bims-novged 2021-11-21 papers

on Non-viral vectors for gene delivery

Issue of 2021–11–21
nine papers selected by
the Merkel lab, Ludwig-Maximilians University and Benjamin Winkeljann, Ludwig-Maximilians University

Ginsenoside as a new stabilizer enhances the transfection efficiency and biocompatibility of cationic liposome.
Modified cyclodextrin-based nanoparticles mediated delivery of siRNA for Huntingtin gene silencing across an in vitro BBB model.
Cationic Glycopolyelectrolytes for RNA Interference in Tick Cells.
PEI-PEG-Coated Mesoporous Silica Nanoparticles Enhance the Antitumor Activity of Tanshinone IIA and Serve as a Gene Transfer Vector.
Poly (β amino esters) Copolymers: Novel Potential Vectors for Delivery of Genes and Related Therapeutics.
pH-Dependent Phase Behavior and Stability of Cationic Lipid-mRNA Nanoparticles.
Stable Loading and Delivery of Melittin with Lipid-Coated Polymeric Nanoparticles for Effective Tumor Therapy with Negligible Systemic Toxicity.
Engineering pH-Sensitive Stable Nanovesicles for Delivery of MicroRNA Therapeutics.
Potential of Mean Force for DNA Wrapping Around a Cationic Nanoparticle.

Biomater Sci. 2021 Nov 17.

Ginsenoside as a new stabilizer enhances the transfection efficiency and biocompatibility of cationic liposome.

Mao-Ze Wang, Yang Xu, Jia-Feng Xie, Zhi-Hong Jiang, Li-Hua Peng.

Nucleic acid drugs have emerged as important therapeutics but their clinical application has been greatly limited by their large molecular weight, high polarity, negative charge and short half-life. Cationic liposomes (CLs) have gained wide attention as non-viral vectors for nucleic acid delivery. However, the absolute transfection efficiency of CLs can still be enhanced while their cytotoxicity should be decreased simultaneously. Ginsenosides, obtained from natural plants, possess a similar steroid structure to cholesterol and might be an alternative to cholesterol for acting as a membrane stabilizer of CLs. Herein, seven kinds of ginsenoside-based compounds were utilized to prepare CLs (GCLs) and their efficacy in siRNA delivery was investigated. The particle sizes of the GCLs were 90-300 nm and the siRNA delivery efficiencies were in the range of 23.6%-78.4%. Rg5-based CLs (Rg5-CLs) exhibited the highest transfection efficiency of 81% and the lowest toxicity, with 82% cell viability obtained even at high concentrations. Ginsenosides are shown as promising biomaterials as membrane stabilizers of CLs. Rg5-CLs have been demonstrated as efficient non-viral vectors with high transfection efficiency and good biocompatibility for gene delivery, possessing great potential for gene therapy.

DOI: https://doi.org/10.1039/d1bm01353j
Eur J Pharm Biopharm. 2021 Nov 15. pii: S0939-6411(21)00276-9. [Epub ahead of print]

Modified cyclodextrin-based nanoparticles mediated delivery of siRNA for Huntingtin gene silencing across an in vitro BBB model.

Monique C P Mendonça, Michael F Cronin, John F Cryan, Caitriona M O'Driscoll.

  Huntington's disease (HD) is a neurodegenerative disorder caused by a mutation in the huntingtin (HTT) gene, leading to a toxic version of the HTT protein. There are currently no disease-modifying therapies available. In this scenario, gene-based treatments for HD aimed at lowering HTT levels have become one of the most promising emerging therapeutic options. To date, however, promising results have only been achieved following direct intrathecal or intracranial injections designed to circumvent the blood-brain barrier (BBB). Consequently, efforts to develop less invasive delivery platforms are highly desirable. Here, we described a novel delivery system based on modified cyclodextrin nanoparticles (CDs) loaded with small interfering RNAs (siRNAs) targeting HTT andcomplexed with the rabies virus glycoprotein(RVG), a BBB-shuttle peptide. Results using an in vitro BBB model, indicate the formulation successfully crosses the brain endothelial cells, releases the encapsulated siRNAs into the cytoplasm of neuronal cells, and mediates downregulation of HTT. In conclusion, the CD platform is a promising option for delivery of siRNA-based therapeutics for HD with wider potential to treat other diseases with a genetically validated target in the central nervous system.

Keywords:  Huntington’s disease; Nanocarriers; RVG; non-viral gene therapy; targeted cyclodextrins

DOI:  https://doi.org/10.1016/j.ejpb.2021.11.003
Biomacromolecules. 2021 Nov 18.

Cationic Glycopolyelectrolytes for RNA Interference in Tick Cells.

Kelli A Stockmal, Latoyia P Downs, Ashley N Davis, Lisa K Kemp, Shahid Karim, Sarah E Morgan.

The black-legged tick (Ixodes scapularis) is the primary vector for bacteria that cause Lyme disease (Borrelia burgdorferi), where numerous glycosylated tick proteins are involved at the interface of vector-host-pathogen interactions. Reducing the expression of key tick proteins, such as selenoprotein K (SelK), through RNA interference is a promising approach to reduce pathogen transmission, but efficient delivery of nucleic acids to arthropods has proven challenging. While cationic glycopolymers have been used as nonviral gene delivery vehicles in mammalian cells, their use in arthropod or insect gene transfection has not been established. In this study, statistical acrylamide-based cationic glycopolymers with glucose or galactose pendant groups were synthesized by reversible addition-fragmentation chain transfer polymerization, and the effects of the saccharide pendant group and cationic monomer loading on polymer cytotoxicity, RNA complexation, and SelK gene knockdown in ISE6 cells were evaluated. All polymers exhibited low cytotoxicity, yet RNA/copolymer complex cell uptake and gene knockdown were highly dependent on the saccharide structure and the N:P (amino to phosphate groups) ratio.

DOI: https://doi.org/10.1021/acs.biomac.1c00824
Evid Based Complement Alternat Med. 2021 ;2021 6756763

PEI-PEG-Coated Mesoporous Silica Nanoparticles Enhance the Antitumor Activity of Tanshinone IIA and Serve as a Gene Transfer Vector.

Yinxing Zhu, Miao Yue, Ting Guo, Fang Li, Zhifeng Li, Dazhuang Yang, Mei Lin.

Tanshinone IIA (TanIIA) and gene therapy both hold promising potentials in hepatocellular carcinoma (HCC) treatment. However, low solubility and poor bioavailability of TanIIA limit its clinical application. Similarly, gene therapy with GPC3-shRNA, a type of short hairpin RNAs (shRNAs) capable of silencing the glypican-3 (GPC3) expression, is seriously limited due to its susceptibility to nuclease degradation and high off-target effects. In the present study, polyethyleneimine (PEI)-polyethylene glycol (PEG)-coated mesoporous silica nanoparticles (MSN-PEG) were used as a drug carrier. By encapsulating TanIIA into MSN-PEG, we synthesized MSN-TanIIA-PEG nanoparticles and observed the involved characteristics. This was followed by exploration of antitumor activity on the HepG2 cell lines in vitro. Meanwhile, in order to construct GPC3-shRNA plasmids, a shRNA sequence targeting GPC3 was synthesized and cloned into the pSLenti-U6 vector. Accordingly, the performance of MSN-PEG as a gene transfer carrier for GPC3-shRNA gene therapy of HCC in vitro was evaluated, including transfection efficiency and DNA binding biological characteristics. The results indicated successful encapsulation of TanIIA in MSN-PEG, which had satisfactory efficacy, favorable dispersity, suitable particle size, and sustained release effect. The in vitro anti-HCC effects of nano-TanIIA were greatly improved, which outperformed free-TanIIA in terms of proliferation and invasion inhibition, as well as apoptosis induction of HCC cells. As expected, MSN-PEG possessed excellent gene delivery capacity with good binding, release, and protection from RNase digestion. Using MSN-PEG as a gene carrier, the plasmids were successfully transfected into HepG2 cells, and both the mRNA and protein expressions of GPC3 were significantly downregulated. It was thus concluded that a sustained release TanIIA delivery system for HCC treatment was synthesized and that MSN-PEG could also serve as a gene transfer carrier for gene therapy. More interestingly, MSN-PEG may be a potential delivery platform that combines TanIIA and GPC3-shRNA together to enhance their synergistic effect.

DOI: https://doi.org/10.1155/2021/6756763
Int J Pharm. 2021 Nov 11. pii: S0378-5173(21)01095-4. [Epub ahead of print] 121289

Poly (β amino esters) Copolymers: Novel Potential Vectors for Delivery of Genes and Related Therapeutics.

Sajid Iqbal, Zhongxi Zhao.

  The unique properties of polymers have performed an essential contribution to the drug delivery system by providing an outstanding platform for the delivery of macromolecules and genes. However, the block copolymers have been the subject of many recently published works whose results have demonstrated excellent performance in drug targeting. Poly(β-amino esters) (PβAEs) copolymers are the synthetic cationic polymers that are tailored by chemically joining PβAEs with other additives to demonstrate extraordinary efficiency in designing pre-defined and pre-programmed nanostructures, site-specific delivery, andovercoming the distinct cellular barriers. Different compositional and structural libraries could be generated by combinatorial chemistry and by the addition of various novel functional additives that fulfill the multiple requirements of targeted delivery. These intriguing attributes allow PβAE-copolymers to have customized therapeutic functions such as excellent encapsulation capacity, high stability, and stimuli-responsive release. Here, we give an overview of PβAE copolymers-based formulations along with focusing on most notable improvements such as structural modifications, bio-conjugations, and stimuli-responsive approaches, for safe and effective nucleic acids delivery.

Keywords:  Copolymers; Cytotoxicity; Nanoparticles; Poly(β-amino esters); Transfection; nucleic acid delivery

DOI:  https://doi.org/10.1016/j.ijpharm.2021.121289
J Pharm Sci. 2021 Nov 11. pii: S0022-3549(21)00610-9. [Epub ahead of print]

pH-Dependent Phase Behavior and Stability of Cationic Lipid-mRNA Nanoparticles.

Nicholas R Larson, Gang Hu, Yangjie Wei, Anthony Tuesesca, M Laird Forrest, C Russell Middaugh.

Lipid nanoparticles (LNPs) containing mRNA can deliver genetic material to cells for use as vaccines or protein replacement therapies. We characterized the effect of solution pH on cationic LNPs containing green fluorescent protein (EGFP) mRNA and their transfection efficiency. We compared the structural and colloidal properties of mRNA LNPs with LNPs not containing mRNA and mRNA free in solution. We used a combination of biophysical technique to build a picture of the structure of the lipids and mRNA across pH and temperature in the form of an empirical phase diagram (EPD). A combination of Fourier-transform infrared (FTIR) spectroscopy and differential scanning calorimetry was used to investigate lipid phase behavior. The mRNA-LNPs transition from an inverse hexagonal phase at pH values below the pKa of the cationic lipid to a lamellar phase above the pKa. At higher temperatures the mRNA-LNPs also transitioned from an inverse hexagonal phase to a lamellar phase indicating the inverse hexagonal phase is more thermodynamically favorable. Based on circular dichroism, the mRNA within the LNP has more A form structure at pH values below the lipid pKa than above it. Optical density, zeta potential and dynamic light scattering measurements were used to probe the colloidal stability of the mRNA-LNPs. The particles were larger and more prone to aggregation below the pKa. A stability study was performed to relate the biophysical characteristics to the storage of the particles in solution at 4 and 25°C. mRNA-LNPs had the highest transfection efficiency and stability at pH values below the pKa. However, there was a trade-off between the stability and aggregation propensity since at very low pH the particles were most prone to aggregation. We performed kinetic experiments to show that the time scale of the pH-dependent phase behavior is slow (6 hour transition) and the transition from lamellar to inverse hexagonal phases is irreversible. This suggests that the lamellar phase is less stable and kinetically trapped. Our findings deepen our structural understanding of mRNA-LNPs and will aid the development of related formulations.

DOI: https://doi.org/10.1016/j.xphs.2021.11.004
ACS Appl Mater Interfaces. 2021 Nov 18.

Stable Loading and Delivery of Melittin with Lipid-Coated Polymeric Nanoparticles for Effective Tumor Therapy with Negligible Systemic Toxicity.

Ran Ye, Yuan Zheng, Yang Chen, Xiaohui Wei, Sanyuan Shi, Yuetan Chen, Wanxin Zhu, Anqi Wang, Liuxin Yang, Yuhong Xu, Jinliang Peng.

  Melittin is a potential anticancer candidate with remarkable antitumor activity and ability to overcome tumor drug resistance. However, the clinical applications of melittin are largely restricted by its severe hemolytic activity and nonspecific cytotoxicity after systemic administration. Here, a biocompatible and stable melittin-loaded lipid-coated polymeric nanoparticle (MpG@LPN) formulation that contains a melittin/poly-γ-glutamic acid nanoparticle inner core, a lipid membrane middle layer, and a polyethylene glycol (PEG) and PEG-targeting molecule outer shell was designed. The formulations were prepared by applying a self-assembly procedure based on intermolecular interactions, including electrostatic attraction and hydrophobic effect. The core-shell MpG@LPN presented high efficiency for melittin encapsulation and high stability in physiological conditions. Hemolysis and cell proliferation assays showed that the PEG-modified MpG@LPN had almost no hemolytic activity and nonspecific cytotoxicity even at high concentrations. The modification of targeting molecules on the MpG@LPNs allowed for the selective binding with target tumor cells and cytolytic activity via apoptosis induction. In vivo experiments revealed that MpG@LPNs can remarkably inhibit the growth of tumors without the occurrence of hemolysis and tissue toxicity. Results suggested that the developed MpG@LPN with a core-shell structure can effectively address the main obstacles of melittin in clinical applications and has great potential in cancer treatment.

Keywords:  core−shell structure; hemolysis; lipid-coated polymeric nanoparticle; melittin; tumor therapy

DOI:  https://doi.org/10.1021/acsami.1c17618
Small. 2021 Nov 16. e2101959

Engineering pH-Sensitive Stable Nanovesicles for Delivery of MicroRNA Therapeutics.

Ariadna Boloix, Natalia Feiner-Gracia, Mariana Köber, Javier Repetto, Rosa Pascarella, Aroa Soriano, Marc Masanas, Nathaly Segovia, Guillem Vargas-Nadal, Josep Merlo-Mas, Dganit Danino, Inbal Abutbul-Ionita, Laia Foradada, Josep Roma, Alba Córdoba, Santi Sala, Josep Sánchez de Toledo, Soledad Gallego, Jaume Veciana, Lorenzo Albertazzi, Miguel F Segura, Nora Ventosa.

  MicroRNAs (miRNAs) are small non-coding endogenous RNAs, which are attracting a growing interest as therapeutic molecules due to their central role in major diseases. However, the transformation of these biomolecules into drugs is limited due to their unstability in the bloodstream, caused by nucleases abundantly present in the blood, and poor capacity to enter cells. The conjugation of miRNAs to nanoparticles (NPs) could be an effective strategy for their clinical delivery. Herein, the engineering of non-liposomal lipid nanovesicles, named quatsomes (QS), for the delivery of miRNAs and other small RNAs into the cytosol of tumor cells, triggering a tumor-suppressive response is reported. The engineered pH-sensitive nanovesicles have controlled structure (unilamellar), size (<150 nm) and composition. These nanovesicles are colloidal stable (>24 weeks), and are prepared by a green, GMP compliant, and scalable one-step procedure, which are all unavoidable requirements for the arrival to the clinical practice of NP based miRNA therapeutics. Furthermore, QS protect miRNAs from RNAses and when injected intravenously, deliver them into liver, lung, and neuroblastoma xenografts tumors. These stable nanovesicles with tunable pH sensitiveness constitute an attractive platform for the efficient delivery of miRNAs and other small RNAs with therapeutic activity and their exploitation in the clinics.

Keywords:  cancer therapy; miRNAs delivery; nanocarriers; nanovesicles; neuroblastoma; pediatric cancer; quatsomes; siRNAs delivery

DOI:  https://doi.org/10.1002/smll.202101959
J Chem Theory Comput. 2021 Nov 18.

Potential of Mean Force for DNA Wrapping Around a Cationic Nanoparticle.

Sehui Bae, Jun Soo Kim.

Sharp bending and wrapping of DNA around proteins and nanoparticles (NPs) has been of extensive research interest. Here, we present the potential of mean force (PMF) for wrapping a DNA double helix around a cationic NP using coarse-grained models of a double-stranded DNA and a cationic NP. Starting from a NP wrapped around by DNA, the PMF was calculated along the distance between the center of the NP and one end of the DNA molecule. A relationship between the distance and the extent of DNA wrapping is used to calculate the PMF as a function of DNA wrapping around a NP. In particular, the PMF was compared for two DNA sequences of (AT)25/(AT)25 and (AC)25/(GT)25, for which the persistence lengths are different by ∼10 nm. The simulation results provide solid evidence of the thermodynamic preference for complex formation of a cationic NP with more flexible DNA over the less flexible DNA. Furthermore, we estimated the elastic energy of DNA bending, which was in good order-of-magnitude agreement with the theoretical prediction of elastic rods. This work suggests that the variation of sequence-dependent DNA flexibility can be utilized in DNA nanotechnologies, in which the position and dynamics of NPs are regulated on large-scale DNA structures, or the structural transformation of DNA is triggered by the sequence-dependent binding of NPs.

DOI: https://doi.org/10.1021/acs.jctc.1c00797