bims-novged 2024-02-25 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2024–02–25
nine papers selected by
the Merkel lab, Ludwig-Maximilians University

Closing the Gap between Experiment and Simulation─A Holistic Study on the Complexation of Small Interfering RNAs with Polyethylenimine.
Physicochemical Targeting of Lipid Nanoparticles to the Lungs Induces Clotting: Mechanisms and Solutions.
Systematic optimization of siRNA productive uptake into resting and activated T cells ex vivo.
Design of charge converting lipid nanoparticles via a microfluidic coating technique.
Lipid Nanoparticle (LNP) Delivery Carrier-Assisted Targeted Controlled Release mRNA Vaccines in Tumor Immunity.
A systematic study of the effect of lipid architecture on cytotoxicity and cellular uptake of cationic cubosomes.
Novel polyurethane-based ionene nanoparticles electrostatically stabilized with hyaluronic acid for effective gene therapy.
Nanoparticles Coated with Brain Microvascular Endothelial Cell Membranes can Target and Cross the Blood-Brain Barrier to Deliver Drugs to Brain Tumors.
Cell Membrane-Coated Nanoparticles for Precision Medicine: A Comprehensive Review of Coating Techniques for Tissue-Specific Therapeutics.

Mol Pharm. 2024 Feb 19.

Closing the Gap between Experiment and Simulation─A Holistic Study on the Complexation of Small Interfering RNAs with Polyethylenimine.

Jonas Binder, Joshua Winkeljann, Katharina Steinegger, Lara Trnovec, Daria Orekhova, Jonas Zähringer, Andreas Hörner, Valentin Fell, Philip Tinnefeld, Benjamin Winkeljann, Wolfgang Frieß, Olivia M Merkel.

  Rational design is pivotal in the modern development of nucleic acid nanocarrier systems. With the rising prominence of polymeric materials as alternatives to lipid-based carriers, understanding their structure-function relationships becomes paramount. Here, we introduce a newly developed coarse-grained model of polyethylenimine (PEI) based on the Martini 3 force field. This model facilitates molecular dynamics simulations of true-sized PEI molecules, exemplified by molecules with molecular weights of 1.3, 5, 10, and 25 kDa, with degrees of branching between 50.0 and 61.5%. We employed this model to investigate the thermodynamics of small interfering RNA (siRNA) complexation with PEI. Our simulations underscore the pivotal role of electrostatic interactions in the complexation process. Thermodynamic analyses revealed a stronger binding affinity with increased protonation, notably in acidic (endosomal) pH, compared to neutral conditions. Furthermore, the molecular weight of PEI was found to be a critical determinant of binding dynamics: smaller PEI molecules closely enveloped the siRNA, whereas larger ones extended outward, facilitating the formation of complexes with multiple RNA molecules. Experimental validations, encompassing isothermal titration calorimetry and single-molecule fluorescence spectroscopy, aligned well with our computational predictions. Our findings not only validate the fidelity of our PEI model but also accentuate the importance of in silico data in the rational design of polymeric drug carriers. The synergy between computational predictions and experimental validations, as showcased here, signals a refined and precise approach to drug carrier design.

Keywords:  Martini 3; RNA interference; polyplex; rationale design

DOI:  https://doi.org/10.1021/acs.molpharmaceut.3c00747
Adv Mater. 2024 Feb 23. e2312026

Physicochemical Targeting of Lipid Nanoparticles to the Lungs Induces Clotting: Mechanisms and Solutions.

Serena Omo-Lamai, Marco E Zamora, Manthan N Patel, Jichuan Wu, Jia Nong, Zhicheng Wang, Alina Peshkova, Aparajeeta Majumder, Jilian R Melamed, Liam S Chase, Eno-Obong Essien, Drew Weissman, Vladimir R Muzykantov, Oscar A Marcos-Contreras, Jacob W Myerson, Jacob S Brenner.

  Lipid nanoparticles (LNPs) have become the dominant drug delivery technology in industry, holding the promise to deliver RNA to up or down-regulate any protein of interest. LNPs have mostly been targeted to specific cell types or organs by physicochemical targeting in which LNP's lipid compositions are adjusted to find mixtures with the desired tropism. Here we examined lung tropic LNPs, whose organ tropism derives from containing either a cationic or ionizable lipid conferring a positive zeta potential. Surprisingly, we found these LNPs induce massive thrombosis. We show such thrombosis in the lungs and other organs, and that it is greatly exacerbated by pre-existing inflammation. This clotting is induced by a variety of formulations with cationic lipids, including LNPs and non-LNP nanoparticles, and even by lung-tropic ionizable lipids that do not have a permanent cationic charge. The mechanism depends on the LNPs binding to and then changing the conformation of fibrinogen, which then activates platelets and thrombin. Based on these mechanisms, we engineered multiple solutions that enable positively charged LNPs to target the lungs while ameliorating thrombosis. Our findings implicate thrombosis as a major barrier that blood erects against LNPs with positive zeta potential and illustrate how physicochemical targeting approaches must be investigated early for risks and re-engineered with a careful understanding of biological mechanisms. This article is protected by copyright. All rights reserved.

Keywords:  Drug Delivery; Lipid nanoparticles; Nanomedicine; Side-effects; Thrombosis

DOI:  https://doi.org/10.1002/adma.202312026
Biomed Pharmacother. 2024 Feb 20. pii: S0753-3322(24)00166-5. [Epub ahead of print]172 116285

Systematic optimization of siRNA productive uptake into resting and activated T cells ex vivo.

A Kremer, T Ryaykenen, R A Haraszti.

  RNA-based medicines are ideally suited for precise modulation of T cell phenotypes in anti-cancer immunity, in autoimmune diseases and for ex vivo modulation of T-cell-based therapies. Therefore, understanding productive siRNA uptake to T cells is of particular importance. Most studies used unmodified siRNAs or commercially available siRNAs with undisclosed chemical modification patterns to show functionality in T cells. Despite being an active field of research, robust siRNA delivery to T cells still represents a formidable challenge. Therefore, a systematic approach is needed to further optimize and understand productive siRNA uptake pathways to T cells. Here, we compared conjugate-mediated and nanoparticle-mediated delivery of siRNAs to T cells in the context of fully chemically modified RNA constructs. We showed that lipid-conjugate-mediated delivery outperforms lipid-nanoparticle-mediated and extracellular-vesicle-mediated delivery in activated T cells ex vivo. Yet, ex vivo manipulation of T cells without the need of activation is of great therapeutic interest for CAR-T, engineered TCR-T and allogeneic donor lymphocyte applications. We are first to report productive siRNA uptake into resting T cells using lipid-conjugate-mediated delivery. Interestingly, we observed strong dependence of silencing activity on lipid-conjugate-identity in resting T cells but not in activated T cells. This phenomenon is consistent with our early uptake kinetics data. Lipid-conjugates also enabled delivery of siRNA to all mononuclear immune cell types, including both lymphoid and myeloid lineages. These findings are expected to be broadly applicable for ex vivo modulation of immune cell therapies.

Keywords:  Extracellular vesicle; Immune cell; Lipid nanoparticle; Lipid-mediated delivery; T cell; siRNA

DOI:  https://doi.org/10.1016/j.biopha.2024.116285
Drug Deliv Transl Res. 2024 Feb 21.

Design of charge converting lipid nanoparticles via a microfluidic coating technique.

Katrin Zöller, Soheil Haddadzadegan, Sera Lindner, Florina Veider, Andreas Bernkop-Schnürch.

  It was the aim of this study to design charge converting lipid nanoparticles (LNP) via a microfluidic mixing technique used for the preparation and coating of LNP. LNP consisting of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol, N-(carbonyl-methoxypolyethyleneglycol-2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine (MPEG-2000-DSPE), and various cationic surfactants were prepared at diverging flow rate ratios (FRR) via microfluidic mixing. Utilizing a second chip in the microfluidic set-up, LNP were coated with polyoxyethylene (9) nonylphenol monophosphate ester (PNPP). LNP were examined for their stability in different physiologically relevant media as well as for hemolytic and cytotoxic effects. Finally, phosphate release and charge conversion of PNPP-coated LNP were evaluated after incubation with alkaline phosphatase and on Caco2-cells. LNP produced at an FRR of 5:1 exhibited a size between 80 and 150 nm and a positive zeta potential. Coating with PNPP within the second chip led to LNP exhibiting a negative zeta potential. After incubation with 1 U/ml alkaline phosphatase for 4 h, zeta potential of the LNP containing 1,2-dioleoyloxy-3-trimethylammonium-propane chloride (DOTAP) as cationic component shifted from - 35 mV to approximately + 5 mV. LNP prepared with other cationic surfactants remained slightly negative after enzymatic phosphate cleavage. Manufacturing of LNP containing PNPP and DOTAP via connection of two chips in a microfluidic instrument proves to show efficient change in zeta potential from negative to positive after incubation with alkaline phosphatase.

Keywords:  DOTAP; Lipid nanoparticles; Microfluidics; Zeta change

DOI:  https://doi.org/10.1007/s13346-024-01538-5
Vaccines (Basel). 2024 Feb 12. pii: 186. [Epub ahead of print]12(2):

Lipid Nanoparticle (LNP) Delivery Carrier-Assisted Targeted Controlled Release mRNA Vaccines in Tumor Immunity.

Liusheng Wu, Xiaoqiang Li, Xinye Qian, Shuang Wang, Jixian Liu, Jun Yan.

  In recent years, lipid nanoparticles (LNPs) have attracted extensive attention in tumor immunotherapy. Targeting immune cells in cancer therapy has become a strategy of great research interest. mRNA vaccines are a potential choice for tumor immunotherapy, due to their ability to directly encode antigen proteins and stimulate a strong immune response. However, the mode of delivery and lack of stability of mRNA are key issues limiting its application. LNPs are an excellent mRNA delivery carrier, and their structural stability and biocompatibility make them an effective means for delivering mRNA to specific targets. This study summarizes the research progress in LNP delivery carrier-assisted targeted controlled release mRNA vaccines in tumor immunity. The role of LNPs in improving mRNA stability, immunogenicity, and targeting is discussed. This review aims to systematically summarize the latest research progress in LNP delivery carrier-assisted targeted controlled release mRNA vaccines in tumor immunity to provide new ideas and strategies for tumor immunotherapy, as well as to provide more effective treatment plans for patients.

Keywords:  delivery carrier; lipid nanoparticles (LNPs); mRNA vaccine; review; tumor immunity

DOI:  https://doi.org/10.3390/vaccines12020186
J Colloid Interface Sci. 2024 Feb 15. pii: S0021-9797(24)00343-6. [Epub ahead of print]663 82-93

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

S Pushpa Ragini, Brendan P Dyett, Sampa Sarkar, Jiali Zhai, Jacinta F White, Rajkumar Banerjee, Calum J Drummond, Charlotte E Conn.

   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.

Keywords:  Cationic lipids; Cellular uptake; Critical packing parameter; Cubosomes; Cytotoxicity; Pharmacophore

DOI:  https://doi.org/10.1016/j.jcis.2024.02.099
Colloids Surf B Biointerfaces. 2024 Feb 16. pii: S0927-7765(24)00060-2. [Epub ahead of print]236 113802

Novel polyurethane-based ionene nanoparticles electrostatically stabilized with hyaluronic acid for effective gene therapy.

Athar Mahdieh, Hamidreza Motasadizadeh, Samane Maghsoudian, Alireza Sabzevari, Fereshte Khalili, Hamid Yeganeh, Bo Nyström.

  Gene therapy is considered to be a valuable strategy for effective cancer treatment. However, the development of effective delivery systems that can specifically deliver gene materials, such as siRNA to tumor tissues plays a critical role in cancer therapy. In the present study, we have developed a novel complex that is based on an electrostatic interaction between cationic polyurethane ionene (CPUI) nanoparticles and an anti-signal transducer and activator of transcription 3 (STAT3) siRNA. For active targeting, hyaluronic acid (HA) was used to coat the complexes, which significantly reduced the cytotoxicity of the blank nanocarriers while demonstrating high transport efficiency of the siRNA via the CD44-mediated endocytosis pathway in MCF-7 breast cancer cells. The targeted nanocarriers (HA/CPUI/siRNA) showed significantly higher cellular internalization in flow cytometry and confocal microscopy compared with the non-targeted system (CPUI/siRNA). In addition, the incorporation of HA on the surface of the complexes resulted in significantly greater suppression of the STAT3 gene compared to the corresponding non-targeted formulation. Whole-body fluorescence images showed more significant tumor accumulation of the targeted nanocarriers in 4T1 breast tumor-bearing mice. Therefore, HA/CPUI/siRNA nanocarriers are an interesting option for the siRNA-targeted treatment of breast cancer cells.

Keywords:  Breast cancer therapy; Hyaluronic acid; Ionene nanoparticles; Polyurethane; STAT3 siRNA; Targeted gene delivery

DOI:  https://doi.org/10.1016/j.colsurfb.2024.113802
Small. 2024 Feb 23. e2306714

Nanoparticles Coated with Brain Microvascular Endothelial Cell Membranes can Target and Cross the Blood-Brain Barrier to Deliver Drugs to Brain Tumors.

Huajian Chen, Jingsen Ji, Li Zhang, Chuangcai Luo, Taoliang Chen, Yuxuan Zhang, Chengcheng Ma, Yiquan Ke, Jihui Wang.

  The blood-brain barrier (BBB) contains tightly connected brain microvascular endothelial cells (BMECs) that hinder drug delivery to the brain, which makes brain tumors difficult to treat. Previous studies have shown that nanoparticles coated with tumor cell membranes selectively target their homologous tumors. Therefore, this study investigated whether bEnd.3-line BMEC membrane-coated nanoparticles with poly(lactide-co-glycolide)-poly(ethylene glycol)-based doxorubicin-loaded cores (BM-PDs) can be used to target BMECs and cross the BBB. In vitro, the BM-PDs effectively target BMECs and cross a BBB model. The BM-PDs enter the BMECs via macropinocytosis, clathrin-mediated endocytosis, caveolin-mediated endocytosis, and membrane fusion, which result in excellent cellular uptake. The BM-PDs also show excellent cellular uptake in brain tumor cells. In vivo, the BM-PDs target BMECs, cross the BBB, accumulate in brain tumors, and efficiently kill tumor cells. Therefore, the proposed strategy has great therapeutic potential owing to its ability to cross the BBB to reach brain tumors.

Keywords:  bEnd.3 cell; blood-brain barrier; brain microvascular endothelial cells; glioblastoma; nanoparticles

DOI:  https://doi.org/10.1002/smll.202306714
Int J Mol Sci. 2024 Feb 08. pii: 2071. [Epub ahead of print]25(4):

Cell Membrane-Coated Nanoparticles for Precision Medicine: A Comprehensive Review of Coating Techniques for Tissue-Specific Therapeutics.

Andrés Fernández-Borbolla, Lorena García-Hevia, Mónica L Fanarraga.

  Nanoencapsulation has become a recent advancement in drug delivery, enhancing stability, bioavailability, and enabling controlled, targeted substance delivery to specific cells or tissues. However, traditional nanoparticle delivery faces challenges such as a short circulation time and immune recognition. To tackle these issues, cell membrane-coated nanoparticles have been suggested as a practical alternative. The production process involves three main stages: cell lysis and membrane fragmentation, membrane isolation, and nanoparticle coating. Cell membranes are typically fragmented using hypotonic lysis with homogenization or sonication. Subsequent membrane fragments are isolated through multiple centrifugation steps. Coating nanoparticles can be achieved through extrusion, sonication, or a combination of both methods. Notably, this analysis reveals the absence of a universally applicable method for nanoparticle coating, as the three stages differ significantly in their procedures. This review explores current developments and approaches to cell membrane-coated nanoparticles, highlighting their potential as an effective alternative for targeted drug delivery and various therapeutic applications.

Keywords:  biomimetic nanoparticle; biomimicry; homotypic targeting; nanomedicine; nanoparticle coating; targeted drug delivery

DOI:  https://doi.org/10.3390/ijms25042071