Drug Loading Techniques Research Articles

The Evolution of Microfluidic-Based Drug-Loading Techniques for Cells and Their Derivatives.

Conventional drug delivery techniques face challenges related to targeting and adverse reactions. Recent years have witnessed significant advancements in nanoparticle-based drug carriers. Nevertheless, concerns persist regarding their safety and insufficient metabolism. Employing cells and their derivatives, such as cell membranes and extracellular vesicles (EVs), as drug carriers effectively addresses the challenges associated with nanoparticle carriers. However, an essential hurdle remains in efficiently loading drugs into these carriers. With the advancement of microfluidic technology and its advantages in precise manipulation at the micro- and nanoscales, as well as minimal sample loss, it has found extensive application in the loading of drugs using cells and their derivatives, thereby fostering the development of drug-loading techniques. This paper outlines the characteristics and benefits of utilizing cells and their derivatives as drug carriers and provides an overview of current drug-loading techniques, particularly those rooted in microfluidic technology. The significant potential for microfluidic technology in targeted disease therapy through drug delivery systems employing cells and their derivatives, is foreseen.

A Systematic Review on Plant-Derived Extracellular Vesicles as Drug Delivery Systems.

This systematic review offers a comprehensive analysis of plant-derived extracellular vesicles (PDEVs) as emerging drug delivery systems, focusing on original research articles published between 2016 and 2024 that exclusively examine the use of PDEVs for drug delivery. After a rigorous search across multiple databases, 20 relevant studies out of 805 initial results were selected for analysis. This review systematically summarizes the critical data on PDEV components, isolation methods, and drug-loading techniques. It highlights the potential of PDEVs to significantly enhance drug safety and efficacy, reduce dosage and toxicity, and align drug development with sustainable and environmentally friendly biotechnological processes. This review also emphasizes the advantages of PDEVs over mammalian-derived vesicles, such as cost-effectiveness, higher yield, and reduced immunogenicity. Additionally, it explores the synergistic potential between encapsulated drugs and bioactive compounds naturally present in PDEVs. This study acknowledges the challenges in standardizing isolation and formulation methods for clinical use. Overall, this review provides valuable insights into the current state and future directions of PDEV-based drug delivery systems, highlighting their promising role in advancing pharmaceutical research and development.

Development of Focused Ultrasound-Assisted Nanoplexes for RNA Delivery.

RNA-based therapeutics, including siRNA, have obtained recognition in recent years due to their potential to treat various chronic and rare diseases. However, there are still limitations to lipid-based drug delivery systems in the clinical use of RNA therapeutics due to the need for optimization in the design and the preparation process. In this study, we propose adaptive focused ultrasound (AFU) as a drug loading technique to protect RNA from degradation by encapsulating small RNA in nanoliposomes, which we term nanoplexes. The AFU method is non-invasive and isothermal, as nanoplexes are produced without direct contact with any external materials while maintaining precise temperature control according to the desired settings. The controllability of sample treatments can be effectively modulated, allowing for a wide range of ultrasound intensities to be applied. Importantly, the absence of co-solvents in the process eliminates the need for additional substances, thereby minimizing the potential for cross-contaminations. Since AFU is a non-invasive method, the entire process can be conducted under sterile conditions. A minimal volume (300 μL) is required for this process, and the treatment is speedy (10 min in this study). Our in vitro experiments with silencer CD44 siRNA, which performs as a model therapeutic drug in different mammalian cell lines, showed encouraging results (knockdown > 80%). To quantify gene silencing efficacy, we employed quantitative polymerase chain reaction (qPCR). Additionally, cryo-electron microscopy (cryo-EM) and atomic force microscopy (AFM) techniques were employed to capture images of nanoplexes. These images revealed the presence of individual nanoparticles measuring approximately 100-200 nm in contrast with the random distribution of clustered complexes observed in ultrasound-untreated samples of liposome nanoparticles and siRNA. AFU holds great potential as a standardized liposome processing and loading method because its process is fast, sterile, and does not require additional solvents.

Extracellular vesicles containing fullerene derivatives prepared by an exchange reaction for photodynamic therapy.

Extracellular vesicles (EVs) have excellent biocompatibility and long retention times in the circulation and have consequently been expected to be useful as drug-delivery systems. However, their applications have been limited because of the inability to introduce hydrophobic compounds to EVs without the use of harmful organic solvents. Herein, we developed an organic-solvent-free drug-loading technique based on the host exchange reaction. We demonstrated that the exchange reaction enabled quantitative loading of EVs with highly concentrated (0.1 mM) hydrophobic fullerene derivatives. Fullerene derivative-loaded EVs (EVs/C60) could eliminate cancer cell lines more efficiently than fullerene derivative-loaded liposomes (Lip/C60). Moreover, the photodynamic activity of EVs/C60 was fivefold higher than that of the clinically available photosensitizer photofrin. EVs/C60 could efficiently suppress tumor growth in tumor-xenograft model mice.

Tackling Esophageal Squamous Cell Carcinoma with ITFn-Pt(IV): A Novel Fusion of PD-L1 Blockade, Chemotherapy, and T-cell Activation.

PD-1/PD-L1 blockade immunotherapy has gained approval for the treatment of a diverse range of tumors; however, its efficacy is constrained by the insufficient infiltration of T lymphocytes into the tumor microenvironment, resulting in suboptimal patient responses. Here, a pioneering immunotherapy ferritin nanodrug delivery system denoted as ITFn-Pt(IV) is introduced. This system orchestrates a synergistic fusion of PD-L1 blockade, chemotherapy, and T-cell activation, aiming to augment the efficacy of tumor immunotherapy. Leveraging genetic engineering approach and temperature-regulated channel-based drug loading techniques, the architecture of this intelligent responsive system is refined. It is adept at facilitating the precise release of T-cell activating peptide Tα1 in the tumor milieu, leading to an elevation in T-cell proliferation and activation. The integration of PD-L1 nanobody KN035 ensures targeted engagement with tumor cells and mediates the intracellular delivery of the encapsulated Pt(IV) drugs, culminating in immunogenic cell death and the subsequent dendritic cell maturation. Employing esophageal squamous cell carcinoma (ESCC) as tumor model, the potent antitumor efficacy of ITFn-Pt(IV) is elucidated, underscored by augmented T-cell infiltration devoid of systemic adverse effects. These findings accentuate the potential of ITFn-Pt(IV) for ESCC treatment and its applicability to other malignancies resistant to established PD-1/PD-L1 blockade therapies.

HPLC-UV evaluation of a microwave assisted method as an active drug loading technique for exosome-based drug delivery system

This paper evaluates the potential of a microwave radiation (MR) assisted method as an active drug loading technique for exosomes using polyphenolic nutraceuticals as model drugs (i.e. resveratrol (RV), rosmarinic acid (RA), pterostilbene (PT) and epigallocatechin gallate (EG)). MR is evaluated as a single step method and as part of a two-step method consisting of incubation (IN) followed by MR. The effect of exposure time, loading method and type of nutraceutical on the loading efficiency were investigated using high performance liquid chromatography (HPLC), X-ray diffraction (XRD) and flow cytometry. Additionally, dynamic light scattering (DLS) was used to determine the size of exosomes. Loading efficiency results indicated that MR is a promising method to be used as loading process. Results also suggested that due to different levels of hydrophobicity, related to the number of OH groups, the absorption of polyphenols into the bilayer of EVs is different for each molecule. According to XRD results, MR could not be used with any cargo drug since radiation could affect the chemical composition and the degree of crystallinity of such molecules, consequently affecting their performance. Flow cytometry results indicated that loading methods negatively affect exosome concentration.

Therapeutic Agent-Loaded Fibrous Scaffolds for Biomedical Applications.

Tissue engineering is a sophisticated field that involves the integration of various disciplines, such as clinical medicine, material science, and life science, to repair or regenerate damaged tissues and organs. To achieve the successful regeneration of damaged or diseased tissues, it is necessary to fabricate biomimetic scaffolds that provide structural support to the surrounding cells and tissues. Fibrous scaffolds loaded with therapeutic agents have shown considerable potential in tissue engineering. In this comprehensive review, we examine various methods for fabricating bioactive molecule-loaded fibrous scaffolds, including preparation methods for fibrous scaffolds and drug-loading techniques. Additionally, we delved into the recent biomedical applications of these scaffolds, such as tissue regeneration, inhibition of tumor recurrence, and immunomodulation. The aim of this review is to discuss the latest research trends in fibrous scaffold manufacturing methods, materials, drug-loading methods with parameter information, and therapeutic applications with the goal of contributing to the development of new technologies or improvements to existing ones.

Microbubbles Contrast Agents: General Overview as Diagnostics and Therapeutic Agent.

Recent discoveries have unfolded many powerful emerging applications in the field of drug delivery science. For the past few years, ultrasound mediated microbubble contrast agents have been an emerging modality for diagnostic and drug delivery applications. Microbubbles are small spherical bubbles composed of a gas core encapsulated by a shell with different materials. The composition of the microbubble determines its stiffness, encapsulation efficiency, stability, and clearance from the system. A gas-filled microbubble, when activated by an acoustic pulse, can produce large volumetric oscillations and, once administered intravenously, can act as a cavitating nuclei, allowing for a wide range of ultrasound-assisted drug delivery applications. Microbubbles offer a fantastic approach to ultrasound triggered drug delivery with various drug loading techniques and targeting strategies for the uptake of bioactive substances such as polynucleotides, proteins, genes, and small-molecule drugs. Microbubbles can be used for several diagnostic and therapeutic purposes for accurate detection and treatment of various life-threatening diseases.

Fungal Keratitis Treatment Using Drug-Loaded Hyaluronic Acid Microgels.

Antifungal drug-loaded hyaluronic acid (HA) microgels using conjugation and encapsulation drug-loading techniques were utilized in the treatment of fungal keratitis. Natamycin (NAT) and amphotericin B (AMB) drugs were chemically linked to HA microgels by employing a chemical coupling agent to obtain conjugated (C-) HA:NAT and HA:AMB microgels. Also, these drugs were loaded into the HA microgel network during HA microgel preparation to attain encapsulated (E-) HA:NAT and HA:AMB microgels. The conjugation of drug molecules was confirmed by FT-IR spectra of bare and drug-loaded HA microgels. It was determined that the AMB loading amount was about 4-fold higher for E-HA:AMB in comparison to C-HA:AMB microgels. Furthermore, the antifungal activity of drug conjugated and encapsulated HA:NAT and HA:AMB microgels was tested on Fusarium sp. and compared with the effect of bare drug molecules as control for up to 15 days of incubation time by means of the disc diffusion technique. The antifungal activity of 200 μL at 20 mg/mL concentration of C-HA:NAT and C-HA:AMB microgels was not found to effectively inhibit Fusarium sp. growth after 1 day of incubation, whereas the same concentration of E-HA:NAT and E-HA:AMB microgels totally killed Fusarium sp. for up to 15 days. These E-HA:NAT and E-HA:AMB microgels show no cytotoxicity on the L929 fibroblast cells up to 1000 μg/mL concentration, whereas the free drug molecules destroy the cells even at 100 μg/mL concentration.

Theragnostic Applications of Mammal and Plant-Derived Extracellular Vesicles: Latest Findings, Current Technologies, and Prospects.

The way cells communicate is not fully understood. However, it is well-known that extracellular vesicles (EVs) are involved. Researchers initially thought that EVs were used by cells to remove cellular waste. It is now clear that EVs function as signaling molecules released by cells to communicate with one another, carrying a cargo representing the mother cell. Furthermore, these EVs can be found in all biological fluids, making them the perfect non-invasive diagnostic tool, as their cargo causes functional changes in the cells upon receiving, unlike synthetic drug carriers. EVs last longer in circulation and instigate minor immune responses, making them the perfect drug carrier. This review sheds light on the latest development in EVs isolation, characterization and, application as therapeutic cargo, novel drug loading techniques, and diagnostic tools. We also address the advancement in plant-derived EVs, their characteristics, and applications; since plant-derived EVs only recently gained focus, we listed the latest findings. Although there is much more to learn about, EV is a wide field of research; what scientists have discovered so far is fascinating. This paper is suitable for those new to the field seeking to understand EVs and those already familiar with it but wanting to review the latest findings.

Exosomes as Emerging Drug Delivery and Diagnostic Modality for Breast Cancer: Recent Advances in Isolation and Application.

Breast cancer (BC) is the most common type of malignancy which covers almost one-fourth of all the cancers diagnosed in women. Conventionally, chemo-, hormonal-, immune-, surgery, and radiotherapy are the clinically available therapies for BC. However, toxicity and other related adverse effects are still the major challenges. A variety of nano platforms have been reported to overcome these limitations, among them, exosomes provide a versatile platform not only for the diagnosis but also as a delivery vehicle for drugs. Exosomes are biological nanovesicles made up of a lipidic bilayer and known for cell-to-cell communication. Exosomes have been reported to be present in almost all bodily fluids, viz., blood, milk, urine, saliva, pancreatic juice, bile, peritoneal, and cerebrospinal fluid. Such characteristics of exosomes have attracted immense interest in cancer diagnosis and therapy. They can deliver bioactive moieties such as protein, lipids, hydrophilic as well as hydrophobic drugs, various RNAs to both distant and nearby recipient cells as well as have specific biological markers. By considering the growing interest of the scientific community in this field, we comprehensively compiled the information about the biogenesis of exosomes, various isolation methods, the drug loading techniques, and their diverse applications in breast cancer diagnosis and therapy along with ongoing clinical trials which will assist future scientific endeavors in a more organized direction.

Preparation and Evaluation of rhINF-α-2b Sodium Hyaluronate Cross-Linked Porous Microspheres: Characterization, Sustained-Release Properties, and Antitumor Activity.

Recombinant human interferon α-2b (rhINF-α-2b), like most proteins, has several shortcomings such as relatively short half-life, low therapeutic index, high circulating drug fluctuations, and rapid degradation which could hinder its effective delivery. Novel electrostatic spray and ion exchange drug-loading techniques were combined to formulate rhINF-α-2b sodium hyaluronate cross-linked porous sustained-release microspheres (rhINF-α-2b-SHCPM), a protein delivery system. The different properties of rhINF-α-2b-SHCPM including the physicochemical nature, in vitro release behavior, and antitumor activity were evaluated. The loading rate (10.31 ± 0.94%) and encapsulation efficiency (89.09 ± 2.37%) of rhINF-α-2b-SHCPM produced acceptable values. The in vitro cumulative release rate of rhINF-α-2b-SHCPM within 24 h was also 86.26 ± 2.11% with a much better sustained release effect. Thus, the half-life (10.763 h) and retention time (14.067 h) of rhIFNα-2b-SHCPM were significantly prolonged with enhanced bioavailability (43,198.387 ng/L*h) and decreased peak concentration (15,266.4 ngL-1) compared with the free rhIFNα-2b protein (0.912 h, 0.952 h, 34,749.048 ng/L*h, and 48,870.2 ngL-1, respectively). The in vitro anti-proliferative activity and in vivo tumor inhibitory rate of rhIFNα-2b-SHCPM also increased by 90 and 55.86%, respectively, compared with the free rhIFNα-2b solution. The findings significantly supported a well-developed protein delivery system with improved sustained release, acceptable bioavailability, and increased antitumor activities. Graphical Abstract.

Mild Hyperthermia Responsive Liposomes for Enhanced In Vitro and In Vivo Anticancer Efficacy of Doxorubicin against Hepatocellular Carcinoma.

The current study is aimed to fabricate doxorubicin (Dox) loaded mild temperature responsive liposomes (MTLs) by thin film hydration technique for enhanced in vitro and in vivo anticancer efficacy against hepatocellular carcinoma. The aforementioned Dox loaded MTLs were developed and optimized with extrusion and drug loading techniques. The optimized MTLs were in optimum size range (118.20 ± 2.81–187.13 ± 4.15 nm), colloidal stability (−13.27 ± 0.04 to −32.34 ± 0.15 mV), and enhanced entrapment of Dox (28.71 ± 2.01–79.24 ± 2.16). Furthermore, the optimized formulation (MTL1-E(AL)) embodied improved physicochemical stability deducted by Fourier transform infra-red (FTIR) spectroscopy and mild hyperthermia-based phase transition demonstrated from differential scanning calorimetry (DSC). An in vitro drug release study revealed mild hyperthermia assisted rapid in vitro Dox release from MTLs-E(AL) (T100% ≈ 1 h) by Korsmeyer–Peppas model based Fickian diffusion (n < 0.45). Likewise, an in vitro cytotoxicity study and lower IC50 values also symbolized mild hyperthermia (40.2 °C) based quick and improved cytotoxicity of MTL1-E(AL) in HepG2 and MCF-7 cells than Dox. The fluorescence microscopy also represented enhanced cellular internalization of MTL1-E(AL) at mild hyperthermia compared to the normothermia (37.2 °C). In addition, an in vivo animal study portrayed the safety, improved anticancer efficacy and healing of hepatocellular carcinoma (HCC) through MTL1-E(AL). In brief, the Dox loaded MTLs could be utilized as safe and effective therapeutic strategy against HCC.

Drug loading techniques for exosome-based drug delivery systems.

Exosomes are bilayer membrane-coated extracellular vesicles measuring between 40 and 100 nm in diameter. As a natural carrier, exosomes have the advantages of low immunogenicity, high stability in blood, and direct delivery of drugs to cells. Exosomes can be transported between cells and thus are conducive to the exchange of substances and information between cells. They change the functional state of recipient cells by loading exogenous drugs (e.g., small-molecule drugs, transmembrane proteins, and nucleic acid drugs). The key to using exosomes as drug carriers is the effective loading of exogenous drugs into exosomes; however, this task poses a challenge in studying the functionalization of exosomes as drug carriers. Currently, sonication, electroporation, transfection, incubation, extrusion, saponin-assisted loading, transgenesis, freeze-thaw cycles, thermal shock, pH gradient method, and hypotonic dialysis have been applied to load these drugs into exosomes. This review aims to provide an overview of the advantages and disadvantages of various drug loading technologies for exosomes.

Halloysite Nanotubes for Nanomedicine: Prospects, Challenges and Applications

Recently, naturally occurring and abundantly available halloysite nanoclay has emerged as a nanomaterial carrier system suitable for both controlled and sustained delivery. These aluminosilicate tubes of 50-nm diameter, with outer silica and inner alumina layers, possess a tubular structure, with excellent features such as large aspect ratio, good biocompatibility, and high mechanical strength entails them to be suitable for drug delivery. The unique features of these nanotubes are the ability to load DNA and enzyme due to opposite charges on both inner and outer surfaces which allow selective drug loading on both surfaces. This review article emphasizes on the drug loading techniques with release characteristics and some important applications in biomedical, environmental fields, and different types of medication. Being most versatile, these nanotubes can be used widely as a carrier system for enzyme bases and different types of the nucleus which can be a most promising step for novel drug delivery systems.

A facile and universal method to achieve liposomal remote loading of non-ionizable drugs with outstanding safety profiles and therapeutic effect

Liposomes have made remarkable achievements as drug delivery vehicles in the clinic. Liposomal products mostly benefited from remote drug loading techniques that succeeded in amphipathic and/or ionizable drugs, but seemed impracticable for nonionizable and poorly water-soluble therapeutic agents, thereby impeding extensive promising drugs to hitchhike liposomal vehicles for disease therapy. In this study, a series of weak acid drug derivatives were designed by a simplistic one step synthesis, which could be remotely loaded into liposomes by pH gradient method. Cabazitaxel (CTX) weak acid derivatives were selected to evaluate regarding its safety profiles, pharmacodynamics, and pharmacokinetics. CTX weak acid derivative liposomes were superior to Jevtana® in terms of safety profiles, including systemic toxicity, hematological toxicity, and potential central nerve toxicity. Specifically, it was demonstrated that liposomes had capacity to weaken potential toxicity of CTX on cortex and hippocampus neurons. Significant advantages of CTX weak acid derivative-loaded liposomes were achieved in prostate cancer and metastatic cancer therapy resulting from higher safety and elevated tolerated doses.

Ultrasound-responsive lipid microbubbles for drug delivery: A review of preparation techniques to optimise formulation size, stability and drug loading.

Lipid-shelled microbubbles have received extensive interest to enhance ultrasound-responsive drug delivery outcomes due to their high biocompatibility. While therapeutic effectiveness of microbubbles is well established, there remain limitations in sample homogeneity, stability profile and drug loading properties which restrict these formulations from seeing widespread use in the clinical setting. In this review, we evaluate and discuss the most encouraging leads in lipid microbubble design and optimisation. We examine current applications in drug delivery for the systems and subsequently detail shell compositions and preparation strategies that improve monodispersity while retaining ultrasound responsiveness. We review how excipients and storage techniques help maximise stability and introduce different characterisation and drug loading techniques and evaluate their impact on formulation performance. The review concludes with current quality control measures in place to ensure lipid microbubbles can be reproducibly used in drug delivery.

Lipoprotein-based drug delivery.

Lipoproteins (LPs) are circulating heterogeneous nanoparticles produced by the liver and intestines. LPs play a major role in the transport of dietary and endogenous lipids to target cells through cell membrane receptors or cell surface-bound lipoprotein lipase. The stability, biocompatibility, and selective transport of LPs make them promising delivery vehicles for various therapeutic and imaging agents. This review discusses isolation, manufacturing, and drug loading techniques used for LP-based drug delivery, as well as recent applications for diagnosis and treatment of cancer, atherosclerosis, and other life-threatening diseases.

Nanostructured ZnO as Multifunctional Carrier for a Green Antibacterial Drug Delivery System-A Feasibility Study.

The physico–chemical and biological properties of nanostructured ZnO are combined with the non-toxic and eco-friendly features of the scCO2-mediated drug loading technique to develop a multifunctional antimicrobial drug delivery system for potential applications in wound healing. Two nanostructured ZnO (NsZnO) with different morphologies were prepared through wet organic-solvent-free processes and characterized by means of powder X-ray diffraction, field emission scanning electron microscopy (FESEM), and nitrogen adsorption analysis. The antimicrobial activity of the two samples against different microbial strains was investigated together with the in vitro Zn2+ release. The results indicated that the two ZnO nanostructures exhibited the following activity: S. aureus > C. albicans > K. pneumoniae. A correlation between the antimicrobial activity, the physico–chemical properties (specific surface area and crystal size) and the Zn2+ ion release was found. Ibuprofen was, for the first time, loaded on the NsZnO carriers with a supercritical CO2-mediated drug impregnation process and in vitro dissolution studies of the loaded drug were performed. A successful loading up to 14% w/w of ibuprofen in its amorphous form was obtained. A preliminary drug release test showed that up to 68% of the loaded ibuprofen could be delivered to a biological medium, confirming the feasibility of using NsZnO as a multifunctional antimicrobial drug carrier.

Biomorphic Ceramics for Drug Delivery in Bone Tissue Regeneration.

Incorporating therapeutic molecules into biomorphic ceramics for in situ drug release can be used to generate novel systems for tissue regeneration. These systems couple the complex hierarchical porous structures of biomorphic ceramics with the therapeutic activity of drugs. There are a large number of natural precursors available to be used as templates to obtain biomorphic silicon carbide ceramics. Additionally, different drug loading techniques can be used for these systems. The high versatility in structures and drugs allows the selection of the right structure-drug fit in each case according to the tissue needs. This paper reviews the utility of biomorphic ceramics for tissue engineering as well as their use for local drug release.