Facile scalable one-flow synthesis of ionizable cationic lipid library as precursors of nanoparticle carriers.

A variety of ionizable and cationic lipids have been synthesized as precursors for nanoparticle carriers. However, the laborious synthetic routes in batch reactors often involve the use of toxic and carcinogenic agents, as well as challenge of removing gaseous byproducts. In this study, we present facile one-flow micro-reaction process that enables the synthesis of 11 ionizable lipids as well as 7 cationic lipids, including the well-known DODAP and DOTAP. These lipids can be scaled up to produce approximately ∼10g/h by using a straightforward size-up approach. The development of the lipid library was involved generating highly moisture-sensitive acyl chloride at 25 °C for 1.5 min. The toxic byproducts such as HCl, CO2 and CO were subsequently removed using a liquid-gas separator. The esterification with dimethylamino-1,2-diol at 25 °C for 3 min, monitored in-line with FTIR, completed the process. Additionally, the synthesized ionizable lipids were converted to cationic lipids with methyl sulfate, chloride ions via dimethyl sulfate and Steglich esterification in a continuous flow system. Finally, the produced DODAP was transformed into a uniform-sized LNPs (64 nm, PDI 0.07) and liposomal nanoparticles (72 nm, PDI 0.05) while DOTAP was converted to liposomes (55 nm, PDI 0.08) using a custom micro-mixer. This efficient platform for lipid synthesis significantly contributes to the practical applications of lipid-based nanomedicines.

Construction of Recombinant Baculoviruses.

The success of using the insect cell-baculovirus expression technology (BEST) relies on the efficient construction of recombinant baculovirus with genetic stability and high productivity, ideally within a short time period. Generation of recombinant baculoviruses requires the transfection of insect cells, harvesting of recombinant baculovirus pools, isolation of plaques, and the expansion of baculovirus stocks for their use for recombinant protein production. Moreover, many options exist for selecting the genetic elements to be present in the recombinant baculovirus. This chapter describes the most commonly used homologous recombination systems for the production of recombinant baculoviruses, as well as strategies to maximize generation efficiency and recombinant protein or baculovirus production. The key steps for generating baculovirus stocks and troubleshooting strategies are described.

One-Component Cationic Lipids for Systemic mRNA Delivery to Splenic T Cells.

Unlocking the full potential of mRNA immunotherapy necessitates targeted delivery to specific cell subsets in the spleen. Four-component lipid nanoparticles (LNPs) utilized in numerous clinical trials are primarily limited in hepatocyte and muscular targeting, highlighting the imperative demand for targeted and simplified non-liver mRNA delivery systems. Herein, we report the rational design of one-component ionizable cationic lipids to selectively deliver mRNA to the spleen and T cells with high efficacy. Unlike the tertiary amine-based ionizable lipids involved in LNPs, the proposed cationic lipids rich in secondary amines can efficiently deliver mRNA both in vitro and in vivo as the standalone carriers. Furthermore, these vectors facilitate efficacious mRNA delivery to the T cell subsets following intravenous administration, demonstrating substantial potential for advancing immunotherapy applications. This straightforward strategy extends the utility of lipid family for extrahepatic mRNA delivery, offering new insights into vector development beyond LNPs to further the field of precise mRNA therapy.

Cationic nanocarriers: A potential approach for targeting negatively charged cancer cell.

Cancer, a widespread and lethal disease, necessitates precise therapeutic interventions to mitigate its devastating impact. While conventional chemotherapy remains a cornerstone of cancer treatment, its lack of specificity towards cancer cells results in collateral damage to healthy tissues, leading to adverse effects. Thus, the quest for targeted strategies has emerged as a critical focus in cancer research. This review explores the development of innovative targeting methods utilizing novel drug delivery systems tailored to recognize and effectively engage cancer cells. Cancer cells exhibit morphological and metabolic traits, including irregular morphology, unchecked proliferation, metabolic shifts, genetic instability, and a higher negative charge, which serve as effective targeting cues. Central to these strategies is the exploitation of the unique negative charge characteristic of cancer cells, attributed to alterations in phospholipid composition and the Warburg effect. Leveraging this distinct feature, researchers have devised cationic carrier systems capable of enhancing the specificity of therapeutic agents towards cancer cells. The review delineates the underlying causes of the negative charge in cancer cells and elucidates various targeting approaches employing cationic compounds for drug delivery systems. Furthermore, it delves into the methods employed for the preparation of these systems. Beyond cancer treatment, the review also underscores the multifaceted applications of cationic carrier systems, encompassing protein and peptide delivery, imaging, photodynamic therapy, gene delivery, and antimicrobial applications. This comprehensive exploration underscores the potential of cationic carrier systems as versatile tools in the fight against cancer and beyond.

Determination of the interior pH of lipid nanoparticles using a pH-sensitive fluorescent dye-based DNA probe.

Lipid nanoparticles (LNPs) containing ionizable cationic lipids are proven delivery systems for therapeutic nucleic acids, such as small interfering RNA (siRNA). It is important to understand the relationship between the interior pH of LNPs and the pH of the external environment to understand LNP formulation and function. Here, we developed a simple and rapid approach for determining the pH of the LNP core using a pH-sensitive fluorescent dye-based DNA probe. LNP siRNA systems containing pH-responsive DNA probes (LNP-siRNA&DNA) were generated by rapid mixing of lipids in ethanol and pH 4 aqueous buffer containing siRNA and DNA probes. We demonstrated that DNA probes were readily encapsulated in LNP systems and were sequestered into an environment at a high concentration as evidenced by an inter-probe FRET signal. It was shown that the pH of LNP encapsulated probes closely follows the pH increase or decrease of the external environment. This indicates that the clinically approved LNP RNA systems with similar lipid compositions (e.g., Onpattro and Comirnaty) are highly permeable to protons and that the pH of the interior environment closely mirrors the external environment. The pH-dependent response of the probe in LNPs was also confirmed under buffer conditions at various pHs. Furthermore, we showed that the pH-sensitive DNA probe can be incorporated into LNP systems at levels that allow the pH response to be monitored at a single LNP level using convex lens-induced confinement (CLiC) confocal microscopy. Direct visualization of the internal pH of single particles with the fluorescent DNA probe was achieved by CLiC for LNP-siRNA&DNA systems formulated under both high and normal ionic strength conditions.

Cationic lipids from multi-component Passerini reaction for non-viral gene delivery: A structure-activity relationship study.

Although many types of cationic lipids have been developed as efficient gene vectors, the construction of lipid molecules with simple procedures remains challenging. Passerini reaction, as a classic multicomponent reaction, could directly give the α-acyloxycarboxamide products with biodegradable ester and amide bonds. Herein, two series of novel cationic lipids with heterocyclic pyrrolidine and piperidine as headgroups were synthesized through Passerini reaction (P-series) and amide condensation (A-series), and relevant structure-activity relationships on their gene delivery capability was studied. It was found that although both of the two series of lipids could form lipid nanoparticles (LNPs) which could effectively condense DNA, the LNP derived from P-series lipids showed higher transfection efficiency, serum tolerance, cellular uptake, and lower cytotoxicity. Unlike the A-series LNPs, the P-series LNPs showed quite different structure-activity relationship, in which the relative site of the secondary amine had significant effect on the transfection performance. The othro-isomers of the P-series lipids had lower cytotoxicity, but poor transfection efficiency, which was probably due to their unstable nature. Taken together, this study not only validated the feasibility of Passerini reaction for the construction of cationic lipids for gene delivery, but also afforded some clues for the rational design of effective non-viral lipidic gene vectors.

A systematic study of the effect of lipid architecture on cytotoxicity and cellular uptake of cationic cubosomes.

HYPOTHESIS: Lipid nanoparticles containing a cationic lipid are increasingly used in drug and gene delivery as they can display improved cellular uptake, enhanced loading for anionic cargo such as siRNA and mRNA or exhibit additional functionality such as cytotoxicity against cancer cells. This research study tests the hypothesis that the molecular structure of the cationic lipid influences the structure of the lipid nanoparticle, the cellular uptake, and the resultant cytotoxicity. EXPERIMENTS: Three potentially cytotoxic cationic lipids, with systematic variations to the hydrophobic moiety, were designed and synthesised. All the three cationic lipids synthesised contain pharmacophores such as the bicyclic coumarin group (CCA12), the tricyclic etodolac moiety (ETD12), or the large pentacyclic triterpenoid "ursolic" group (U12) conjugated to a quaternary ammonium cationic lipid containing twin C12 chains. The cationic lipids were doped into monoolein cubosomes at a range of concentrations from 0.1 mol% to 5 mol% and the effect of the lipid molecular architecture on the cubosome phase behaviour was assessed using a combination of Small Angle X-Ray Scattering (SAXS), Dynamic Light Scattering (DLS), zeta-potential and cryo-Transmission Electron Microscopy (Cryo-TEM). The resulting cytotoxicity of these particles against a range of cancerous and non-cancerous cell-lines was assessed, along with their cellular uptake. FINDINGS: The molecular architecture of the cationic lipid was linked to the internal nanostructure of the resulting cationic cubosomes with a transition to more curved cubic and hexagonal phases generally observed. Cubosomes formed from the cationic lipid CCA12 were found to have improved cellular uptake and significantly higher cytotoxicity than the cationic lipids ETD12 and U12 against the gastric cancer cell-line (AGS) at lipid concentrations ≥ 75 µg/mL. CCA12 cationic cubosomes also displayed reasonable cytotoxicity against the prostate cancer PC-3 cell-line at lipid concentrations ≥ 100 µg/mL. In contrast, 2.5 mol% ETD12 and 2.5 mol% U12 cubosomes were generally non-toxic against both cancerous and non-cancerous cell lines over the entire concentration range tested. The molecular architecture of the cationic lipid was found to influence the cubosome phase behaviour, the cellular uptake and the toxicity although further studies are necessary to determine the exact relationship between structure and cellular uptake across a range of cell lines.

Histidine-based ionizable cationic surfactants: novel biodegradable agents for hydrophilic macromolecular drug delivery.

The aim of this study was to design surfactants based on histidine (His) for hydrophobic ion-pairing and evaluate their safety and efficacy. Lauryl, palmitoyl and oleyl alcohol, as well as 2-hexyl-1-decanol were converted into surfactants with histidine as head-group via esterification. The synthesized His-surfactants were characterized regarding pKa, critical micellar concentration (CMC), biodegradability, toxicity on Caco-2 cells, and ability to provide endosomal escape. Furthermore, the suitability of these agents to be employed as counterions in hydrophobic ion pairing was evaluated. Chemical structures were confirmed by 1H-NMR, FT-IR, and MS. The synthesized surfactants showed pKa values ranging from 4.9 to 6.0 and CMC values in the range of 0.3 to 7.0 mM. Their biodegradability was proven by enzymatic cleavage within 24 h. Below the CMC, His-surfactants did not show cytotoxic effects on Caco-2 cells (cell viability > 80%). All His-surfactants showed the ability to provide endosomal escape in a pH-dependent manner in the range of 5.2 to 6.8. Complexes formed between His-surfactants and heparin or plasmid DNA (pDNA) via hydrophobic ion pairing showed at least 100-fold higher lipophilicity than the correspondent model drugs. According to these results, His-surfactants might be a promising safe tool for delivering hydrophilic macromolecular drugs and nucleic acids.

Synthesis of Amino Acid-Based Cationic Lipids and Study of the Role of the Cationic Head Group for Enhanced Drug and Nucleic Acid Delivery.

Leveraging liposomes for drug and nucleic acid delivery, though promising due to reduced toxicity and ease of preparation, faces challenges in stability and efficiency. To address this, we synthesized cationic amphiphiles from amino acids (arginine, lysine, and histidine). Histidine emerged as the superior candidate, leading to the development of three histidine-rich cationic amphiphiles for liposomes. Using the hydration method, we have prepared the liposomes and determined the optimal N/P ratios for lipoplex formation via gel electrophoresis. In vitro transfection assays compared the efficacy of our lipids to Fugene, while MTT assays gauged biocompatibility across cancer cell lines (MDA-MB 231 and MCF-7). The histidine-based lipid demonstrated marked potential in enhancing drug and nucleic acid delivery. This improvement stemmed from increased zeta potential, enhancing electrostatic interactions with nucleic acids and cellular uptake. Our findings underscore histidine's crucial role over lysine and arginine for effective delivery, revealing a significant correlation between histidine abundance and optimal performance. This study paves the way for histidine-enriched lipids as promising candidates for efficient drug and nucleic acid delivery, addressing key challenges in the field.

Study of the correlation between the structure of selected triester of phosphatidylcholine and their impact on physicochemical properties of model mammalian membranes.

Cationic lipids are synthetic compounds of amphiphilic character used in Drug Delivery Systems (DDS), especially in gene therapy, as the carriers of genetic material. As it is known, the main limitation of the application of cationic lipids in DDS is their high cytotoxicity after in vivo administration and low bioactivity. This is probably related to not fully known the relationship between the lipid structure and its activity as well as the mechanism of lipofection or drug delivery. Therefore, in this work we determined the impact of a selected group of cationic lipids - triesters of phosphatidylcholine (Et-PCs) - differing in their hydrophobic structure on model mammalian membranes. In the research, as model systems, Langmuir monolayers and liposomes were applied. It was shown that the incorporation of Et-PCs into model mammalian membranes weakens interactions between lipids, causing the increase of fluidity, disordering degree and permeability of membrane. The destabilization of the membrane in this way can facilitate the entry of drugs, carried inside cationic liposomes, into the pathological cell. Moreover, the studies prove that the structure of the hydrophobic part of cationic lipids also affects the properties of lipid membranes.

On Fusogenicity of Positively Charged Phased-Separated Lipid Vesicles: Experiments and Computational Simulations.

This paper studies the fusogenicity of cationic liposomes in relation to their surface distribution of cationic lipids and utilizes membrane phase separation to control this surface distribution. It is found that concentrating the cationic lipids into small surface patches on liposomes, through phase-separation, can enhance liposome's fusogenicity. Further concentrating these lipids into smaller patches on the surface of liposomes led to an increased level of fusogenicity. These experimental findings are supported by numerical simulations using a mathematical model for phase-separated charged liposomes. Findings of this study may be used for design and development of highly fusogenic liposomes with minimal level of toxicity.

Inner leaflet cationic lipid increases nucleic acid loading independently of outer leaflet lipid charge in asymmetric liposomes.

Use of cationic lipid vesicles (liposomes) can yield large amounts of nucleic acid entrapped inside the vesicles and/or bound to the external surface of the vesicles. To show a method to prepare asymmetric lipid vesicles (liposomes) with high amounts of entrapped nucleic acid is possible, symmetric and asymmetric lipid vesicles composed of mixtures of neutral (zwitterionic), anionic, and/or cationic phospholipids were formed in the presence of oligo DNA. For symmetric large unilamellar vesicles nucleic acid association with vesicles was roughly 100 times greater for vesicles with a net cationic charge than for vesicles having a net neutral or anionic net charge. A high degree of association between nucleic acid and lipid was also achieved using asymmetric large unilamellar vesicles with a net cationic charge in their inner leaflet, even when they had an anionic charge in their outer leaflet. In contrast, asymmetric vesicles in which only the outer leaflet had a net cationic charge had only low amounts of vesicle-associated nucleic acid, similar in amount to the amount of nucleic acid associated with asymmetric vesicles with an outer leaflet having a net anionic charge. These results indicate that in asymmetric vesicles with cationic lipid enriched inner leaflets nucleic acid is largely entrapped inside the vesicle lumen rather than bound to their external surface, and that asymmetric vesicles can be used to trap high amounts of nucleic acid even when using a lipid composition in the outer leaflet of a lipid vesicle that does not associate with nucleic acids. Such asymmetrically charged vesicles should have applications in studies of membrane protein-nucleic acid interactions as well as in studies of how membrane charge asymmetry can influence membrane protein structure, orientation, and function.

Evaluation of lipophosphoramidates-based amphiphilic compounds on the formation of biofilms of marine bacteria.

The bacteriostatic and/or bactericidal properties of few phosphoramide-based amphiphilic compounds on human pathogenic bacteria were previously reported. In this study, the potential of two cationic (BSV36 and KLN47) and two zwitterionic (3 and 4) amphiphiles as inhibitors of marine bacterial growth and biofilm formation were investigated. Results showed that the four compounds have little impact on the growth of a panel of 18 selected marine bacteria at a concentration of 200 µM, and up to 700 µM for some bacterial strains. Interestingly, cationic lipid BSV36 and zwitterionic lipids 3 and 4 effectively disrupt biofilm formation of Paracoccus sp. 4M6 and Vibrio sp. D02 at 200 µM and to a lesser extent of seven other bacterial strains tested. Moreover, ecotoxicological assays on four species of microalgae highlighted that compounds 3 and 4 have little impact on microalgae growth with EC50 values of 51 µM for the more sensitive species and up to 200 µM for most of the others. Amphiphilic compounds, especially zwitterionic amphiphiles 3 and 4 seem to be promising candidates against biofilm formation by marine bacteria.

Novel bioactive cationic cubosomes enhance the cytotoxic effect of paclitaxel against a paclitaxel resistant prostate cancer cell-line.

Hypothesis The study aimed to use molecular hybridization of a cationic lipid with a known pharmacophore to produce a bifunctional lipid having a cationic charge to enhance fusion with the cancer cell surface and biological activity via the pharmacophoric head group. Experiments The novel cationic lipid DMP12 [N-(2-(3-(3,4-dimethoxyphenyl) propanamido) ethyl)-N-dodecyl-N-methyldodecan-1-aminium iodide] was synthesised by conjugating 3-(3,4-dimethoxyphenyl) propanoic acid (or 3,4-dimethoxyhydrocinnamic acid) to twin 12 carbon chains bearing a quaternary ammonium group [N-(2-aminoethyl)-N-dodecyl-N-methyldodecan-1-aminium iodide]. The physicochemical and biological properties of DMP12 were investigated. Cubosome particles consisting of monoolein (MO) doped with DMP12 and paclitaxel were characterized using Small-angle X-ray Scattering (SAXS), Dynamic Light Scattering (DLS), and Cryo-Transmission Electron Microscopy (Cryo-TEM). Combination therapy using these cubosomes was assessed in vitro against the gastric (AGS) and prostate (DU-145 and PC-3) cancer cell lines using cytotoxicity assay. Findings Monoolein (MO) cubosomes doped with DMP12 were observed to be toxic against the AGS and DU-145 cell-lines at higher cubosome concentrations (≥100 µg/ml) but had limited activity against the PC-3 cell-line. However, combination therapy consisting of 5 mol% DMP12 and 0.5 mol% paclitaxel (PTX) significantly increased the cytotoxicity against the PC-3 cell-line which was resistant to either DMP12 or PTX individually. The results demonstrate that DMP12 has a prospective role as a bioactive excipient in cancer therapy.

Design of Cationic Lipids with Acetal Linkers: Conformational Preferences, Hydrolytic Stability, and High Drug-Loading Abilities.

Lipids are key constituents of numerous biomedical drug delivery technologies. Here, we present the design, synthesis and biophysical characterizations of a library of cationic lipids containing an acetal residue in their linker region. These cationic acetal lipids (CALs) were conveniently prepared through a trans-acetalization protocol from commercially available precursors. NMR studies highlighted the conformational rigidity at the acetal residue and the high hydrolytic stability of these CALs. Fluorescence anisotropy studies revealed that the CAL with a pyridinium headgroup (CAL1) formed highly cohesive vesicular aggregates in water. These structural and self-assembly features of the CAL1 allowed up to 196 % w/w loading of curcumin (Cur) as a representative hydrophobic drug. A reconstitutable formulation of Cur was obtained as a result, which could deliver the drug inside mammalian cells with very high efficiency. The hemocompatibility and cytocompatibility of CAL1 was significantly enhanced by creating a coating of polydopamine (PDA) onto its vesicular assemblies to produce hybrid lipid-polymer nanocapsules. This work demonstrates rapid access to the useful synthetic lipid formulations with high potential in drug and gene delivery applications.

In vitro and in vivo evaluation of clinically-approved ionizable cationic lipids shows divergent results between mRNA transfection and vaccine efficacy.

Ionizable cationic lipids (ICLs) play an essential role in the effectiveness of lipid nanoparticles (LNPs) for delivery of mRNA therapeutics and vaccines; therefore, critical evaluations of their biological performance would extend the existing knowledge in the field. In the present study, we examined the effects of the three clinically-approved ICLs, Dlin-MC3-DMA, ALC-0315 and SM-102, as well as DODAP, on the in vitro and in vivo performance of LNPs for mRNA delivery and vaccine efficacy. mRNA-LNPs containing these lipids were successfully prepared, which were all found to be very similar in their physicochemical properties and mRNA encapsulation efficiencies. Furthermore, the results of the in vitro studies indicated that these mRNA-LNPs were efficiently taken up by immortalized and primary immune cells with comparable efficiency; however, SM-102-based LNPs were superior in inducing protein expression and antigen-specific T cell proliferation. In contrast, in vivo studies revealed that LNPs containing ALC-0315 and SM-102 yielded almost identical protein expression levels in zebrafish embryos, which were significantly higher than Dlin-MC3-DMA-based LNPs. Additionally, a mouse immunization study demonstrated that a single-dose subcutaneous administration of the mRNA-LNPs resulted in a high production of intracellular cytokines by antigen-specific T cells, but no significant differences among the three clinically-approved ICLs were observed, suggesting a weak correlation between in vitro and in vivo outcomes. This study provides strong evidence that ICLs modulate the performance of mRNA-LNPs and that in vitro data does not adequately predict their behavior in vivo.

Synthesis and Characterization of a New Cationic Lipid: Efficient siRNA Delivery and Anticancer Activity of Survivin-siRNA Lipoplexes for the Treatment of Lung and Breast Cancers.

Survivin has been shown to be widely expressed in most tumor cells, including lung and breast cancers. Due to limited siRNA delivery, it is more challenging to target survivin using knockdown-based techniques. Designing and developing new, bifunctional chemical molecules with both selective anti-proliferative activity and effective siRNA transfection capabilities by targeting a particular gene is important to treat aggressive tumors like triple-negative breast tumors (TNBC). The cationic lipids deliver small interfering RNA (siRNA) and also display inherent anti-cancer activities; therefore, cationic lipid therapies have become very popular for treating malignant cancers. In the current study, we attempted to synthesize a series of acid-containing cationic lipids, anthranilic acid-containing mef lipids, and indoleacetic acid-containing etodo lipids etc. Further, we elucidated their bi-functional activity for their anticancer activity and survivin siRNA-mediated anti-cancer activity. Our results showed that lipoplexes with siRNA-Etodo: Dotap (ED) and siRNA-Mef: Dotap (MD) exhibited homogeneous particle size and positive zeta potential. Further, biological investigations resulted in enhanced survivin siRNA delivery with high stability, improved transfection efficiency, and anti-cancer activity. Additionally, our findings showed that survivin siRNA lipoplexes (ED and MD) in A549 cells and 4T1 cells exhibited stronger survivin knockdown, enhanced apoptosis, and G1 or G2/M phase arrest in both cell types. In vivo results revealed that treatment with survivin complexed lipoplexes significantly reduced tumor growth and tumor weight compared to control. Thus, our novel quaternary amine-based liposome formulations are predicted to open up new possibilities in the development of a simple and widely utilized platform for siRNA delivery and anti-cancer activities.

Recent advances and prospects for lipid-based nanoparticles as drug carriers in the treatment of human retinal diseases.

The delivery of cures for retinal diseases remains problematic. There are four main challenges: passing through multiple barriers of the eye, the delivery to particular retinal cell types, the capability to carry different forms of therapeutic cargo and long-term therapeutic efficacy. Lipid-based nanoparticles (LBNPs) are potent to overcome these challenges due to their unique merits: amphiphilic nanoarchitectures to pass biological barriers, vary modifications with specific affinity to target cell types, flexible capacity for large and mixed types of cargos and slow-release formulations for long-term treatment. We have reviewed the latest research on the applications of LBNPs for treating retinal diseases and categorized them by different payloads. Furthermore, we identified technical barriers and discussed possible future development for LBNPs to expand the therapeutic potential in treating retinal diseases.

Versatility of Liposomes for Antisense Oligonucleotide Delivery: A Special Focus on Various Therapeutic Areas.

Nucleic acid therapeutics, specifically antisense oligonucleotides (ASOs), can effectively modulate gene expression and protein function, leading to long-lasting curative effects. The hydrophilic nature and large size of oligonucleotides present translational challenges, which have led to the exploration of various chemical modifications and delivery systems. The present review provides insights into the potential role of liposomes as a drug delivery system for ASOs. The potential benefits of liposomes as an ASO carrier, along with their method of preparation, characterization, routes of administration, and stability aspects, have been thoroughly discussed. A novel perspective in terms of therapeutic applications of liposomal ASO delivery in several diseases such as cancer, respiratory disease, ophthalmic delivery, infectious diseases, gastrointestinal disease, neuronal disorders, hematological malignancies, myotonic dystrophy, and neuronal disorders remains the major highlights of this review.

Multicomponent Lipid Nanoparticles for RNA Transfection.

Despite the wide variety of available cationic lipid platforms for the delivery of nucleic acids into cells, the optimization of their composition has not lost its relevance. The purpose of this work was to develop multi-component cationic lipid nanoparticles (LNPs) with or without a hydrophobic core from natural lipids in order to evaluate the efficiency of LNPs with the widely used cationic lipoid DOTAP (1,2-dioleoyloxy-3-[trimethylammonium]-propane) and the previously unstudied oleoylcholine (Ol-Ch), as well as the ability of LNPs containing GM3 gangliosides to transfect cells with mRNA and siRNA. LNPs containing cationic lipids, phospholipids and cholesterol, and surfactants were prepared according to a three-stage procedure. The average size of the resulting LNPs was 176 nm (PDI 0.18). LNPs with DOTAP mesylate were more effective than those with Ol-Ch. Core LNPs demonstrated low transfection activity compared with bilayer LNPs. The type of phospholipid in LNPs was significant for the transfection of MDA-MB-231 and SW 620 cancer cells but not HEK 293T cells. LNPs with GM3 gangliosides were the most efficient for the delivery of mRNA to MDA-MB-231 cells and siRNA to SW620 cells. Thus, we developed a new lipid platform for the efficient delivery of RNA of various sizes to mammalian cells.