Difference in Endocytosis Pathways Used by Differentiated Versus Nondifferentiated Epithelial Caco-2 Cells to Internalize Nanosized Particles.

Understanding the internalization of nanosized particles by mucosal epithelial cells is essential in a number of areas including viral entry at mucosal surfaces, nanoplastic pollution, as well as design and development of nanotechnology-type medicines. Here, we report our comparative study on pathways of cellular internalization in epithelial Caco-2 cells cultured in vitro as either a polarized, differentiated cell layer or as nonpolarized, nondifferentiated cells. The study reveals a number of differences in the extent that endocytic processes are used by cells, depending on their differentiation status and the nature of applied nanoparticles. In polarized cells, actin-driven and dynamin-independent macropinocytosis plays a prominent role in the internalization of both positively and negatively charged nanoparticles, contrary to its modest contribution in nonpolarized cells. Clathrin-mediated cellular entry plays a prominent role in the endocytosis of positive nanoparticles and cholesterol inhibition in negative nanoparticles. However, in nonpolarized cells, dynamin-dependent endocytosis is a major pathway in the internalization of both positive and negative nanoparticles. Cholesterol depletion affects both nonpolarized and polarized cells' internalization of positive and negative nanoparticles, which, in addition to the effect of cholesterol-binding inhibitors on the internalization of negative nanoparticles, indicates the importance of membrane cholesterol in endocytosis. The data collectively provide a new contribution to understanding endocytic pathways in epithelial cells, particularly pointing to the importance of the cell differentiation stage and the nature of the cargo.

Macrophage Membrane-Encapsulated Dopamine-Modified Poly Cyclodextrin Multifunctional Biomimetic Nanoparticles for Atherosclerosis Therapy.

Atherosclerotic plaques exhibit high cholesterol deposition and oxidative stress resulting from high reactive oxygen species (ROS). These are the major components in plaques and the main pro-inflammatory factor. Therefore, it is crucial to develop an effective therapeutic strategy that can simultaneously address the multiple pro-inflammatory factors via removing cholesterol and inhibiting the overaccumulated ROS. In this study, we constructed macrophage membrane-encapsulated biomimetic nanoparticles (MM@DA-pCD@MTX), which not only alleviate cholesterol deposition at the plaque lesion via reverse cholesterol transport but also scavenge the overaccumulated ROS. ß-Cyclodextrin (ß-CD) and the loaded methotrexate (MTX) act synergistically to induce cholesterol efflux for inhibiting the formation of foam cells. Among them, MTX up-regulated the expression of ABCA1, CYP27A1, and SR-B1. ß-CD increased the solubility of cholesterol crystals. In addition, the ROS scavenging property of dopamine (DA) was perfectly preserved in MM@DA-pCD@MTX, which could scavenge the overaccumulated ROS to alleviate the oxidative stress at the plaque lesion. Last but not least, MM-functionalized "homing" targeting of atherosclerotic plaques not only enables the targeted drug delivery but also prolongs in vivo circulation time and drug half-life. In summary, MM@DA-pCD@MTX emerges as a potent, multifunctional therapeutic platform for AS treatment, offering a high degree of biosafety and efficacy in addressing the complex pathophysiology of atherosclerosis.

Unnatural Peptide Assemblies Rapidly Deplete Cholesterol and Potently Inhibit Cancer Cells.

Cholesterol-rich membranes play a pivotal role in cancer initiation and progression, necessitating innovative approaches to target these membranes for cancer inhibition. Here we report the first case of unnatural peptide (1) assemblies capable of depleting cholesterol and inhibiting cancer cells. Peptide 1 self-assembles into micelles and is rapidly taken up by cancer cells, especially when combined with an acute cholesterol-depleting agent (MßCD). Click chemistry has confirmed that 1 depletes cell membrane cholesterol. It localizes in membrane-rich organelles, including the endoplasmic reticulum, Golgi apparatus, and lysosomes. Furthermore, 1 potently inhibits malignant cancer cells, working synergistically with cholesterol-lowering agents. Control experiments have confirmed that C-terminal capping and unnatural amino acid residues (i.e., BiP) are essential for both cholesterol depletion and potent cancer cell inhibition. This work highlights unnatural peptide assemblies as a promising platform for targeting the cell membrane in controlling cell fates.

Cholesterol-Modified Anti-Il6 siRNA Reduces the Severity of Acute Lung Injury in Mice.

Small interfering RNA (siRNA) holds significant therapeutic potential by silencing target genes through RNA interference. Current clinical applications of siRNA have been primarily limited to liver diseases, while achievements in delivery methods are expanding their applications to various organs, including the lungs. Cholesterol-conjugated siRNA emerges as a promising delivery approach due to its low toxicity and high efficiency. This study focuses on developing a cholesterol-conjugated anti-Il6 siRNA and the evaluation of its potency for the potential treatment of inflammatory diseases using the example of acute lung injury (ALI). The biological activities of different Il6-targeted siRNAs containing chemical modifications were evaluated in J774 cells in vitro. The lead cholesterol-conjugated anti-Il6 siRNA after intranasal instillation demonstrated dose-dependent therapeutic effects in a mouse model of ALI induced by lipopolysaccharide (LPS). The treatment significantly reduced Il6 mRNA levels, inflammatory cell infiltration, and the severity of lung inflammation. IL6 silencing by cholesterol-conjugated siRNA proves to be a promising strategy for treating inflammatory diseases, with potential applications beyond the lungs.

Superior Drug Delivery Performance of Multifunctional Bilosomes: Innovative Strategy to Kill Skin Cancer Cells for Nanomedicine Application.

Purpose: Numerous failures in melanoma treatment as a highly aggressive form of skin cancer with an unfavorable prognosis and excessive resistance to conventional therapies are prompting an urgent search for more effective therapeutic tools. Consequently, to increase the treatment efficiency and to reduce the side effects of traditional administration ways, herein, it has become crucial to combine photodynamic therapy as a promising therapeutic approach with the selectivity and biocompatibility of a novel colloidal transdermal nanoplatform for effective delivery of hybrid cargo with synergistic effects on melanoma cells. Methods: The self-assembled bilosomes, co-stabilized with L-α-phosphatidylcholine, sodium cholate, Pluronic® P123, and cholesterol, were designated, and the stability of colloidal vesicles was studied using dynamic and electrophoretic light scattering, also provided in cell culture medium (Dulbecco's Modified Eagle's Medium). The hybrid compounds - a classical photosensitizer (Methylene Blue) along with a complementary natural polyphenolic agent (curcumin), were successfully co-loaded, as confirmed by UV-Vis, ATR-FTIR, and fluorescent spectroscopies. The biocompatibility and usefulness of the polymer functionalized bilosome with loaded double cargo were demonstrated in vitro cyto- and phototoxicity experiments using normal keratinocytes and melanoma cancer cells. Results: The in vitro bioimaging and immunofluorescence study upon human skin epithelial (A375) and malignant (Me45) melanoma cell lines established the protective effect of the PEGylated bilosome surface. This effect was confirmed in cytotoxicity experiments, also determined on human cutaneous (HaCaT) keratinocytes. The flow cytometry experiments indicated the enhanced uptake of the encapsulated hybrid cargo compared to the non-loaded MB and CUR molecules, as well as a selectivity of the obtained nanocarriers upon tumor cell lines. The phyto-photodynamic action provided 24h-post irradiation revealed a more significant influence of the nanoplatform on Me45 cells in contrast to the A375 cell line, causing the cell viability rate below 20% of the control. Conclusion: As a result, we established an innovative and effective strategy for potential metastatic melanoma treatment through the synergism of phyto-photodynamic therapy and novel bilosomal-origin nanophotosensitizers.

Relative biomembrane fusogenicities of the tumor-selective liposomes of RGDK- and CGKRK-lipopeptides.

Cancer is the second leading cause of death globally after heart diseases. Currently used highly cytotoxic anti-cancer drugs not only kill cancer cells but also often kill non-cancerous healthy body cells, causing adverse side effects. Efforts are now being directed towards developing tumor-selective chemotherapy. Tumor/tumor endothelial cell selective peptide ligands are being covalently grafted onto the exo-surfaces of drug carriers such as liposomes, polymers, etc. A number of prior studies used conjugation of tumor/tumor endothelial cell-selective RGDK- or CGKRK-peptide ligands on the outer surfaces of liposomes, metal-based nanoparticles, single walled carbon nanotubes (SWNTs), etc. However, studies aimed at examining the relative cell membrane fusogenicities and the relative degrees of cellular uptake for the RGDK- and CGKRK-ligand-grafted nanometric drug carriers have not yet been undertaken. Herein, using the widely used liposomes of DOPC, DOPE, DOPS and cholesterol (45 : 25 : 20 : 15, w/w ratio) as the model biomembranes and the fluorescence resonance energy transfer (FRET) assay for measuring membrane fusogenicities, we show that the liposomes of the RGDK-lipopeptide are more biomembrane fusogenic than the liposomes of the CGKRK-lipopeptide. Notably, such FRET assay-derived relative biomembrane fusogenicities of the liposomes of RGDK- and CGKRK-lipopeptides were found to be consistent with their relative degrees of cellular uptake in cultured cancer cells. The present findings open the door for undertaking in-depth in vivo studies aimed at evaluating the relative therapeutic potential of different nanocarriers of drugs/genes/siRNA having tumor-targeting RGDK- and CGKRK-peptides on their exo-surfaces.

Comparative Study of Gas and Liquid Chromatography Methods for the Determination of Underivatised Neutral and Acidic Cannabinoids and Cholesterol.

The aim of our study was to develop a gas chromatographic method coupled with mass spectrometry (GC-MS) for the determination of underivatised neutral (CBDs-N) and acidic (CBDs-A) cannabinoids (CBDs) and cholesterol (Chol). Emphasis was also placed on comparing our original GC-MS method with the currently developed C18-high-performance liquid chromatography with photodiode detection (C18-HPLC-DAD). A combination of a long GC column, shallow temperature column programme, and mass-spectrometry was employed to avoid issues arising from the overlap between CBDs and Chol and background fluctuations. The pre-column procedure for CBDs and Chol in egg yolks consisted of hexane extractions, whereas the pre-column procedure for CBDs in non-animal samples involved methanol and hexane extractions. CBDs-A underwent decarboxylation to CBDs during GC-MS analyses, and pre-column extraction of the processed sample with NaOH solution allowed for CBD-A removal. No losses of CBDs-N were observed in the samples extracted with NaOH solution. GC-MS analyses of the samples before and after extraction with NaOH solution enabled the quantification of CBDs-A and CBDs-N. CBDs-A did not undergo decarboxylation to CBDs-N during C18-HPLC-DAD runs. The use of the C18-HPLC-DAD method allowed simultaneous determination of CBDs-N and CBDs-A. In comparison to the C18-HPLC-DAD method, our GC-MS technique offered improved sensitivity, precision, specificity, and satisfactory separation of underivatised CBDs and Chol from biological materials of endogenous species, especially in hemp and hen egg yolk. The scientific novelty of the present study is the application of the GC-MS method for quantifying underivatised CBDs-A, CBDs-N, and Chol in the samples of interest.

Tracking Plasma Membrane Damage Using a Ruthenium(II) Complex Phosphorescent Indicator Paired with Cholesterol.

Long-term in situ plasma membrane-targeted imaging is highly significant for investigating specific biological processes and functions, especially for the imaging and tracking of apoptosis processes of cells. However, currently developed membrane probes are rarely utilized to monitor the in situ damage of the plasma membrane. Herein, a transition-metal complex phosphorescent indicator, Ru-Chol, effectively paired with cholesterol, exhibits excellent properties on staining the plasma membrane, with excellent antipermeability, good photostability, large Stokes shift, and long luminescence lifetime. In addition, Ru-Chol not only has the potential to differentiate cancerous cells from normal cells but also tracks in real time the entire progression of cisplatin-induced plasma membrane damage and cell apoptosis. Therefore, Ru-Chol can serve as an efficient tool for the monitoring of morphological and physiological changes in the plasma membrane, providing assistance for drug screening and early diagnosis and treatment of diseases, such as immunodeficiency, diabetes, cirrhosis, and tumors.

A Coronin 1-Derived Peptide Inhibits Membrane Fusion by Modulating Membrane Organization and Dynamics.

Membrane fusion is considered the first step in the entry of enveloped viruses into the host cell. Several targeted strategies have been implemented to block viral entry by limiting the fusion protein to form a six-helix bundle, which is a prerequisite for fusion. Nonetheless, the development of broad-spectrum fusion inhibitors is essential to combat emerging and re-emerging viral infections. TG-23, a coronin 1, a tryptophan-aspartate-rich phagosomal protein-derived peptide, demonstrated inhibition of fusion between small unilamellar vesicles (SUVs) by modulating the membrane's physical properties. However, its inhibitory efficacy reduces with an increasing concentration of membrane cholesterol. The present work aims to develop a fusion inhibitor whose efficacy would be unaltered in the presence of membrane cholesterol. A stretch of the tryptophan-aspartic acid-containing peptide with a similar secondary structure and hydrophobicity profile of TG-23 from coronin 1 was synthesized, and its ability to inhibit SUV-SUV fusion with varying concentrations of membrane cholesterol was evaluated. Our results demonstrate that the GG-21 peptide inhibits fusion irrespective of the cholesterol content of the membrane. We have further evaluated the peptide-induced change in the membrane organization and dynamics utilizing arrays of steady-state and time-resolved fluorescence measurements and correlated these results with their effect on fusion. Interestingly, GG-21 displays inhibitory efficacy in a wide variety of lipid compositions despite having a secondary structure and physical properties similar to those of TG-23. Overall, our results advocate that the secondary structure and physical properties of the peptide may not be sufficient to predict its inhibitory efficacy.

Formulation design and characterization of silymarin liposomes for enhanced antitumor activity.

Liposomes, a nanoscale carrier, plays an important role in the delivery of drug, affects the in vivo efficacy of drugs. In this paper, silymarin(SM)-loaded liposomes was optimized using the response surface method (RSM), with entrapment efficiency (EE%) as an index. The formulation was optimized as follow: lecithin (7.8mg/mL), SM/lecithin (1/26) and lecithin/cholesterol (10/1). The optimized SM liposomes had a high EE (96.58 ±3.06%), with a particle size of 290.3 ±10.5nm and a zeta potential of +22.98 ±1.73mV. In vitro release tests revealed that SM was released in a sustained-release manner, primarily via diffusion mechanism. In vitro cytotoxicity studies demonstrated that the prepared SM liposomes had stronger inhibitory effects than the model drug. Overall, these results indicate that this liposome system is suitable for intravenous delivery to enhance the antitumor effects of SM.

Macrophages and Natural Killers Degrade α-Synuclein Aggregates.

Amyloid oligomers and fibrils are protein aggregates that exert a high cell toxicity. Efficient degradation of these protein aggregates can minimize the spread and progression of neurodegeneration. In this study, we investigate the properties of natural killer (NK) cells and macrophages in the degradation of α-synuclein (α-Syn) aggregates grown in a lipid-free environment and in the presence of phosphatidylserine and cholesterol (PS/Cho), which are lipids that are directly associated with the onset and progression of Parkinson's disease. We found that both types of α-Syn aggregates were endocytosed by neurons, which caused strong damage to cell endosomes. Our results also indicated that PS/Cho vesicles drastically increased the toxicity of α-Syn fibrils formed in their presence compared to the toxicity of α-Syn aggregates grown in a lipid-free environment. Both NK cells and macrophages were able to degrade α-Syn and α-Syn/Cho monomers, oligomers, and fibrils. Quantitative analysis of protein degradation showed that macrophages demonstrated substantially more efficient internalization and degradation of amyloid aggregates in comparison to NK cells. We also found that amyloid aggregates induced the proliferation of macrophages and NK cells and significantly changed the expression of their cytokines and chemokines.

Insight into the interaction of 5,6 epoxy-cholesterols with human serum albumin.

5,6-Epoxy-cholesterols has been recently revealed to control metabolic pathway in breast cancer, which makes investigating their binding interaction with human serum albumin (HSA) an attractive field of research. The main aim of this article is to examine the binding interaction of 5,6 α-epoxy-cholesterol (5,6 α EC) and 5,6 ß-epoxy-cholesterol (5,6 ß- EC) with HSA using different spectroscopic methods and molecular modeling. These compounds interact with HSA via hydrophobic interactions and hydrogen bonds with binding constants 6.3 × 105 M-1 for 5,6 α-epoxy-cholesterol and 6.9 × 105 M-1 for 5,6 ß-epoxy-cholesterol besides, the mechanism of the interaction can be attributed to static quenching. Circular dichroism data indicated that the α-helical content of HSA increased from 50.5 to 59.8 and 61.1 % after the addition of 5,6 α-ECs and 5,6 ß-EC, respectively, with a ratio of 1:2. Thermodynamic analysis revealed that binding between 5,6-epoxy-cholesterols and HSA is spontaneous and entropy-driven. The molecular docking and esterase-like activity experiments were performed to envision a link between the experimental and theoretical results. The optimal binding site of 5,6-epoxy-cholesterols with HSA was located in subdomain IIA. Moreover, theoretical calculations were performed using the B3LYP function with the 6-311++G (d,p) basis set, indicating the HOMO-LUMO energy gap of 7.874 eV for 5,6 α-epoxy-cholesterol and 7.873 eV for 5,6 ß-epoxy-cholesterol. The obtained findings are assumed to provide basic data for understanding the binding interactions of HSA with oxysterol compounds, which could help explore the pharmacokinetics and pharmacodynamics of oxysterol compounds.

Bitter taste receptor activation by cholesterol and an intracellular tastant.

Bitter taste sensing is mediated by type 2 taste receptors (TAS2Rs (also known as T2Rs)), which represent a distinct class of G-protein-coupled receptors1. Among the 26 members of the TAS2Rs, TAS2R14 is highly expressed in extraoral tissues and mediates the responses to more than 100 structurally diverse tastants2-6, although the molecular mechanisms for recognizing diverse chemicals and initiating cellular signalling are still poorly understood. Here we report two cryo-electron microscopy structures for TAS2R14 complexed with Ggust (also known as gustducin) and Gi1. Both structures have an orthosteric binding pocket occupied by endogenous cholesterol as well as an intracellular allosteric site bound by the bitter tastant cmpd28.1, including a direct interaction with the α5 helix of Ggust and Gi1. Computational and biochemical studies validate both ligand interactions. Our functional analysis identified cholesterol as an orthosteric agonist and the bitter tastant cmpd28.1 as a positive allosteric modulator with direct agonist activity at TAS2R14. Moreover, the orthosteric pocket is connected to the allosteric site via an elongated cavity, which has a hydrophobic core rich in aromatic residues. Our findings provide insights into the ligand recognition of bitter taste receptors and suggest activities of TAS2R14 beyond bitter taste perception via intracellular allosteric tastants.

Thermal evaporation as sample preparation for silver-assisted laser desorption/ionization mass spectrometry imaging of cholesterol in amyloid tissues.

Cholesterol plays an important biological role in the body, and its disruption in homeostasis and synthesis has been implicated in several diseases. Mapping the locations of cholesterol is crucial for gaining a better understanding of these conditions. Silver deposition has proven to be an effective method for analyzing cholesterol using mass spectrometry imaging (MSI). We optimized and evaluated thermal evaporation as an alternative deposition technique to sputtering for silver deposition in MSI of cholesterol. A silver layer with a thickness of 6 nm provided an optimal combination of cholesterol signal intensity and mass resolution. The deposition of an ultrathin nanofilm of silver enabled high-resolution MSI with a pixel size of 10 µm. We used this optimized method to visualize the distribution of cholesterol in the senile plaques in the brains of APP/PS1 mice, a model that resembles Alzheimer's disease pathology. We found that cholesterol was evenly distributed across the frontal cortex tissue, with no evidence of plaque-like accumulation. Additionally, we investigated the presence and distribution of cholesterol in myocardial sections of a human heart affected by wild-type ATTR amyloidosis. We identified the presence of cholesterol in areas with amyloid deposition, but complete colocalization was not observed.

A blood-brain barrier- and blood-brain tumor barrier-penetrating siRNA delivery system targeting gliomas for brain tumor immunotherapy.

Glioma is recognized as the most infiltrative and lethal form of central nervous system tumors and is known for its limited response to standard therapeutic interventions, high recurrence rate, and unfavorable prognosis. Recent progress in gene and immunotherapy presents a renewed sense of optimism in the treatment of glioblastoma. However, the barriers to overcome include the blood-brain barrier (BBB) and the blood-brain tumor barrier (BBTB), as well as the suppressive immune microenvironment. Overcoming these barriers remains a significant challenge. Here, we developed a lipid nanoparticle platform incorporating a dual-functional peptide (cholesterol-DP7-ACP-T7-modified DOTAP or DAT-LNP) capable of targeting glioma across the BBB and BBTB for brain tumor immunotherapy. This system was designed to achieve two key functions. First, the system could effectively penetrate the BBB during accumulation within brain tissue following intravenous administration. Second, this system enhances the maturation of dendritic cells, the polarization of M1 macrophages, and the activation of cytotoxic CD8+ T cells. This multifaceted approach effectively mitigates the immunosuppressive tumor microenvironment of glioma and promotes robust antitumor immune responses. Overall, the intravenous administration of the delivery system designed in this study demonstrates significant therapeutic potential for glioma and holds promising applications in the field of cancer immunotherapy.

Phosphatidylglycerol Acts as a Switch for Cholesterol-Dependent Membrane Binding of ApoE Signal Peptide.

The apolipoprotein E (ApoE) signal peptide is a short stretch of N-terminal amino acids that direct the ApoE protein to the endoplasmic reticulum after synthesis. Previous studies have shown that this peptide can bind to lipid membranes in a cholesterol-dependent manner; however, the mechanism of this interaction is yet to be clarified. In this study, we aimed to investigate how the composition of neighboring lipids affects the membrane-binding of the ApoE signal peptide. We found that a negatively charged lipid, such as phosphatidylglycerol, can act as a switch that reduces the binding efficiency of the peptide to cholesterol-rich membranes. Interestingly, phosphatidylethanolamine does not activate the cholesterol-dependent binding of the ApoE signal peptide yet acts synergistically to enhance the cholesterol sensitivity in phosphatidylglycerol-containing membranes. To the best of our knowledge, this is the first report of modulation of the affinity of a peptide for a membrane by a neighboring lipid rather than by the lipid-binding domain of the peptide. Our findings revealed a novel role of lipid diversity in modulating the membrane binding of the ApoE signal peptide and its potential implications in the unidirectional trafficking of a newly synthesized protein from the ribosomes to the endoplasmic reticulum.

Conventional vs PEGylated loaded liposomal formulations by microfluidics for delivering hydrophilic chemotherapy.

Developing drug delivery systems (DDSs) is one of the approaches used to improve cancer treatment, with the main goal of loading cancer drugs into a carrier targeting a specific organ and avoiding the distribution to healthy tissues. Nanoparticles (NPs) have been shown to be one of the optimum carriers that can be used as DDSs. Lipid-based NPs, such as liposomes, have been investigated in the current study due to their low toxicity and ability to carry hydrophilic and hydrophobic molecules. In the current studies, conventional liposomes composed of DPPC, and cholesterol and PEGylated liposomes composed of DPPC, cholesterol, and DSPE-PEG2000 are manufactured and loaded with Carboplatin. The study focused on investigating and comparing the impact of modifying the carboplatin-loaded liposomes with different concentrations of DSPE-PEG2000 on the NP diameter, polydispersity, ζ-potential, encapsulation efficiency (EE%), and drug release. The hydrodynamic microfluidic system was used to investigate any possible improvement in the EE% over other conventional methods. The results showed the microfluidic system's promising effect in enhancing the EE% of the Carboplatin. Moreover, the results showed a smaller diameter and higher stability of the PEGylated liposome. However, conventional liposomes represent better homogeneity and higher encapsulation efficiency for hydrophilic molecules.

Cholesterol-modified sphingomyelin chimeric lipid bilayer for improved therapeutic delivery.

Cholesterol (Chol) fortifies packing and reduces fluidity and permeability of the lipid bilayer in vesicles (liposomes)-mediated drug delivery. However, under the physiological environment, Chol is rapidly extracted from the lipid bilayer by biomembranes, which jeopardizes membrane stability and results in premature leakage for delivered payloads, yielding suboptimal clinic efficacy. Herein, we report a Chol-modified sphingomyelin (SM) lipid bilayer via covalently conjugating Chol to SM (SM-Chol), which retains membrane condensing ability of Chol. Systemic structure activity relationship screening demonstrates that SM-Chol with a disulfide bond and longer linker outperforms other counterparts and conventional phospholipids/Chol mixture systems on blocking Chol transfer and payload leakage, increases maximum tolerated dose of vincristine while reducing systemic toxicities, improves pharmacokinetics and tumor delivery efficiency, and enhances antitumor efficacy in SU-DHL-4 diffuse large B-cell lymphoma xenograft model in female mice. Furthermore, SM-Chol improves therapeutic delivery of structurally diversified therapeutic agents (irinotecan, doxorubicin, dexamethasone) or siRNA targeting multi-drug resistant gene (p-glycoprotein) in late-stage metastatic orthotopic KPC-Luc pancreas cancer, 4T1-Luc2 triple negative breast cancer, lung inflammation, and CT26 colorectal cancer animal models in female mice compared to respective FDA-approved nanotherapeutics or lipid compositions. Thus, SM-Chol represents a promising platform for universal and improved drug delivery.

Lithocholic acid-based oligomers as drug delivery candidates targeting model of lipid raft.

This study presents a new approach to designing a lithocholic acid functionalized oligomer (OLithocholicAA-X) that can be used as a drug carrier with additional, beneficial activity. Namely, this novel oligomer can incorporate an anti-cancer drug due to the application of an effective backbone as its component (lithocholic acid) alone is known to have anticancer activity. The oligomer was synthesized and characterized in detail by nuclear magnetic resonance, attenuated total reflectance Fourier-transform infrared spectroscopy, ultraviolet-visible spectroscopy, thermal analysis, and mass spectrometry analysis. We selected lipid rafts as potential drug carrier-membrane binding sites. In this respect, we investigated the effects of OLithocholicAA-X on model lipid raft of normal and altered composition, containing an increased amount of cholesterol (Chol) or sphingomyelin (SM), using Langmuir monolayers and liposomes. The surface topography of the studied monolayers was additionally investigated by atomic force microscopy (AFM). The obtained results showed that the investigated oligomer has affinity for a system that mimics a normal lipid raft (SM:Chol 2:1). On the other hand, for systems with an excess of SM or Chol, thermodynamically unfavorable fluidization of the films occurs. Moreover, AFM topographies showed that the amount of SM determines the bioavailability of the oligomer, causing fragmentation of its lattice.

Enhanced Oral Bioavailability of Progesterone in Bilosome Formulation: Fabrication, Statistical Optimization, and Pharmacokinetic Study.

Progesterone, a female sex steroid hormone, is highly lipophilic, leading to poor oral bioavailability. This study aimed to develop a progesterone bilosome system to enhance its oral bioavailability and retain it longer in the body. Progesterone vesicles were formulated with bile salts by thin film hydration method to prevent enzymatic and bile acid degradation. The Box-Behnken experimental design was used to statistically optimize progesterone bilosomes by checking the effect of phosphatidylcholine, cholesterol, and sodium deoxycholate on vesicle size, zeta potential, and entrapment efficiency. The optimum batch showed 239.5 nm vesicle size, -28.2 mV zeta potential and 84.08% entrapment efficiency, respectively, which were significantly affected by phosphatidylcholine and cholesterol concentration. The successful incorporation of progesterone in the system was evident from ATR-FTIR analysis that revealed no sharp progesterone peaks in bilosomes. TEM analysis confirmed the spherical structure and uniform bilosome vesicles. Furthermore, the in vitro drug release of progesterone bilosomes revealed a sustained pattern exhibiting 90% drug release in 48 h. The pharmacokinetic study in female ovariectomized Wistar rats confirmed the 4.287- and 9.75-fold enhanced oral bioavailability of the progesterone bilosomes than marketed capsules and progesterone API, respectively. Therefore, progesterone bilosome formulation can be further explored for improved oral administration in chronic treatments.