CHARMM-GUI Membrane Builder for Lipid Nanoparticles with Ionizable Cationic Lipids and PEGylated Lipids

A lipid nanoparticle (LNP) formulation is a state-of-the-art delivery system for genetic drugs such as DNA, mRNA, and siRNA, which is successfully applied to COVID-19 vaccines and gains tremendous interest in therapeutic applications. Despite its importance, a molecular-level understanding of the LNP structures and dynamics is still lacking, which makes a rational LNP design almost impossible. In this work, we present an extension of CHARMM-GUI Membrane Builder to model and simulate all-atom LNPs with various (ionizable) cationic lipids and PEGylated lipids (PEG-lipids). These new lipid types can be mixed with any existing lipid types with or without a biomolecule of interest, and the generated systems can be simulated using various molecular dynamics engines. As a first illustration, we considered model LNP membranes with DLin-KC2-DMA (KC2) or DLin-MC3-DMA (MC3) without PEG-lipids. The results from these model membranes are consistent with those from the two previous studies albeit with mild accumulation of neutral MC3 in the bilayer center. To demonstrate Membrane Builder's capability of building a realistic LNP patch, we generated KC2- or MC3-containing LNP membranes with high concentrations of cholesterol and ionizable cationic lipids together with 2 mol% PEG-lipids. We observe that PEG-chains are flexible, which can be more preferentially extended laterally in the presence of cationic lipids due to the attractive interactions between their head groups and PEG oxygen. The presence of PEG-lipids also relaxes the lateral packing in LNP membranes, and the area compressibility modulus (KA) of LNP membranes with cationic lipids fit into typical KA of fluid-phase membranes. Interestingly, the interactions between PEG oxygen and head group of ionizable cationic lipids induce a negative curvature. We hope that this LNP capability in Membrane Builder can be useful to better characterize various LNPs with or without genetic drugs for a rational LNP design.

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Soft Cationic Nanoparticles for Drug Delivery: Production and Cytotoxicity of Solid Lipid Nanoparticles (SLNs)

Applied Sciences ◽

10.3390/app9204438 ◽

2019 ◽

Vol 9 (20) ◽

pp. 4438 ◽

Cited By ~ 14

Author(s):

Amélia Silva ◽

Carlos Martins-Gomes ◽

Tiago Coutinho ◽

Joana Fangueiro ◽

Elena Sanchez-Lopez ◽

...

Keyword(s):

Drug Delivery ◽

Surface Properties ◽

Cell Lines ◽

Solid Lipid Nanoparticles ◽

Lipid Nanoparticles ◽

Cationic Lipids ◽

Solid Lipid ◽

Mcf 7

The surface properties of nanoparticles have decisive influence on their interaction with biological barriers (i.e., living cells), being the concentration and type of surfactant factors to have into account. As a result of different molecular structure, charge, and degree of lipophilicity, different surfactants may interact differently with the cell membrane exhibiting different degrees of cytotoxicity. In this work, the cytotoxicity of two cationic solid lipid nanoparticles (SLNs), differing in the cationic lipids used as surfactants CTAB (cetyltrimethylammonium bromide) or DDAB (dimethyldioctadecylammonium bromide), referred as CTAB-SLNs and DDAB-SLNs, respectively, was assessed against five different human cell lines (Caco-2, HepG2, MCF-7, SV-80, and Y-79). Results showed that the cationic lipids used in SLN production highly influenced the cytotoxic profile of the particles, with CTAB-SLNs being highly cytotoxic even at low concentrations (IC50 < 10 µg/mL, expressed as CTAB amount). DDAB-SLNs produced much lower cytotoxicity, even at longer exposure time (IC50 from 284.06 ± 17.01 µg/mL (SV-80) to 869.88 ± 62.45 µg/mL (MCF-7), at 48 h). To the best of our knowledge, this is the first report that compares the cytotoxic profile of CTAB-SLNs and DDAB-SLNs based on the concentration and time of exposure, using different cell lines. In conclusion, the choice of the right surfactant for biological applications influences the biocompatibility of the nanoparticles. Regardless the type of drug delivery system, not only the cytotoxicity of the drug-loaded nanoparticles should be assessed, but also the blank (non-loaded) nanoparticles as their surface properties play a decisive role both in vitro and in vivo.

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Tuning the Immunostimulation Properties of Cationic Lipid Nanocarriers for Nucleic Acid Delivery

Frontiers in Immunology ◽

10.3389/fimmu.2021.722411 ◽

2021 ◽

Vol 12 ◽

Author(s):

Arindam K. Dey ◽

Adrien Nougarède ◽

Flora Clément ◽

Carole Fournier ◽

Evelyne Jouvin-Marche ◽

...

Keyword(s):

Nucleic Acid ◽

Messenger Rna ◽

Cationic Lipid ◽

Lipid Nanoparticles ◽

Cationic Lipids ◽

Fine Tuning ◽

Target Cells ◽

Substantial Impact ◽

Lipid Nanocarriers ◽

The Impact

Nonviral systems, such as lipid nanoparticles, have emerged as reliable methods to enable nucleic acid intracellular delivery. The use of cationic lipids in various formulations of lipid nanoparticles enables the formation of complexes with nucleic acid cargo and facilitates their uptake by target cells. However, due to their small size and highly charged nature, these nanocarrier systems can interact in vivo with antigen-presenting cells (APCs), such as dendritic cells (DCs) and macrophages. As this might prove to be a safety concern for developing therapies based on lipid nanocarriers, we sought to understand how they could affect the physiology of APCs. In the present study, we investigate the cellular and metabolic response of primary macrophages or DCs exposed to the neutral or cationic variant of the same lipid nanoparticle formulation. We demonstrate that macrophages are the cells affected most significantly and that the cationic nanocarrier has a substantial impact on their physiology, depending on the positive surface charge. Our study provides a first model explaining the impact of charged lipid materials on immune cells and demonstrates that the primary adverse effects observed can be prevented by fine-tuning the load of nucleic acid cargo. Finally, we bring rationale to calibrate the nucleic acid load of cationic lipid nanocarriers depending on whether immunostimulation is desirable with the intended therapeutic application, for instance, gene delivery or messenger RNA vaccines.

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Tuning the immunostimulation properties of cationic lipid nanocarriers for nucleic acid delivery

10.1101/2021.06.16.448666 ◽

2021 ◽

Author(s):

Arindam K Dey ◽

Adrien Nougarede ◽

Flora Clement ◽

Carole Fournier ◽

Evelyne Jouvin-Marche ◽

...

Keyword(s):

Nucleic Acid ◽

Messenger Rna ◽

Cationic Lipid ◽

Lipid Nanoparticles ◽

Cationic Lipids ◽

Fine Tuning ◽

Target Cells ◽

Substantial Impact ◽

Lipid Nanocarriers ◽

The Impact

Nonviral systems, such as lipid nanoparticles, have emerged as reliable methods to enable nucleic acid intracellular delivery. The use of cationic lipids in various formulations of lipid nanoparticles enables the formation of complexes with nucleic acid cargo and facilitates their uptake by target cells. However, due to their small size and highly charged nature, these nanocarrier systems can interact in vivo with antigen-presenting cells (APCs), such as dendritic cells (DCs) and macrophages. As this might prove to be a safety concern for developing therapies based on lipid nanocarriers, we sought to understand how they could affect the physiology of APCs. In the present study, we investigate the cellular and metabolic response of primary macrophages or DCs exposed to the neutral or cationic variant of the same lipid nanoparticle formulation. We demonstrate that macrophages are the cells affected most significantly and that the cationic nanocarrier has a substantial impact on their physiology, depending on the positive surface charge. Our study provides a first model explaining the impact of charged lipid materials on immune cells and demonstrates that the primary adverse effects observed can be prevented by fine-tuning the load of nucleic acid cargo. Finally, we bring rationale to calibrate the nucleic acid load of cationic lipid nanocarriers depending on whether immunostimulation is desirable with the intended therapeutic application, for instance, gene delivery or messenger RNA vaccines.

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Sorafenib Repurposing for Ophthalmic Delivery by Lipid Nanoparticles: A Preliminary Study

Pharmaceutics ◽

10.3390/pharmaceutics13111956 ◽

2021 ◽

Vol 13 (11) ◽

pp. 1956

Author(s):

Angela Bonaccorso ◽

Veronica Pepe ◽

Cristina Zappulla ◽

Cinzia Cimino ◽

Angelo Pricoco ◽

...

Keyword(s):

Uveal Melanoma ◽

Physical Stability ◽

Malignant Tumour ◽

Lipid Nanoparticles ◽

Resistance Mechanisms ◽

Cationic Lipids ◽

Ophthalmic Delivery ◽

Preliminary Study ◽

Prolonged Drug Release ◽

Potential Use

Uveal melanoma is the second most common melanoma and the most common intraocular malignant tumour of the eye. Among various treatments currently studied, Sorafenib was also proposed as a promising drug, often administered with other compounds in order to avoid resistance mechanisms. Despite its promising cellular activities, the use of Sorafenib by oral administration is limited by its severe side effects and the difficulty to reach the target. The encapsulation into drug delivery systems represents an interesting strategy to overcome these limits. In this study, different lipid nanoparticulate formulations were prepared and compared in order to select the most suitable for the encapsulation of Sorafenib. In particular, two solid lipids (Softisan or Suppocire) at different concentrations were used to produce solid lipid nanoparticles, demonstrating that higher amounts were able to achieve smaller particle sizes, higher homogeneity, and longer physical stability. The selected formulations, which demonstrated to be biocompatible on Statens Seruminstitut Rabbit Cornea cells, were modified to improve their mucoadhesion, evaluating the effect of two monovalent cationic lipids with two lipophilic chains. Sorafenib encapsulation allowed obtaining a sustained and prolonged drug release, thus confirming the potential use of the developed strategy to topically administer Sorafenib in the treatment of uveal melanoma.

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Understanding structure-activity relationships of pH-sensitive cationic lipids facilitates the rational identification of promising lipid nanoparticles for delivering siRNAs in vivo

Journal of Controlled Release ◽

10.1016/j.jconrel.2019.01.001 ◽

2019 ◽

Vol 295 ◽

pp. 140-152 ◽

Cited By ~ 36

Author(s):

Yusuke Sato ◽

Kazuki Hashiba ◽

Kosuke Sasaki ◽

Masatoshi Maeki ◽

Manabu Tokeshi ◽

...

Keyword(s):

Lipid Nanoparticles ◽

Ph Sensitive ◽

Cationic Lipids ◽

Structure Activity Relationships ◽

Structure Activity

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Hydrophobic scaffolds of pH-sensitive cationic lipids contribute to miscibility with phospholipids and improve the efficiency of delivering short interfering RNA by small-sized lipid nanoparticles

Acta Biomaterialia ◽

10.1016/j.actbio.2019.11.022 ◽

2020 ◽

Vol 102 ◽

pp. 341-350 ◽

Cited By ~ 8

Author(s):

Yusuke Sato ◽

Nana Okabe ◽

Yusuke Note ◽

Kazuki Hashiba ◽

Masatoshi Maeki ◽

...

Keyword(s):

Lipid Nanoparticles ◽

Ph Sensitive ◽

Cationic Lipids ◽

Short Interfering Rna ◽

Interfering Rna

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Advances in the Formulation and Assembly of Non-Cationic Lipid Nanoparticles for the Medical Application of Gene Therapeutics

Nanomaterials ◽

10.3390/nano11030825 ◽

2021 ◽

Vol 11 (3) ◽

pp. 825

Author(s):

Richard K. Fisher ◽

Phillip C. West ◽

Samuel I. Mattern-Schain ◽

Michael D. Best ◽

Stacy S. Kirkpatrick ◽

...

Keyword(s):

Gene Therapy ◽

Nucleic Acid ◽

Cationic Lipid ◽

Medical Application ◽

Lipid Nanoparticles ◽

Cell Penetrating Peptides ◽

Cationic Lipids ◽

Injection Technique ◽

Ethanol Injection ◽

Assembly Technique

Lipid nanoparticles have become increasingly popular delivery platforms in the field of gene therapy, but bench-to-bedside success has been limited. Many liposomal gene vectors are comprised of synthetic cationic lipids, which are associated with lipid-induced cytotoxicity and immunogenicity. Natural, non-cationic PEGylated liposomes (PLPs) demonstrate favorable biocompatibility profiles but are not considered viable gene delivery vehicles due to inefficient nucleic acid loading and reduced cellular uptake. PLPs can be modified with cell-penetrating peptides (CPPs) to enhance the intracellular delivery of liposomal cargo but encapsulate leakage upon CPP-PLP assembly is problematic. Here, we aimed to identify parameters that overcome these performance barriers by incorporating nucleic acid condensers during CPP-PLP assembly and screening variable ethanol injection parameters for optimization. CPP-PLPs were formed with R8-amphiphiles via pre-insertion, post-insertion and post-conjugation techniques and liposomes were characterized for size, surface charge, homogeneity, siRNA encapsulation efficiency and retention and cell associative properties. Herein we demonstrate that pre-insertion of stearylated R8 into PLPs is an efficient method to produce non-cationic CPP-PLPs and we provide additional assembly parameter specifications for a modified ethanol injection technique that is optimized for siRNA encapsulation/retention and enhanced cell association. This assembly technique could provide improved clinical translation of liposomal based gene therapy applications.

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