Water-responsive supercontractile polymer films for bioelectronic interfaces.

Connecting different electronic devices is usually straightforward because they have paired, standardized interfaces, in which the shapes and sizes match each other perfectly. Tissue-electronics interfaces, however, cannot be standardized, because tissues are soft1-3 and have arbitrary shapes and sizes4-6. Shape-adaptive wrapping and covering around irregularly sized and shaped objects have been achieved using heat-shrink films because they can contract largely and rapidly when heated7. However, these materials are unsuitable for biological applications because they are usually much harder than tissues and contract at temperatures higher than 90 °C (refs. 8,9). Therefore, it is challenging to prepare stimuli-responsive films with large and rapid contractions for which the stimuli and mechanical properties are compatible with vulnerable tissues and electronic integration processes. Here, inspired by spider silk10-12, we designed water-responsive supercontractile polymer films composed of poly(ethylene oxide) and poly(ethylene glycol)-α-cyclodextrin inclusion complex, which are initially dry, flexible and stable under ambient conditions, contract by more than 50% of their original length within seconds (about 30% per second) after wetting and become soft (about 100 kPa) and stretchable (around 600%) hydrogel thin films thereafter. This supercontraction is attributed to the aligned microporous hierarchical structures of the films, which also facilitate electronic integration. We used this film to fabricate shape-adaptive electrode arrays that simplify the implantation procedure through supercontraction and conformally wrap around nerves, muscles and hearts of different sizes when wetted for in vivo nerve stimulation and electrophysiological signal recording. This study demonstrates that this water-responsive material can play an important part in shaping the next-generation tissue-electronics interfaces as well as broadening the biomedical application of shape-adaptive materials.

Membrane nanotubes transform into double-membrane sheets at condensate droplets.

Cellular membranes exhibit a multitude of highly curved morphologies such as buds, nanotubes, cisterna-like sheets defining the outlines of organelles. Here, we mimic cell compartmentation using an aqueous two-phase system of dextran and poly(ethylene glycol) encapsulated in giant vesicles. Upon osmotic deflation, the vesicle membrane forms nanotubes, which undergo surprising morphological transformations at the liquid-liquid interfaces inside the vesicles. At these interfaces, the nanotubes transform into cisterna-like double-membrane sheets (DMS) connected to the mother vesicle via short membrane necks. Using super-resolution (stimulated emission depletion) microscopy and theoretical considerations, we construct a morphology diagram predicting the tube-to-sheet transformation, which is driven by a decrease in the free energy. Nanotube knots can prohibit the tube-to-sheet transformation by blocking water influx into the tubes. Because both nanotubes and DMSs are frequently formed by cellular membranes, understanding the formation and transformation between these membrane morphologies provides insight into the origin and evolution of cellular organelles.

Polysaccharide functionalization reduces lipid vesicle stiffness.

The biophysical properties of lipid vesicles are important for their stability and integrity, key parameters that control the performance when these vesicles are used for drug delivery. The vesicle properties are determined by the composition of lipids used to form the vesicle. However, for a given lipid composition, they can also be tailored by tethering polymers to the membrane. Typically, synthetic polymers like polyethyleneglycol are used to increase vesicle stability, but the use of polysaccharides in this context is much less explored. Here, we report a general method for functionalizing lipid vesicles with polysaccharides by binding them to cholesterol. We incorporate the polysaccharides on the outer membrane leaflet of giant unilamellar vesicles (GUVs) and investigate their effect on membrane mechanics using micropipette aspiration. We find that the presence of the glycolipid functionalization produces an unexpected softening of GUVs with fluid-like membranes. By contrast, the functionalization of GUVs with polyethylene glycol does not reduce their stretching modulus. This work provides the potential means to study membrane-bound meshworks of polysaccharides similar to the cellular glycocalyx; moreover, it can be used for tuning the mechanical properties of drug delivery vehicles.

Enhancing protein dynamics analysis with hydrophilic polyethylene glycol cross-linkers.

Cross-linkers play a critical role in capturing protein dynamics in chemical cross-linking mass spectrometry techniques. Various types of cross-linkers with different backbone features are widely used in the study of proteins. However, it is still not clear how the cross-linkers' backbone affect their own structure and their interactions with proteins. In this study, we systematically characterized and compared methylene backbone and polyethylene glycol (PEG) backbone cross-linkers in terms of capturing protein structure and dynamics. The results indicate the cross-linker with PEG backbone have a better ability to capture the inter-domain dynamics of calmodulin, adenylate kinase, maltodextrin binding protein and dual-specificity protein phosphatase. We further conducted quantum chemical calculations and all-atom molecular dynamics simulations to analyze thermodynamic and kinetic properties of PEG backbone and methylene backbone cross-linkers. Solution nuclear magnetic resonance was employed to validate the interaction interface between proteins and cross-linkers. Our findings suggest that the polarity distribution of PEG backbone enhances the accessibility of the cross-linker to the protein surface, facilitating the capture of sites located in dynamic regions. By comprehensively benchmarking with disuccinimidyl suberate (DSS)/bis-sulfosuccinimidyl-suberate(BS3), bis-succinimidyl-(PEG)2 revealed superior advantages in protein dynamic conformation analysis in vitro and in vivo, enabling the capture of a greater number of cross-linking sites and better modeling of protein dynamics. Furthermore, our study provides valuable guidance for the development and application of PEG backbone cross-linkers.

Consequences of poly(ethylene oxide) and poloxamer P188 on transcription in healthy and stressed myoblasts.

Poly(ethylene oxide) (PEO) and poloxamers, a class of poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) (PEO-PPO-PEO) triblock copolymers, have many personal and medical care applications, including the stabilization of stressed cellular membranes. Despite the widespread use, the cellular transcriptional response to these molecules is relatively unknown. C2C12 myoblasts, a model muscle cell, were subjected to short-term Poloxamer 188 (P188) and PEO181 (8,000 g/mol) treatment in culture. RNA was extracted and sequenced to quantify transcriptomic impact. The addition of moderate concentrations (14 µM) of either polymer to unstressed cells caused substantial differential gene expression, including at least twofold modulation of 357 and 588 genes, respectively. In addition, evaluation of the transcriptome response to osmotic stress without polymer treatment revealed dramatic change in RNA expression. Interestingly, the addition of polymer to stressed cells-at concentrations that provide physiological protection-did not yield a significant difference in expression of any gene relative to stress alone. Genome-scale expression analysis was corroborated by single-gene quantitative real-time PCR. Changes in protein expression were measured via western blot, which revealed partial alignment with the RNA results. Collectively, the significant changes to expression of multiple genes and resultant protein translation demonstrates an unexpectedly broad biochemical response to these polymers in healthy myoblasts in vitro. Meanwhile, the lack of substantial transcriptional response to polymer treatment in stressed cells highlights the physical nature of that protective mechanism.

Drinkable in situ-forming tough hydrogels for gastrointestinal therapeutics.

Pills are a cornerstone of medicine but can be challenging to swallow. While liquid formulations are easier to ingest, they lack the capacity to localize therapeutics with excipients nor act as controlled release devices. Here we describe drug formulations based on liquid in situ-forming tough (LIFT) hydrogels that bridge the advantages of solid and liquid dosage forms. LIFT hydrogels form directly in the stomach through sequential ingestion of a crosslinker solution of calcium and dithiol crosslinkers, followed by a drug-containing polymer solution of alginate and four-arm poly(ethylene glycol)-maleimide. We show that LIFT hydrogels robustly form in the stomachs of live rats and pigs, and are mechanically tough, biocompatible and safely cleared after 24 h. LIFT hydrogels deliver a total drug dose comparable to unencapsulated drug in a controlled manner, and protect encapsulated therapeutic enzymes and bacteria from gastric acid-mediated deactivation. Overall, LIFT hydrogels may expand access to advanced therapeutics for patients with difficulty swallowing.

Increased efficiency of charge-mediated fusion in polymer/lipid hybrid membranes.

Due to their augmented properties, biomimetic polymer/lipid hybrid compartments are a promising substitute for natural liposomes in multiple applications, but the protein-free fusion of those semisynthetic membranes is unexplored to date. Here, we study the charge-mediated fusion of hybrid vesicles composed of poly(dimethylsiloxane)-graft-poly(ethylene oxide) and different lipids and analyze the process by size distribution and the mixing of membrane species at µm and nano scales. Remarkably, the membrane mixing of oppositely charged hybrids surpasses by far the degree in liposomes, which we correlate with properties like membrane disorder, rigidity, and ability of amphiphiles for flip-flop. Furthermore, we employ the integration of two respiratory proteins as a functional content mixing assay for different membrane compositions. This reveals that fusion is also attainable with neutral and cationic hybrids and that the charge is not the sole determinant of the final adenosine triphosphate synthesis rate, substantiating the importance of reconstitution environment. Finally, we employ this fusion strategy for the delivery of membrane proteins to giant unilamellar vesicles as a way to automate the assembly of synthetic cells.

The influence of molecular design on structure-property relationships of a supramolecular polymer prodrug.

Supramolecular self-assemblies of hydrophilic macromolecules functionalized with hydrophobic, structure-directing components have long been used for drug delivery. In these systems, loading of poorly soluble compounds is typically achieved through physical encapsulation during or after formation of the supramolecular assembly, resulting in low encapsulation efficiencies and limited control over release kinetics, which are predominately governed by diffusion and carrier degradation. To overcome these limitations, amphiphilic prodrugs that leverage a hydrophobic drug as both the therapeutic and structure-directing component can be used to create supramolecular materials with higher loading and controlled-release kinetics using biodegradable or enzymatically cleavable linkers. Here, we report the design, synthesis, and characterization of a library of supramolecular polymer prodrugs based on poly(ethylene glycol) (PEG) and the proregenerative drug 1,4-dihydrophenonthrolin-4-one-3-carboxylic acid (DPCA). Structure-property relationships were elucidated through experimental characterization of prodrug behavior in both the wet and dry states using scattering techniques and electron microscopy and corroborated by coarse-grained modeling. Molecular architecture and the hydrophobic-to-hydrophilic ratio of PEG-DPCA conjugates strongly influenced their physical state in water, ranging from fully soluble to supramolecular spherical assemblies and nanofibers. Molecular design and supramolecular structure, in turn, were shown to dramatically alter hydrolytic and enzymatic release and cellular transport of DPCA. In addition to potentially expanding therapeutic options for DPCA through control of supramolecular assemblies, the design principles elaborated here may inform the development of other supramolecular prodrugs based on hydrophobic small-molecule compounds.

Gd2O3-mesoporous silica/gold nanoshells: A potential dual T1/T2 contrast agent for MRI-guided localized near-IR photothermal therapy.

A promising clinical trial utilizing gold-silica core-shell nanostructures coated with polyethylene glycol (PEG) has been reported for near-infrared (NIR) photothermal therapy (PTT) of prostate cancer. The next critical step for PTT is the visualization of therapeutically relevant nanoshell (NS) concentrations at the tumor site. Here we report the synthesis of PEGylated Gd2O3-mesoporous silica/gold core/shell NSs (Gd2O3-MS NSs) with NIR photothermal properties that also supply sufficient MRI contrast to be visualized at therapeutic doses (≥108 NSs per milliliter). The nanoparticles have r1 relaxivities more than three times larger than those of conventional T1 contrast agents, requiring less concentration of Gd3+ to observe an equivalent signal enhancement in T1-weighted MR images. Furthermore, Gd2O3-MS NS nanoparticles have r2 relaxivities comparable to those of existing T2 contrast agents, observed in agarose phantoms. This highly unusual combination of simultaneous T1 and T2 contrast allows for MRI enhancement through different approaches. As a rudimentary example, we demonstrate T1/T2 ratio MR images with sixfold contrast signal enhancement relative to its T1 MRI and induced temperature increases of 20 to 55 °C under clinical illumination conditions. These nanoparticles facilitate MRI-guided PTT while providing real-time temperature feedback through thermal MRI mapping.

Ultrasound controlled mechanophore activation in hydrogels for cancer therapy.

Mechanophores are molecular motifs that respond to mechanical perturbance with targeted chemical reactions toward desirable changes in material properties. A large variety of mechanophores have been investigated, with applications focusing on functional materials, such as strain/stress sensors, nanolithography, and self-healing polymers, among others. The responses of engineered mechanophores, such as light emittance, change in fluorescence, and generation of free radicals (FRs), have potential for bioimaging and therapy. However, the biomedical applications of mechanophores are not well explored. Herein, we report an in vitro demonstration of an FR-generating mechanophore embedded in biocompatible hydrogels for noninvasive cancer therapy. Controlled by high-intensity focused ultrasound (HIFU), a clinically proven therapeutic technique, mechanophores were activated with spatiotemporal precision to generate FRs that converted to reactive oxygen species (ROS) to effectively kill tumor cells. The mechanophore hydrogels exhibited no cytotoxicity under physiological conditions. Upon activation with HIFU sonication, the therapeutic efficacies in killing in vitro murine melanoma and breast cancer tumor cells were comparable with lethal doses of H2O2 This process demonstrated the potential for mechanophore-integrated HIFU combination as a noninvasive cancer treatment platform, named "mechanochemical dynamic therapy" (MDT). MDT has two distinct advantages over other noninvasive cancer treatments, such as photodynamic therapy (PDT) and sonodynamic therapy (SDT). 1) MDT is ultrasound based, with larger penetration depth than PDT. 2) MDT does not rely on sonosensitizers or the acoustic cavitation effect, both of which are necessary for SDT. Taking advantage of the strengths of mechanophores and HIFU, MDT can provide noninvasive treatments for diverse cancer types.

PEGylated liposomes for diagnosis of polyethylene glycol allergy.

BACKGROUND: Polyethylene glycol (PEG) is a nonprotein polymer that is present in its native (unbound) form as an excipient in a range of products. It is increasingly being utilized clinically in the form of PEGylated liposomal medications and vaccines. PEG is the cause of anaphylaxis in a small percentage of drug reactions; however, diagnosis of PEG allergy is complicated by the variable and poor diagnostic performance of current skin testing protocols. OBJECTIVE: We assessed the diagnostic performance of PEGylated lipid medications as an alternative to currently described tests that use medications containing PEG excipients. METHODS: Nine patients with a strong history of PEG allergy were evaluated by skin testing with a panel of PEG-containing medications and with a PEGylated lipid nanoparticle vaccine (BNT162b2). Reactivity of basophils to unbound and liposomal PEG was assessed ex vivo, and specificity of basophil responses to PEGylated liposomes was investigated with a competitive inhibition assay. More detailed information is provided in this article's Methods section in the Online Repository available at www.jacionline.org. RESULTS: Despite compelling histories of anaphylaxis to PEG-containing medications, only 2 (22%) of 9 patients were skin test positive for purified PEG or their index reaction-indicated PEG-containing compound. Conversely, all 9 patients were skin test positive or basophil activation test positive to PEGylated liposomal BNT162b2 vaccine. Concordantly, PEGylated liposomal drugs (BNT162b2 vaccine and PEGylated liposomal doxorubicin), but not purified PEG2000, consistently induced basophil activation ex vivo in patients with PEG allergy but not in nonallergic controls. Basophil reactivity to PEGylated nanoparticles competitively inhibited by preincubation of basophils with native PEG2000. CONCLUSION: Presentation of PEG on the surface of a lipid nanoparticle increases its in vivo and ex vivo allergenicity, and improves diagnosis of PEG allergy.

Sensing PEGylated Peptide Conformations Using a Protein Nanopore.

Membrane pores are exploited for the stochastic sensing of various analytes, and here, we use electrical recordings to explore the interaction of PEGylated peptides of different sizes with a protein pore, CymA. This wide-diameter natural pore comprises densely filled charged residues, facilitating electrophoretic binding of polyethylene glycol (PEG) tagged with a nonaarginine peptide. The small PEG 200 peptide conjugates produced monodisperse blockages and exhibited voltage-dependent translocation across the pores. Notably, the larger PEG 1000 and 2000 peptide conjugates yielded heterogeneous blockages, indicating a multitude of PEG conformations hindering their translocation through the pore. Furthermore, a much larger PEG 5000 peptide occludes the pore entrance, resulting in complete closure. The competitive binding of different PEGylated peptides with the same pore produced specific blockage signals reflecting their identity, size, and conformation. Our proposed model of sensing distinct polypeptide conformations corresponds to disordered protein unfolding, suggesting that this pore can find applications in proteomics.

Therapeutic Peptides Are Preferentially Solubilized in Specific Microenvironments within PEG-PLGA Polymer Nanoparticles.

Polymeric nanoparticles are a highly promising drug delivery formulation. However, a lack of understanding of the molecular mechanisms that underlie their drug solubilization and controlled release capabilities has hindered the efficient clinical translation of such technologies. Polyethylene glycol-poly(lactic-co-glycolic) acid (PEG-PLGA) nanoparticles have been widely studied as cancer drug delivery vehicles. In this letter, we use unbiased coarse-grained molecular dynamics simulations to model the self-assembly of a PEG-PLGA nanoparticle and its solubulization of the anticancer peptide, EEK, with good agreement with previously reported experimental structural data. We applied unsupervised machine learning techniques to quantify the conformations that polymers adopt at various locations within the nanoparticle. We find that the local microenvironments formed by the various polymer conformations promote preferential EEK solubilization within specific regions of the NP. This demonstrates that these microenvironments are key in controlling drug storage locations within nanoparticles, supporting the rational design of nanoparticles for therapeutic applications.

Lifting the Concentration Limit of Mass Photometry by PEG Nanopatterning.

Mass photometry (MP) is a rapidly growing optical technique for label-free mass measurement of single biomolecules in solution. The underlying measurement principle provides numerous advantages over ensemble-based methods but has been limited to low analyte concentrations due to the need to uniquely and accurately quantify the binding of individual molecules to the measurement surface, which results in diffraction-limited spots. Here, we combine nanoparticle lithography with surface PEGylation to substantially lower surface binding, resulting in a 2 orders of magnitude improvement in the upper concentration limit associated with mass photometry. We demonstrate the facile tunability of degree of passivation, enabling measurements at increased analyte concentrations. These advances provide access to protein-protein interactions in the high nanomolar to low micromolar range, substantially expanding the application space of mass photometry.

Regulating Second Coordination Shell of Ce Atom Site and Reshaping of Carrier Enable Single-Atom Nanozyme to Efficiently Express Oxidase-like Activity.

Single-atom nanozymes (SANs) are considered to be ideal substitutes for natural enzymes due to their high atom utilization. This work reported a strategy to manipulate the second coordination shell of the Ce atom and reshape the carbon carrier to improve the oxidase-like activity of SANs. Internally, S atoms were symmetrically embedded into the second coordination layer to form a Ce-N4S2-C structure, which reduced the energy barrier for O2 reduction, promoted the electron transfer from the Ce atom to O atoms, and enhanced the interaction between the d orbital of the Ce atom and p orbital of O atoms. Externally, in situ polymerization of mussel-inspired polydopamine on the precursor helps capture metal sources and protects the 3D structure of the carrier during pyrolysis. On the other hand, polyethylene glycol (PEG) modulated the interface of the material to enhance water dispersion and mass transfer efficiency. As a proof of concept, the constructed PEG@P@Ce-N/S-C was applied to the multimodal assay of butyrylcholinesterase activity.

Surface Passivation of Norland Optical Adhesive Improves the Guiding Efficiency of Gliding Microtubules in Microfluidic Devices.

The microtubule-kinesin biomolecular motor system, which is vital for cellular function, holds significant promise for nanotechnological applications. In vitro gliding assays have demonstrated the ability to transport microcargo by propelling microtubules across kinesin-coated surfaces. However, the uncontrolled directional motion of microtubules has posed significant challenges, limiting the system's application for precise cargo delivery. Microfluidic devices provide a means to direct microtubule movement through their geometric features. Norland Optical Adhesive (NOA) is valued for its mold-free application in microfluidic device fabrication; however, microtubules often climb up channel walls, limiting controlled movement. In this study, a surface passivation method for NOA is introduced, using polyethylene glycol via a thiol-ene click reaction. This technique significantly improved the directional control and concentration of microtubules within NOA microchannels. This approach presents new possibilities for the precise application of biomolecular motors in nanotechnology, enabling advancements in the design of microfluidic systems for complex biomolecular manipulations.

The molecular basis for hydrodynamic properties of PEGylated human serum albumin.

Polyethylene glycol (PEG) conjugation provides a protective modification that enhances the pharmacokinetics and solubility of proteins for therapeutic use. A knowledge of the structural ensemble of these PEGylated proteins is necessary to understand the molecular details that contribute to their hydrodynamic and colligative properties. Because of the large size and dynamic flexibility of pharmaceutically important PEGylated proteins, the determination of structure is challenging. In addition, the hydration of these conjugates that contain large polymers is difficult to determine with traditional methods that identify only first shell hydration water, which does not account for the complete hydrodynamic volume of a macromolecule. Here, we demonstrate that structural ensembles, generated by coarse-grained simulations, can be analyzed with HullRad and used to predict sedimentation coefficients and concentration-dependent hydrodynamic and diffusion nonideality coefficients of PEGylated proteins. A knowledge of these concentration-dependent properties enhances the ability to design and analyze new modified protein therapeutics. HullRad accomplishes this analysis by effectively accounting for the complete hydration of a macromolecule, including that of flexible polymers.

Hydrodynamic and thermodynamic analysis of PEGylated human serum albumin.

Covalent labeling of therapeutic drugs and proteins with polyethylene glycol (PEGylation) is an important modification for improving stability, solubility, and half-life. PEGylation alters protein solution behavior through its impact on thermodynamic nonideality by increasing the excluded volume, and on hydrodynamic nonideality by increasing the frictional drag. To understand PEGylation's impact, we investigated the thermodynamic and hydrodynamic properties of a model system consisting of PEGylated human serum albumin derivatives using analytical ultracentrifugation (AUC) and dynamic light scattering (DLS). We constructed PEGylated human serum albumin derivatives of single, linear 5K, 10K, 20K, and 40K PEG chains and a single branched-chain PEG of 40K (2 × 20K). Sedimentation velocity (SV) experiments were analyzed using SEDANAL direct boundary fitting to extract ideal sedimentation coefficients so, hydrodynamic nonideality ks, and thermodynamic nonideality 2BM1SV terms. These quantities allow the determination of the Stokes radius Rs, the frictional ratio f/fo, and the swollen or entrained volume Vs/v, which measure size, shape, and solvent interaction. We performed sedimentation equilibrium experiments to obtain independent measurements of thermodynamic nonideality 2BM1SE. From DLS measurements, we determined the interaction parameter, kD, the concentration dependence of the apparent diffusion coefficient, D, and from extrapolation of D to c = 0 a second estimate of Rs. Rs values derived from SV and DLS measurements and ensemble model calculations (see complementary study) are then used to show that ks + kD = theoretical 2B22M1. In contrast, experimental BM1 values from SV and sedimentation equilibrium data collectively allow for similar analysis for protein-PEG conjugates and show that ks + kD = 1.02-1.07∗BM1, rather than the widely used ks + kD = 2BM1 developed for hard spheres. The random coil behavior of PEG dominates the colloidal properties of PEG-protein conjugates and exceeds the sum of a random coil and hard-sphere volume due to excess entrained water.

Polyethylene Glycol Impacts Conformation and Dynamics of Escherichia coli Prolyl-tRNA Synthetase Via Crowding and Confinement Effects.

Polyethylene glycol (PEG) is a flexible, nontoxic polymer commonly used in biological and medical research, and it is generally regarded as biologically inert. PEG molecules of variable sizes are also used as crowding agents to mimic intracellular environments. A recent study with PEG crowders revealed decreased catalytic activity of Escherichia coli prolyl-tRNA synthetase (Ec ProRS), where the smaller molecular weight PEGs had the maximum impact. The molecular mechanism of the crowding effects of PEGs is not clearly understood. PEG may impact protein conformation and dynamics, thus its function. In the present study, the effects of PEG molecules of various molecular weights and concentrations on the conformation and dynamics of Ec ProRS were investigated using a combined experimental and computational approach including intrinsic tryptophan fluorescence spectroscopy, atomic force microscopy, and atomistic molecular dynamic simulations. Results of the present study suggest that lower molecular weight PEGs in the dilute regime have modest effects on the conformational dynamics of Ec ProRS but impact the catalytic function primarily via the excluded volume effect; they form large clusters blocking the active site pocket. In contrast, the larger molecular weight PEGs in dilute to semidilute regimes have a significant impact on the protein's conformational dynamics; they wrap on the protein surface through noncovalent interactions. Thus, lower-molecular-weight PEG molecules impact protein dynamics and function via crowding effects, whereas larger PEGs induce confinement effects. These results have implications for the development of inhibitors for protein targets in a crowded cellular environment.

Dual drug-loaded polymeric mixed micelles for ovarian cancer: Approach to enhanced therapeutic efficacy of albendazole and paclitaxel.

Chemotherapy resistance remains a significant challenge in treating ovarian cancer effectively. This study addresses this issue by utilizing a dual drug-loaded nanomicelle system comprising albendazole (ABZ) and paclitaxel (PTX), encapsulated in a novel carrier matrix of D-tocopheryl polyethylene glycol 1000 succinate vitamin E (TPGS), soluplus and folic acid. Our objective was to develop and optimize this nanoparticulate delivery system using solvent evaporation techniques to enhance the therapeutic efficacy against ovarian cancer. The formulation process involved pre-formulation, formulation, optimization, and comprehensive characterization of the micelles. Optimization was conducted through a 32 factorial design, focusing on the effects of polymer ratios on particle size, zeta potential, polydispersity index (PDI) and entrapment efficiency (%EE). The optimal formulation demonstrated improved dilution stability, as indicated by a critical micelle concentration (CMC) of 0.0015 mg/mL for the TPGS-folic acid conjugate (TPGS-FOL). Extensive characterization included differential scanning calorimetry (DSC), nuclear magnetic resonance (NMR), and Fourier-transform infrared spectroscopy (FTIR). The release profile exhibited an initial burst followed by sustained release over 90 h. The cytotoxic potential of the formulated micelles was superior to that of the drugs alone, as assessed by MTT assays on SKOV3 ovarian cell lines. Additionally, in vivo studies confirmed the presence of both drugs in plasma and tumour tissues, suggesting effective targeting and penetration. In conclusion, the developed TPGS-Fol-based nanomicelles for co-delivering ABZ and PTX show promising results in overcoming drug resistance, enhancing solubility, sustaining drug release, and improving therapeutic outcomes in ovarian cancer treatment.