Methotrexate, a structural analogue of folic acid, is one of the most effective and extensively used drugs for treating many kinds of cancer or severe and resistant forms of autoimmune diseases. In this paper, we take an overview of the present state of knowledge with regards to complex mechanisms of methotrexate action and its applications as immunosuppressive drug or chemotherapeutic agent in oncological combination therapy. In addition, the issue of the potential benefits of methotrexate in the development of neurological disorders in Alzheimer's disease or myasthenia gravis will be discussed.

https://www.mdpi.com/1422-0067/21/10/3483/pdf

Overview of Dual-Acting Drug Methotrexate in Different Neurological Diseases, Autoimmune Pathologies and Cancers.

From influenza to COVID-19: Lipid nanoparticle mRNA vaccines at the frontiers of infectious diseases.

Methotrexate (MTX) is the first line agent for therapy against rheumatoid arthritis (RA); however, orally its efficacy is hampered by poor solubility, less permeability, short plasma half-life, and reduced bioavailability. Meanwhile, parenteral formulations are associated with severe adverse effects. In an attempt to improve the efficacy of MTX, we synthesized polycaprolactone-polyethylene glycol-polycaprolactone (PCL-PEG-PCL) triblock copolymer by a ring-opening copolymerization reaction and used it as a carrier for the fabrication of MTX-loaded nanomicelles. Surfactant-free, self-assembled nanomicelles were prepared by nanoprecipitation technique and optimized through central composite design. The optimized nanomicelles exhibited a size distribution of 31 nm and an encapsulation efficiency of 91%. In vitro, the nanomicelles exhibited hemocompatibility, sustained release, and significantly high uptake in lipopolysaccharide activated macrophages. To facilitate application on the skin, optimized nanomicelles were loaded into a Carbopol 934-based hydrogel with eucalyptus oil as a penetration enhancer. Eucalyptus oil significantly improved the permeation of nanomicelles through the skin (p < 0.001). When the hydrogel was applied on the RA mice model, nanomicelles exhibited preferentially highest accumulation in the inflamed joints than other organs. As compared with the free MTX, MTX nanomicelles significantly improved the pharmacokinetic (4.34-fold greater half-life, 3.68-fold higher AUC0-t, and 3.15-fold higher mean residence time) and pharmacodynamic profile ascertained through low inflammatory cytokines expression, improved oxidation protection, recovered behavioral responses, and radiological analysis. MTX nanomicelles-based hydrogel also significantly reduced the hepatotoxicity and did not activate the immune system. These results suggest that the MTX-loaded nanomicelles-based transdermal hydrogel can prove to be a promising agent against RA.

Surfactant-Free, Self-Assembled Nanomicelles-Based Transdermal Hydrogel for Safe and Targeted Delivery of Methotrexate against Rheumatoid Arthritis.

The lymphatic system has an important defensive role in the human body. The metastasis of most tumors initially spreads through the surrounding lymphatic tissue and eventually forms lymphatic metastatic tumors; the tumor cells may even transfer to other organs to form other types of tumors. Clinically, lymphatic metastatic tumors develop rapidly. Given the limitations of surgical resection and the low effectiveness of radiotherapy and chemotherapy, the treatment of lymphatic metastatic tumors remains a great challenge. Lymph node metastasis may lead to the further spread of tumors and may be predictive of the endpoint event. Under these circumstances, novel and effective lymphatic targeted drug delivery systems have been explored to improve the specificity of anticancer drugs to tumor cells in lymph nodes. In this review, we summarize the principles of lymphatic targeted drug delivery and discuss recent advances in the development of lymphatic targeted carriers.

Recent advances in lymphatic targeted drug delivery system for tumor metastasis

Methotrexate (MTX) is a folic acid antagonist used as an effective drug to treat various kinds of cancers. However, MTX has limited use in cancer chemotherapy due to its adverse effects such as poor bioavailability, low specificity, drug resistance, and dose-dependent side effects. To improve lymphatic delivery and reduce toxicity of MTX, MTX-loaded nanoparticles (NPs) were prepared in the present study. NPs were prepared with double emulsion solvent evaporation method using poly(lactide-co-glycolide) (PLGA). NPs were assessed for size, encapsulation efficiency, morphology, Fourier-transform infrared spectroscopy, X-ray diffraction, and thermal characterization. In vitro release profiles and cytotoxicity of these NPs were also evaluated. Prepared NPs and free MTX were administered orally or intravenously (5 mg/kg as MTX) to rats to evaluate their pharmacokinetic characteristics and lymphatic delivery effects. Mean particle size and encapsulation efficiency of NPs were 163.7 ± 10.25 nm and 93.3 ± 0.5%, respectively. Prepared NPs showed a sustained release profile of MTX in vitro and may be effective to cancer cells. Area under the blood concentration-time curve, total clearance, half-life, and lymphatic targeting efficiency were significantly different (p < 0.05) between prepared NPs and free MTX. These results demonstrate that MTX-loaded PLGA NPs are good candidates for targeted delivery of MTX to the lymphatic system.

Preparation and In Vitro/In Vivo Characterization of Polymeric Nanoparticles Containing Methotrexate to Improve Lymphatic Delivery.

Diethyl phthalate (DEP) belongs to phthalates with short alkyl chains. It is a substance frequently used to make various products. Thus, humans are widely exposed to DEP from the surrounding environment such as food, soil, air, and water. As previously reported in many studies, DEP is an endocrine disruptor with reproductive toxicity. Monoethyl phthalate (MEP), a major metabolite of DEP in vivo, is a biomarker for DEP exposure assessment. It is also an endocrine disruptor with reproductive toxicity, similar to DEP. However, toxicokinetic studies on both MEP and DEP have not been reported in detail yet. Therefore, the objective of this study was to evaluate and develop physiologically based pharmacokinetic (PBPK) model for both DEP and MEP in rats and extend this to human risk assessment based on human exposure. This study was conducted in vivo after intravenous or oral administration of DEP into female (2 mg/kg dose) and male (0.1–10 mg/kg dose) rats. Biological samples consisted of urine, plasma, and 11 different tissues. These samples were analyzed using UPLC–ESI–MS/MS method. For DEP, the tissue to plasma partition coefficient was the highest in the kidney, followed by that in the liver. For MEP, the tissue to plasma partition coefficient was the highest in the liver. It was less than unity in all other tissues. Plasma, urine, and fecal samples were also obtained after IV administration of MEP (10 mg/kg dose) to male rats. All results were reflected in a model developed in this study, including in vivo conversion from DEP to MEP. Predicted concentrations of DEP and MEP in rat urine, plasma, and tissue samples using the developed PBPK model fitted well with observed values. We then extrapolated the PBPK model in rats to a human PBPK model of DEP and MEP based on human physiological parameters. Reference dose of 0.63 mg/kg/day (or 0.18 mg/kg/day) for DEP and external doses of 0.246 μg/kg/day (pregnant), 0.193 μg/kg/day (fetus), 1.005–1.253 μg/kg/day (adults), 0.356–0.376 μg/kg/day (adolescents), and 0.595–0.603 μg/kg/day (children) for DEP for human risk assessment were estimated using Korean biomonitoring values. Our study provides valuable insight into human health risk assessment regarding DEP exposure.

Risk assessment for humans using physiologically based pharmacokinetic model of diethyl phthalate and its major metabolite, monoethyl phthalate

Methotrexate, which is widely used in the treatment of cancer and immune-related diseases, has limitations in use because of its low bioavailability, short half-life, and tissue toxicity. Thus, in this study, a nano-sized water-in-oil-in-water (W/O/W) double emulsion containing methotrexate was prepared to enhance its lymphatic delivery and bioavailability. Based on the results from solubility testing and a pseudo-ternary diagram study, olive oil as the oil, Labrasol as a surfactant, and ethanol as a co-surfactant, were selected as the optimal components for the nanoemulsion. The prepared nanoemulsion was evaluated for size, zeta potential, encapsulation efficiency, pH, morphology, and in vitro release profiles. Furthermore, pharmacokinetics and lymphatic targeting efficiency were assessed after oral and intravenous administration of methotrexate-loaded nanoemulsion to rats. Mean droplet size, zeta potential, encapsulation efficiency, and pH of formulated nanoemulsion were 173.77 ± 5.76 nm, -35.63 ± 0.78 mV, 90.37 ± 0.96%, and 4.07 ± 0.03, respectively. In vitro release profile of the formulation indicated a higher dissolution and faster rate of methotrexate than that of free drug. The prepared nanoemulsion showed significant increases in maximum plasma concentration, area under the plasma concentration-time curve, half-life, oral bioavailability, and lymphatic targeting efficiency in both oral and intravenous administration. Therefore, our research proposes a methotrexate-loaded nanoemulsion as a good candidate for enhancing targeted lymphatic delivery of methotrexate.

https://www.mdpi.com/1999-4923/12/10/978/pdf

Enhanced Lymphatic Delivery of Methotrexate Using W/O/W Nanoemulsion: In Vitro Characterization and Pharmacokinetic Study.

With the current advance in nanotechnology, the development has accelerated of a number of nanoparticle-type drugs such as nano-emulsions, lipid emulsions, liposomes, and cell therapeutics. With these developments, attempts are being made to apply these new drugs to healing many intractable diseases related to antibody production, autoimmune disorders, cancer, and organ transplantation in both clinical and nonclinical trials. Drug delivery to the lymphatic system is indispensable for treating these diseases, but the core technologies related to the in vivo distribution characteristics and lymphatic delivery evaluation of these particle-type drugs have not yet been established. Additionally, the core technologies for setting up the pharmacotherapeutic aspects such as their usage and dosages in the development of new drugs do not meet the needs of the market. Therefore, it is necessary to consider dividing these particle-type drugs into soft-lipid nanoparticles that can change size in the process of body distribution and hard-lipid nanoparticles whose surfaces are hardened and whose sizes do not easily change in vivo; these soft- and hard-lipid nanoparticles likely possess different biodistribution characteristics including delivery to the lymphatic system. In this review, we summarize the different types, advantages, limitations, possible remedies, and body distribution characteristics of soft- and hard-lipid nanoparticles based on their administration routes. We also emphasize that it will be necessary to fully understand the differences in distribution between these soft- and hard-lipid nanoparticle-type drugs and to establish pharmacokinetic models for their more ideal lymphatic delivery.

Soft- and hard-lipid nanoparticles: a novel approach to lymphatic drug delivery

The purpose of this study was to develop a sensitive method for quantifying cinnamic acid in human plasma using UPLC–ESI–MS/MS. Cinnamic acid was separated using water containing 0.005% (v/v) formic acid and acetonitrile as a mobile phase by gradient elution at a flow rate 0.2 mL/min, equipped with a HALO-C18 column (2.1 × 100 mm, 2.7 μm). Quantitation of this analysis was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in multiple reaction monitoring negative ion mode. The mass transitions were m/z 146.8 → 103.0 for cinnamic acid, and 432.9 → 225.0 for geniposide as internal standard. Liquid–liquid extraction and protein precipitation with ethyl acetate–methanol-1.2% acetic acid (4/16/1, v/v/v) were used in the sample extraction. The chromatograms showed high resolution, sensitivity, and selectivity with no interference with plasma constituents. The calibration curves for cinnamic acid in human plasma were 0.1–500 ng/mL and displayed excellent linearity with correlation coefficients (r) greater than 0.99. Both the intra- and inter-day precisions (CV%) were within 3.88% for human plasma. The accuracy was 99.34–106.69% for human plasma. In vitro plasma protein binding of cinnamic acid was 64.26 ± 1.89 and 65.50 ± 1.78% for the spiked human plasma concentrations of 100 and 1000 ng/mL, respectively. The developed analytical method satisfied the criteria of international guidance and could be successfully applied to the pharmacokinetic study of cinnamic acid after oral administration of Socheongryong-tang tablets to humans.

A sensitive UPLC–ESI–MS/MS method for the quantification of cinnamic acid in vivo and in vitro: application to pharmacokinetic and protein binding study in human plasma

Seung-Hyun Jeong

Papers

Preparation and In Vitro/In Vivo Characterization of Polymeric Nanoparticles Containing Methotrexate to Improve Lymphatic Delivery.

Risk assessment for humans using physiologically based pharmacokinetic model of diethyl phthalate and its major metabolite, monoethyl phthalate

Enhanced Lymphatic Delivery of Methotrexate Using W/O/W Nanoemulsion: In Vitro Characterization and Pharmacokinetic Study.

Soft- and hard-lipid nanoparticles: a novel approach to lymphatic drug delivery

A sensitive UPLC–ESI–MS/MS method for the quantification of cinnamic acid in vivo and in vitro: application to pharmacokinetic and protein binding study in human plasma