bims-novged 2021-12-26 papers

on Non-viral vectors for gene delivery

Issue of 2021–12–26
ten papers selected by
the Merkel lab, Ludwig-Maximilians University and Benjamin Winkeljann, Ludwig-Maximilians University

Phenylboronic Acid-Modified Polyamidoamine Mediated the Transfection of Polo-Like Kinase-1 siRNA to Achieve an Anti-Tumor Efficacy.
The Effect of Repeat Administration of Lipoplexes on Gene Delivery, Biodistribution, and Cytokine Response in Immunocompetent Tumor-bearing Mice.
Phosphonodithioester-Amine Coupling as a Key Reaction Step for the Design of Cationic Amphiphiles Used for Gene Delivery.
On the mechanism of tissue-specific mRNA delivery by selective organ targeting nanoparticles.
Dynamic Stabilization of Unit Polyion Complexes Incorporating Small Interfering RNA by Fine-Tuning of Cationic Block Length in Two-Branched Poly(ethylene glycol)-b-poly(l-lysine).
Dual targeted delivery of statins and nucleic acids by chitosan-based nanoparticles for enhanced antiatherosclerotic efficacy.
Challenges and Opportunities for Nucleic Acid Therapeutics.
Imaging of the Entry Pathway of a Cell-Penetrating Peptide-DNA Complex From the Extracellular Space to Chloroplast Nucleoids Across Multiple Membranes in Arabidopsis Leaves.
Anti-MicroRNA-21 Oligonucleotide Loaded Spermine-Modified Acetalated Dextran Nanoparticles for B1 Receptor-Targeted Gene Therapy and Antiangiogenesis Therapy.
Insights into nucleic acid-based self-assembling nanocarriers for targeted drug delivery and controlled drug release.

Int J Nanomedicine. 2021 ;16 8037-8048

Phenylboronic Acid-Modified Polyamidoamine Mediated the Transfection of Polo-Like Kinase-1 siRNA to Achieve an Anti-Tumor Efficacy.

Gu Gong, Xiuhui Tang, Jiayuan Zhang, Xiao Liang, Jiebing Yang, Quanshun Li.

   Background: The construction of tumor-targeting carriers with favorable transfection efficiency was of great significance to achieve the tumor gene therapy. The phenylboronic acid-modified polyamidoamine (namely PP) was employed as a carrier for the delivery of Polo-like kinase-1 siRNA (siPlk-1), inducing an obvious anti-tumor response.
Materials and Methods: The interaction between PP and siPlk-1 was evaluated by gel retardation assay. The transfection efficiency and tumor-targeting ability were analyzed by flow cytometry and confocal laser scanning microscopy, using hepatocarcinoma cell line HepG2 as a model. The anti-proliferation effect of PP/siPlk-1 and related mechanism were studied using the strategies of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell apoptosis and cell cycle arrest. The anti-migration effect induced by PP/siPlk-1 delivery was assayed by wound healing and Transwell migration techniques. Finally, quantitative real-time PCR and Western blotting were performed to measure the expression level of Plk-1 and other key targets.
Results: The derivative PP could achieve the condensation of siPlk-1 into stable nanoparticles at nitrogen/phosphate groups ratio (N/P ratio) of >3.0, and it could facilitate the transfection of siPk-1 in a phenylboronic acid-dependent manner. The PP/siPlk-1 nanoparticles exhibited obvious anti-proliferation effect owing to the gene silence of Plk-1, which was identified to be associated with the cell apoptosis and cell cycle arrest at G2 phase. Meanwhile, PP/siPlk-1 transfection could efficiently suppress the migration and invasion of tumor cells.
Conclusion: The derivative PP has been demonstrated to be an ideal tumor-targeting carrier for the delivery of Plk-1 siRNA, exhibiting great potential in the gene therapy of malignant tumors.

Keywords:  gene therapy; phenylboronic acid; polyamidoamine; siPlk-1; tumor-targeting ability

DOI:  https://doi.org/10.2147/IJN.S329433
J Pharm Sci. 2021 Dec 17. pii: S0022-3549(21)00695-X. [Epub ahead of print]

The Effect of Repeat Administration of Lipoplexes on Gene Delivery, Biodistribution, and Cytokine Response in Immunocompetent Tumor-bearing Mice.

Jamie L Betker, Thomas J Anchordoquy.

  It is becoming increasingly clear that the intravenous administration of nanoparticles elicits an immune response that compromises delivery efficiency and can be life threatening. This study investigated both the systemic and tissue-level cytokine response to repeat administration of lipoplexes coated with either lactose or PEG. We report that blood cytokine levels differ significantly from that observed in individual tissues. While we consistently observed a reduced cytokine response to lactosylated particles, this did not result in enhanced delivery or expression as compared to PEGylated formulations. We also document that repeat injection did not increase plasmid levels in the liver, lung, or spleen, but delivery to the tumor was enhanced under these conditions. In addition, we show that changes in neither blood nor tissue cytokines correlated strongly with reporter gene expression, and we observed relatively constant expression efficiencies (RLU/ng plasmid) across all tissues despite a considerably reduced cytokine response in the tumor. Together, these results indicate that both biodistribution and cytokine responses are dramatically altered by a repeat intravenous injection of lipoplexes, and that the mechanisms regulating reporter gene expression are not straightforward.

Keywords:  Cancer; Cationic lipid; DNA delivery; Immune Response; Lipid nanoparticle; Lipoplex; Non-viral gene delivery; Pegylation; Transfection, Plasmid DNA

DOI:  https://doi.org/10.1016/j.xphs.2021.12.017
Molecules. 2021 Dec 11. pii: 7507. [Epub ahead of print]26(24):

Phosphonodithioester-Amine Coupling as a Key Reaction Step for the Design of Cationic Amphiphiles Used for Gene Delivery.

Montassar Khalil, Alexis Hocquigny, Mathieu Berchel, Tristan Montier, Paul-Alain Jaffrès.

  A convergent synthesis of cationic amphiphilic compounds is reported here with the use of the phosphonodithioester-amine coupling (PAC) reaction. This versatile reaction occurs at room temperature without any catalyst, allowing binding of the lipid moiety to a polar head group. This strategy is illustrated with the use of two lipid units featuring either two oleyl chains or two-branched saturated lipid chains. The final cationic amphiphiles were evaluated as carriers for plasmid DNA delivery in four cell lines (A549, Calu3, CFBE and 16HBE) and were compared to standards (BSV36 and KLN47). These new amphiphilic derivatives, which were formulated with DOPE or DOPE-cholesterol as helper lipids, feature high transfection efficacies when associated with DOPE. The highest transfection efficacies were observed in the four cell lines at low charge ratios (CR = 0.7, 1 or 2). At these CRs, no toxic effects were detected. Altogether, this new synthesis scheme using the PAC reaction opens up new possibilities for investigating the effects of lipid or polar head groups on transfection efficacies.

Keywords:  cationic lipids; click reaction; liposomes; nucleic acids delivery; phospholipids

DOI:  https://doi.org/10.3390/molecules26247507
Proc Natl Acad Sci U S A. 2021 Dec 28. pii: e2109256118. [Epub ahead of print]118(52):

On the mechanism of tissue-specific mRNA delivery by selective organ targeting nanoparticles.

Sean A Dilliard, Qiang Cheng, Daniel J Siegwart.

  Lipid nanoparticles (LNPs) are a clinically mature technology for the delivery of genetic medicines but have limited therapeutic applications due to liver accumulation. Recently, our laboratory developed selective organ targeting (SORT) nanoparticles that expand the therapeutic applications of genetic medicines by enabling delivery of messenger RNA (mRNA) and gene editing systems to non-liver tissues. SORT nanoparticles include a supplemental SORT molecule whose chemical structure determines the LNP's tissue-specific activity. To understand how SORT nanoparticles surpass the delivery barrier of liver hepatocyte accumulation, we studied the mechanistic factors which define their organ-targeting properties. We discovered that the chemical nature of the added SORT molecule controlled biodistribution, global/apparent pKa, and serum protein interactions of SORT nanoparticles. Additionally, we provide evidence for an endogenous targeting mechanism whereby organ targeting occurs via 1) desorption of poly(ethylene glycol) lipids from the LNP surface, 2) binding of distinct proteins to the nanoparticle surface because of recognition of exposed SORT molecules, and 3) subsequent interactions between surface-bound proteins and cognate receptors highly expressed in specific tissues. These findings establish a crucial link between the molecular composition of SORT nanoparticles and their unique and precise organ-targeting properties and suggest that the recruitment of specific proteins to a nanoparticle's surface can enable drug delivery beyond the liver.

Keywords:  endogenous targeting; gene editing; lipid nanoparticles; mRNA delivery

DOI:  https://doi.org/10.1073/pnas.2109256118
Biomacromolecules. 2021 Dec 22.

Dynamic Stabilization of Unit Polyion Complexes Incorporating Small Interfering RNA by Fine-Tuning of Cationic Block Length in Two-Branched Poly(ethylene glycol)-b-poly(l-lysine).

Hiroyuki Chaya, Mitsuru Naito, Masaru Cho, Kazuko Toh, Kotaro Hayashi, Shigeto Fukushima, Yuichi Yamasaki, Kazunori Kataoka, Kanjiro Miyata.

To stabilize small interfering RNA (siRNA) in the bloodstream for systemic RNAi therapeutics, we previously fabricated ultrasmall siRNA nanocarriers that were sub-20 nm in hydrodynamic diameter, named as unit polyion complexes (uPICs), using two-branched poly(ethylene glycol)-b-poly(l-lysine) (bPEG-PLys). The blood retention time of uPICs is dramatically increased in the presence of free bPEG-PLys, suggesting dynamic stabilization of uPICs by free bPEG-PLys based on their equilibrium. Herein, we examined how the degree of polymerization of PLys (DPPLys) affected the dynamic stability of uPICs in the bloodstream during prolonged circulation. We prepared a series of bPEG-PLys with DPPLys values of 10, 13, 20, 40, and 80 for the uPIC formation and siRNA with 40 negative charges. These bPEG-PLys were then evaluated in physicochemical characterization and pharmacokinetic analyses. Structural analyses revealed that the uPIC size and association numbers were mainly determined by the molecular weights of PEG and DPPLys, respectively. Under bPEG-PLys-rich conditions, the hydrodynamic diameters of uPICs were 15-20 nm, which were comparable to that of the bPEG block (i.e., ∼18 nm). Importantly, DPPLys significantly affected the association constant of bPEG-PLys to siRNA (Ka) and blood retention of free bPEG-PLys. A smaller DPPLys resulted in a lower Ka and a longer blood retention time of free bPEG-PLys. Thus, DPPLys can control the dynamic stability of uPICs, i.e., the balance between Ka and blood concentration of free bPEG-PLys. Ultimately, the bPEG-PLys with DPPLys values of 14 and 19 prolonged the blood circulation of siRNA-loaded uPICs with relatively small amounts of free bPEG-PLys. This study revealed that the uPIC formation between siRNA and bPEG-PLys can be controlled by their charges, which may be helpful for designing PIC-based delivery systems.

DOI: https://doi.org/10.1021/acs.biomac.1c01344
Biomaterials. 2021 Dec 15. pii: S0142-9612(21)00680-3. [Epub ahead of print]280 121324

Dual targeted delivery of statins and nucleic acids by chitosan-based nanoparticles for enhanced antiatherosclerotic efficacy.

Ting Jiang, Lu Xu, Mengxin Zhao, Fei Kong, Xinrong Lu, Cui Tang, Chunhua Yin.

  Cardiovascular disease caused by atherosclerosis is a leading cause of morbidity and mortality worldwide. Owing to the synergistic regulation of cholesterol metabolism and lesion inflammation, the simultaneous administration of statins and nucleic acids is expected to alleviate atherosclerosis. In this work, we prepared atorvastatin- and galactose-modified trimethyl chitosan nanoparticles (GTANPs) with dual targeting to hepatocytes and lesional macrophages for encapsulating Baf60a siRNA (siBaf60a) and anti-miR-33 pDNA (pAnti-miR-33), attaining the effective codelivery of statins and nucleic acids. We demonstrated that GTANPs/siBaf60a and GTANPs/pAnti-miR-33 had in vitro antiinflammatory and lipid regulating efficacy. In ApoE-knockout atherosclerotic mice, intravenously injected GTANPs/siBaf60a synergistically reduced the plasma cholesterol and atherosclerotic plaque area; more importantly, orally delivered GTANPs/pAnti-miR-33 synergistically increased the levels of plasma high-density lipoprotein cholesterol (HDL-C) and antiinflammatory cytokines, resulting in a satisfactory antiatherosclerotic outcome. Our results suggest that codelivery of statins and nucleic acids provides a promising strategy for the treatment of atherosclerosis.

Keywords:  Atherosclerosis; Atorvastatin; Chitosan-based nanoparticles; Hepatocytes; Lesional macrophages; Nucleic acids

DOI:  https://doi.org/10.1016/j.biomaterials.2021.121324
Nucleic Acid Ther. 2021 Dec 17.

Challenges and Opportunities for Nucleic Acid Therapeutics.

David R Corey, Masad J Damha, Muthiah Manoharan.



Keywords:  antisense; siRNA; therapeutic

DOI:  https://doi.org/10.1089/nat.2021.0085
Front Plant Sci. 2021 ;12 759871

Imaging of the Entry Pathway of a Cell-Penetrating Peptide-DNA Complex From the Extracellular Space to Chloroplast Nucleoids Across Multiple Membranes in Arabidopsis Leaves.

Kazusato Oikawa, Ayaka Tateishi, Masaki Odahara, Yutaka Kodama, Keiji Numata.

  Each plant cell has hundreds of copies of the chloroplast genome and chloroplast transgenes do not undergo silencing. Therefore, chloroplast transformation has many powerful potential agricultural and industrial applications. We previously succeeded in integrating exogenous genes into the chloroplast genome using peptide-DNA complexes composed of plasmid DNA and a fusion peptide consisting of a cell-penetrating peptide (CPP) and a chloroplast transit peptide (cpPD complex). However, how cpPD complexes are transported into the chloroplast from outside the cell remains unclear. Here, to characterize the route by which these cpPD complexes move into chloroplasts, we tracked their movement from the extracellular space to the chloroplast stroma using a fluorescent label and confocal laser scanning microscopy (CLSM). Upon infiltration of cpPD complexes into the extracellular space of Arabidopsis thaliana leaves, the complexes reached the chloroplast surface within 6h. The cpPD complexes reached were engulfed by the chloroplast outer envelope membrane and gradually integrated into the chloroplast. We detected several cpPD complexes localized around chloroplast nucleoids and observed the release of DNA from the cpPD. Our results thus define the route taken by the cpPD complexes for gene delivery from the extracellular space to the chloroplast stroma.

Keywords:  Arabidopsis thaliana; DNA delivery; cell wall; cell-penetrating peptide; chloroplast nucleoid; confocal laser scanning microscopy; field emission-scanning electron microscope; plasma membrane

DOI:  https://doi.org/10.3389/fpls.2021.759871
Adv Sci (Weinh). 2021 Dec 22. e2103812

Anti-MicroRNA-21 Oligonucleotide Loaded Spermine-Modified Acetalated Dextran Nanoparticles for B1 Receptor-Targeted Gene Therapy and Antiangiogenesis Therapy.

Tao Zheng, Wentao Wang, Mohsen Mohammadniaei, Jon Ashley, Ming Zhang, Ninglin Zhou, Jian Shen, Yi Sun.

  The use of nanoparticles (NPs) to deliver small inhibiting microRNAs (miRNAs) has shown great promise for treating cancer. However, constructing a miRNA delivery system that targets brain cancers, such as glioblastoma multiforme (GBM), remains technically challenging due to the existence of the blood-tumor barrier (BTB). In this work, a novel targeted antisense miRNA-21 oligonucleotide (ATMO-21) delivery system is developed for GBM treatment. Bradykinin ligand agonist-decorated spermine-modified acetalated dextran NPs (SpAcDex NPs) could temporarily open the BTB by activating G-protein-coupled receptors that are expressed in tumor blood vessels and tumor cells, which increase transportation to and accumulation in tumor sites. ATMO-21 achieves high loading in the SpAcDex NPs (over 90%) and undergoes gradual controlled release with the degradation of the NPs in acidic lysosomal compartments. This allows for cell apoptosis and inhibition of the expression of vascular endothelial growth factor by downregulating hypoxia-inducible factor (HIF-1α) protein. An in vivo orthotopic U87MG glioma model confirms that the released ATMO-21 shows significant therapeutic efficacy in inhibiting tumor growth and angiogenesis, demonstrating that agonist-modified SpAcDex NPs represent a promising strategy for GBM treatment combining targeted gene therapy and antiangiogenic therapy.

Keywords:  anti-microRNA-21 oligonucleotide; antiangiogenesis therapy; gene therapy; glioblastoma multiforme; targeted delivery

DOI:  https://doi.org/10.1002/advs.202103812
J Control Release. 2021 Dec 21. pii: S0168-3659(21)00673-8. [Epub ahead of print]

Insights into nucleic acid-based self-assembling nanocarriers for targeted drug delivery and controlled drug release.

Keren Chen, Yangzi Zhang, Longjiao Zhu, Huashuo Chu, Kunlun Huang, Xiangli Shao, Charles Asakiya, Kunlun Huang, Wentao Xu.

  Over the past few decades, rapid advances of nucleic acid nanotechnology always drive the development of nanoassemblies with programmable design, powerful functionality, excellent biocompatibility and outstanding biosafety. Nowadays, nucleic acid-based self-assembling nanocarriers (NASNs) play an increasingly greater role in the research and development in biomedical studies, particularly in drug delivery, release and targeting. In this review, NASNs are systematically summarized the strategies cooperated with their broad applications in drug delivery. We first discuss the self-assembling methods of nanocarriers comprised of DNA, RNA and composite materials, and summarize various categories of targeting media, including aptamers, small molecule ligands and proteins. Furthermore, drug release strategies by smart-responding multiple kinds of stimuli are explained, and various applications of NASNs in drug delivery are discussed, including protein drugs, nucleic acid drugs, small molecule drugs and nanodrugs. Lastly, we propose limitations and potential of NASNs in the future development, and expect that NASNs enable facilitate the development of new-generation drug vectors to assist in solving the growing demands on disease diagnosis and therapy or other biomedicine-related applications in the real world.

Keywords:  Drug delivery; Nucleic acid-based self-assembling nanocarriers; Stimulation response modes; Target recognition

DOI:  https://doi.org/10.1016/j.jconrel.2021.12.020