eur. J. Pharmaceut. Biopharmaceut. (2023), doi: 10.1016/j.ejpb.2023.10.019]. The data identify the filgrastim buffer (which is practically the same in filgrastim and pegfilgrastim products) as the main driver of human plasma and blood aggregation. Kinetic experiments showed differences in the extent of plasma aggregation induced by a filgrastim product manufactured in EU and one manufactured in USA. Human donor variability in the plasma aggregation induced by filgrastim was observed. To study the effect of PEGylation of the filgrastim protein on plasma aggregation we compared filgrastim (Neupogen®) with pegfilgrastim (Neulasta®) solutions at the same protein concentration. These data show that PEGylation has a beneficial effect in inhibiting to an extent plasma aggregation. Interestingly, 20 kDa polyethylene glycol in the filgrastim buffer induced more plasma aggregation compared to the buffer, similar to the aggregation induced by pegfilgrastim. For intravenous infusion filgrastim solutions (300 µg/ml, vials only) may be diluted in 5 % dextrose from a concentration of 300 µg/ml to 5 µg/ml. Aggregation of human plasma was also induced by filgrastim solutions diluted in 5 % dextrose to 50 µg/ml, 15 µg/ml and 5 µg/ml filgrastim, as well as by the filgrastim buffer similarly diluted in 5 % dextrose (1:6, 1:20 and 1:60 dilution). These data show that filgrastim solutions used for intravenous administration in patients induce human plasma aggregation in vitro. Such aggregation phenomena may be related to known infusion side effects of filgrastim therapy. [ABSTRACT FROM AUTHOR]

Polymeric nanoparticles (NPs) are widely used as drug delivery systems in nanomedicine. Despite their widespread application, a comprehensive understanding of their intracellular trafficking remains elusive. In the present study, we focused on exploring the impact of a 20 nm difference in size on NP performance, including drug delivery capabilities and intracellular trafficking. For that, poly(ethylene glycol) methyl ether-block-poly(lactide-co-glycolide) (PLGA-PEG) NPs with sizes of 50 and 70 nm were precisely tailored. To assess their prowess in encapsulating and releasing therapeutic agents, we have employed doxorubicin (Dox), a well-established anticancer drug widely utilized in clinical settings, as a model drug. Then, the beneficial effect of the developed nanoformulations was evaluated in breast cancer cells. Finally, we performed a semiquantitative analysis of both NPs' uptake and intracellular localization by immunostaining lysosomes, early endosomes, and recycling endosomes. The results show that the smaller NPs (50 nm) were able to reduce the metabolic activity of cancer cells more efficiently than NPs of 70 nm, in a time and concentration-dependent manner. These findings are corroborated by intracellular trafficking studies that reveal an earlier and higher uptake of NPs, with 50 nm compared to the 70 nm ones, by the breast cancer cells. Consequently, this study demonstrates that NP size, even in small increments, has an important impact on their therapeutic effect. [ABSTRACT FROM AUTHOR]

The covalent attachment of polyethylene glycol (PEG) to therapeutic agents, termed PEGylation, is a well‐established and clinically proven drug delivery approach to improve the pharmacokinetics and pharmacodynamics of drugs. Specifically, PEGylation can improve the parent drug's solubility, extend its circulation time, and reduce its immunogenicity, with minimal undesirable properties. PEGylation technology has been applied to various therapeutic modalities including small molecules, aptamers, peptides, and proteins, leading to over 30 PEGylated drugs currently used in the clinic and many investigational PEGylated agents under clinical trials. Here, we summarize the diverse types of PEGylation strategies, the key advantages of PEGylated therapeutics over their parent drugs, and the broad applications and impacts of PEGylation in clinical settings. A particular focus has been given to the size, topology, and functionalities of PEG molecules utilized in clinically used PEGylated drugs, as well as those under clinical trials. An additional section has been dedicated to analyzing some representative PEGylated drugs that were discontinued at different stages of clinical studies. Finally, we critically discuss the current challenges faced in the development and clinical translation of PEGylated agents. [ABSTRACT FROM AUTHOR]

[Display omitted] Nanoparticles (NPs) adsorb serum proteins when exposed to biological fluids, forming a dynamic protein corona that has a profound impact on their overall biological profile and fate. Polyethylene glycol (PEG) modification is the most widely used strategy to mitigate and inhibit protein corona formation. Nevertheless, the accurate mapping and quantification of PEG inhibition effects on protein corona formation have scarcely been reported. Herein, we demonstrate the direct observation and quantification of protein corona adsorbed onto PEGylated mesoporous silica particles by direct stochastic optical reconstruction microscopy (dSTORM). The variation tendency of protein penetration depth in terms of PEG molecular weights and incubated time is investigated for the first time. The maximum penetration depths present slight increase with the prolonged incubation time, while they tend to remarkably decrease with increased chain length of modified PEG. Moreover, the co-localization of preformed protein corona with lysosomes and the destination of adsorbed protein are demonstrated. Our method provides important technical characterization information and in-depth understanding of protein corona adsorbed onto PEGylated mesoporous silica particles. This shines new light on the behaviors of silica materials in cells and may promote their practical applications in biomedicine. [ABSTRACT FROM AUTHOR]

PEGylated chitosan nanoparticles as oral delivery system for protein drugs were constructed by polyelectrolyte composite method. BSA-loaded nanoparticles were prepared by optimizing the conditions using bovine serum albumin (BSA) as protein drug model. The average particle size was 205.74±1.96 nm with PDI of 0.090 and the zeta potential was –36.76±9.4 mV, when the molar ratio was 0.20, the concentration was 0.6 mg/mL and polyethylene glycol grafting degree was 16%. The system showed high loading capacity and encapsulation efficiency for BSA which was up to 22.8±5.8% and 82.3±6.2% respectively. The cumulative release rate decreased with increasing initial amount and BSA was released slower in strong acid environment, such as simulated gastric pH 1.4, with cumulative release rate of 45% after 180 h, indicating that the nanoparticles can protect BSA from degradation. The Korsmeyer-Peppas and first-order kinetic models were used to fit well, dominated by the diffusion of BSA adsorbed on the surface, the dissolution of nanoparticles and the diffusion of BSA from the nanoparticles. The results suggested that PEGylated chitosan nanoparticles show excellent biocompatibility, with the potential as promising nanocarriers for oral delivery of protein drugs. [ABSTRACT FROM AUTHOR]

The development of novel radiocontrast agents, mainly used for the visualization of blood vessels, is still an emerging task due to the variety of side effects of conventional X-ray contrast media. Recently, we have shown that octahedral chalcogenide rhenium clusters with phosphine ligands—Na2H14[{Re6Q8}(P(C2H4COO)3)6] (Q = S, Se)—can be considered as promising X-ray contrast agents if their relatively high toxicity related to the high charge of the complexes can be overcome. To address this issue, we propose one of the most widely used methods for tuning the properties of proteins and peptides—PEGylation (PEG is polyethylene glycol). The reaction between the clusters and PEG-400 was carried out in acidic aqueous media and resulted in the binding of up to five carboxylate groups with PEG. The study of cytotoxicity against Hep-2 cells and acute toxicity in mice showed a twofold reduction in toxicity after PEGylation, demonstrating the success of the strategy chosen. Finally, the compound obtained has been used for the visualization of blood vessels of laboratory rats by angiography and computed tomography. [ABSTRACT FROM AUTHOR]

A PEGylated Tween 80–functionalized chitosan–lipidic (PEG-T-Chito-Lip) nano-vesicular hybrid was developed for intranasal administration as an alternative delivery route to help improve the poor oral bioavailability of BCS class-III model/antiemetic (metoclopramide hydrochloride; MTC). The influence of varying levels of chitosan, cholesterol, PEG 600, and Tween 80 on the stability/release parameters of the formulated nanovesicles was optimized using Draper-Lin Design. Two optimized formulations (Opti-Max and Opti-Min) with both maximized and minimized MTC-release goals, were predicted, characterized, and proved their vesicular outline via light/electron microscopy, along with the mutual prompt/extended in-vitro release patterns. The dual-optimized MTC–loaded PEG-T-Chito-Lip nanovesicles were loaded in intranasal in-situ gel (ISG) and further underwent in-vivo pharmacokinetics/nose-to-brain delivery valuation on Sprague-Dawley rats. The absorption profiles in plasma (plasma-AUC0-∞) of the intranasal dual-optimized MTC–loaded nano-vesicular ISG formulation in pretreated rats were 2.95-fold and 1.64-fold more than rats pretreated with orally administered MTC and intranasally administered raw MTC-loaded ISG formulation, respectively. Interestingly, the brain-AUC0-∞ of the intranasal dual-optimized MTC–loaded ISG was 10 and 3 times more than brain-AUC0-∞ of the MTC-oral tablet and the intranasal raw MTC-loaded ISG, respectively. It was also revealed that the intranasal dual-optimized ISG significantly had the lowest liver-AUC0-∞ (862.19 ng.g−1.h−1) versus the MTC-oral tablet (5732.17 ng.g−1.h−1) and the intranasal raw MTC-loaded ISG (1799.69 ng.g−1.h−1). The brain/blood ratio profile for the intranasal dual-optimized ISG was significantly enhanced over all other MTC formulations (P < 0.05). Moreover, the 198.55% drug targeting efficiency, 75.26% nose-to-brain direct transport percentage, and 4.06 drug targeting index of the dual-optimized formulation were significantly higher than those of the raw MTC-loaded ISG formulation. The performance of the dual-optimized PEG-T-Chito-Lip nano-vesicular hybrids for intranasal administration evidenced MTC-improved bioavailability, circumvented hepatic metabolism, and enhanced brain targetability, with increased potentiality in heightening the convenience and compliance for patients. [ABSTRACT FROM AUTHOR]

In this study, eight nonconserved residues with exposed surfaces and flexible conformations of the homotetrameric PGUS (β‐glucuronidase from Aspergillus oryzae Li‐3) were identified. Single‐point mutation into cysteine enabled the thiol‐maleimide reaction and site‐specific protein assembly using a two‐arm polyethylene glycol (PEG)‐maleimide crosslinker (Mal2). The Mal2(1k) (with 1 kDa PEG spacer)‐crosslinked PGUS assemblies showed low crosslinking efficiency and unimproved thermostability except for G194C‐Mal2(1k). To improve the crosslinking efficiency, a lengthened crosslinker Mal2(2k) (with 2 kDa PEG spacer) was used to produce PGUS assembly and a highly improved thermostability was achieved with a half‐life of 47.2–169.2 min at 70°C, which is 1.04–3.74 times that of wild type PGUS. It is found that the thermostability of PGUS assembly was closely associated with the formation of inter‐tetramer assembly and intratetramer crosslinking, rather than the PEGylation of the enzyme. Therefore, the four‐arm PEG‐maleimide crosslinker Mal4(2k) (with 2 kDa PEG spacer) was employed to simultaneously increase the inter‐tetramer assembly and intratetramer crosslinking, and the resulting PGUS assemblies showed further improved thermostabilities compared with Mal2(2k)‐crosslinked assemblies. Finally, the application of PGUS assemblies with significantly improved thermostability to the bioconversion of GL proved that the PGUS assembly is a strong catalyst for glycyrrhizin (GL) hydrolysis in industrial applications. [ABSTRACT FROM AUTHOR]

[Display omitted] Tissue-type plasminogen activator (tPA) is the gold standard for emergency treatment of ischemic stroke, which is the third leading cause of death worldwide. Major challenges of tPA therapy are its rapid elimination by plasminogen activator inhibitor-1 (PAI-1) and hepatic clearance, leading to the use of high doses and consequent serious side effects, including internal bleeding, swelling and low blood pressure. In this regard, we developed three polyethylene glycol (PEG)ylated tPA bioconjugates based on the recombinant human tPA drug Alteplase using site-specific conjugation strategies. The first bioconjugate with PEGylation at the N-terminus of tPA performed by reductive alkylation showed a reduced proteolytic activity of 68 % compared to wild type tPA. PEGylation at the single-free cysteine of tPA with linear and branched PEG revealed similar proteolytic activities as the wild-type protein. Moreover, both bioconjugates with PEG-cysteine-modification showed 2-fold slower inhibition kinetics by PAI-1. All bioconjugates increased in hydrodynamic size as a critical requirement for half-life extension. [ABSTRACT FROM AUTHOR]

Chronic wounds have become a major concern for healthcare systems, as they have been related to diabetic foot ulcers, venous leg ulcers and pressure ulcers. Oleuropein is an active compound that is extracted from olive leaves and it has the ability to reduce injury to tissues owing to its antioxidant effect, hence improving wound healing. The poor pharmacokinetics of oleuropein have limited its use clinically. This work is aimed toward studying the impact of PEGylated and non‐PEGylated nanoliposomes loaded with oleuropein, as a carrier model, on wound‐healing activity. The thin film hydration method was used to compose PEGylated and non‐PEGylated liposomes, both loaded with oleuropein. The results indicated that each free, PEGylated and non‐PEGylated composition was within the limit of optimum nanoliposome characterization. The results showed that non‐PEGylated compositions produced higher efficiency in encapsulation (47.09 ± 10.06%) than the PEGylated ones (20.97 ± 10.52%). The PEG‐nanoliposomes loaded with oleuropein (PEG‐oleu) had mean size, charge and polydispersity index of 129.35 nm, −9.55 mV and 0.1010, respectively. The scratch assay results proved that PEGylated liposomal compositions have a more rapid wound‐healing activity than non‐PEGylated ones at different time intervals at 0, 2, 24 and 28 h. [ABSTRACT FROM AUTHOR]

Fabry disease, a rare X-linked genetic disorder, results from pathogenic variants in GLA, leading to deficient lysosomal α-galactosidase A enzyme activity and multi-organ manifestations. Since 2001, enzyme replacement therapy (ERT), using agalsidase alfa or agalsidase beta, has been the mainstay treatment, albeit with limitations such as rapid clearance and immunogenicity. Pegunigalsidase alfa, a novel PEGylated recombinant alpha-galactosidase, offers promise as an alternative. Produced in plant cells, pegunigalsidase alfa exhibits enhanced stability, prolonged half-life, and reduced immunogenicity due to pegylation. A phase 1/2 clinical trial demonstrated Gb3 clearance from renal capillary endothelial cells and its 48-month extension study revealed notable outcomes in renal function preservation. Three phase 3 clinical trials (BRIDGE, BRIGHT, and BALANCE) have shown favorable efficacy and safety profile, although caution is warranted in interpreting the results of BRIDGE and BRIGHT which lacked control groups. In BALANCE, the pivotal phase 3 trial comparing pegunigalsidase alfa with agalsidase beta, an intention-to-treat analysis of the eGFR decline over 2 years showed that the intergroup difference [95%confidence interval] in the median slope was −0.36 mL/min/1.73 m2/year [−2.44; 1.73]. The confidence interval had a lower limit above the prespecified value of −3 mL/min/1.73 m2/year and included zero. Despite challenges such as occasional hypersensitivity reactions and immune-complex-mediated glomerulonephritis, pegunigalsidase alfa approval by the European Medicines Agency and the Food and Drug Administration represents a significant addition to Fabry disease therapeutic landscape providing an option for patients in whom enzyme replacement therapy with current formulations is poorly tolerated or poorly effective.

Oligomerization of antibody fragments via modification with polyethylene glycol (pegylation) may alter their function and properties, leading to a multivalent interaction of the resulting constructs with the target antigen. In a recent study, we generated pegylated monomers and multimers of scFv fragments of GD2-specific antibodies using maleimide–thiol chemistry. Multimerization enhanced the antigen-binding properties and demonstrated a more efficient tumor uptake in a syngeneic GD2-positive mouse cancer model compared to monomeric antibody fragments, thereby providing a rationale for improving the therapeutic characteristics of GD2-specific antibody fragments. In this work, we obtained pegylated conjugates of scFv fragments of GD2-specific antibodies with maytansinoids DM1 or DM4 using tetravalent PEG-maleimide (PEG4). The protein products from the two-stage thiol–maleimide reaction resolved by gel electrophoresis indicated that pegylated scFv fragments constituted the predominant part of the protein bands, and most of the scFv formed pegylated monomers and dimers. The conjugates retained the ability to bind ganglioside GD2 comparable to that of the parental scFv fragment and to specifically interact with GD2-positive cells. Both induced significant inhibitory effects in the GD2-positive B78-D14 cell line, in contrast to the GD2-negative B16 cell line. The decrease in the B78-D14 cell viability when treated with scFv-PEG4-DM4 was more prominent than that for scFv-PEG4-DM1, and was characterized by a twofold lower half-maximal inhibitory concentration (IC50). Unlike the parental scFv fragment, the product of scFv and PEG4 conjugation (scFv–PEG4), consisting predominantly of pegylated scFv multimers and monomers, induced direct cell death in the GD2-positive B78-D14 cells. However, the potency of scFv–PEG4 was low in the selected concentration range, thus demonstrating that the cytotoxic effect of DM1 and DM4 within the antibody fragment–drug conjugates was primary. The suggested approach may contribute to development of novel configurations of antibody fragment–drug conjugates for cancer treatment.

Nanoparticle delivery of drugs to the brain is hindered by the blood-brain barrier (BBB). In malignant glioma (MG), small disruptions in the BBB may allow nanoparticles smaller than 20 nm to penetrate the dysfunctional barrier. We previously developed ultra-small nanoparticles called hyper-cell permeable micelles (HCPMis) with a radius of ∼12 nm and found that a PEGylated HCPMi system showed enhanced cell permeability and cellular uptake, and remarkable anti-tumor properties in MG treatment. However, no study had examined the delivery of temozolomide (TMZ), the first-line drug for MG, with the HCPMi platform. Herein, we use a simple PEGylation increment system (30 wt % PEG, 40 wt % PEG and 50 wt % PEG) to develop a robust optimized HCPMi nanoplatform for TMZ delivery. All optimized HCPMi systems showed greater stability than the non-PEGylated parent formulation. Compared with commercially available micelles (DSPE-PEG2000), all optimized HCPMi systems showed greater cellular uptake in vitro. Although a higher percentage of PEGylation was associated with better cellular uptake and anti-cancer properties, the difference was statistically insignificant. Furthermore, in vitro cytotoxicity assays revealed that all optimized HCPMi-encapsulated TMZ formulations showed significantly stronger anti-cancer properties than the parent drug TMZ and TMZ encapsulated DSPE-PEG2000, thus indicating the feasibility of using this nanoplatform for the delivery of TMZ to treat brain malignancies.

Abstract Background Implant infections caused by biofilm forming bacteria are a major threat in orthopedic surgery. Delivering antibiotics directly to an implant affected by a bacterial biofilm via superparamagnetic nanoporous silica nanoparticles could present a promising approach. Nevertheless, short blood circulation half-life because of rapid interactions of nanoparticles with the host’s immune system hinder them from being clinically used. The aim of this study was to determine the temporal in vivo resolution of magnetic nanoporous silica nanoparticle (MNPSNP) distribution and the effect of PEGylation and clodronate application using PET/CT imaging and gamma counting in an implant mouse model. Methods PEGylated and non-PEGylated MNPSNPs were radiolabeled with gallium-68 (68Ga), implementing the chelator tris(hydroxypyridinone). 36 mice were included in the study, 24 mice received a magnetic implant subcutaneously on the left and a titanium implant on the right hind leg. MNPSNP pharmacokinetics and implant accumulation was analyzed in dependence on PEGylation and additional clodronate application. Subsequently gamma counting was performed for further final analysis. Results The pharmacokinetics and biodistribution of all radiolabeled nanoparticles could clearly be visualized and followed by dynamic PET/CT imaging. Both variants of 68Ga-labeled MNPSNP accumulated mainly in liver and spleen. PEGylation of the nanoparticles already resulted in lower liver uptakes. Combination with macrophage depletion led to a highly significant effect whereas macrophage depletion alone could not reveal significant differences. Although MNPSNP accumulation around implants was low in comparison to the inner organs in PET/CT imaging, gamma counting displayed a significantly higher %I.D./g for the tissue surrounding the magnetic implants compared to the titanium control. Additional PEGylation and/or macrophage depletion revealed no significant differences regarding nanoparticle accumulation at the implantation site. Conclusion Tracking of 68Ga-labeled nanoparticles in a mouse model in the first critical hours post-injection by PET/CT imaging provided a better understanding of MNPSNP distribution, elimination and accumulation. Although PEGylation increases circulation time, nanoparticle accumulation at the implantation site was still insufficient for infection treatment and additional efforts are needed to increase local accumulation.