bims-novged 2022-05-08 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2022–05–08
thirteen papers selected by
the Merkel lab, Ludwig-Maximilians University

pH-sensitive zwitterionic polycarboxybetaine as a potential non-viral vector for small interfering RNA delivery.
Regulation of microglia polarization via mannose receptor-mediated delivery of siRNA by ligand-functionalized DoGo LNP.
Linear Poly(ethylenimine-propylenimine) Random Copolymers for Gene Delivery: From Polymer Synthesis to Efficient Transfection with High Serum Tolerance.
Aerosol delivery of star polymer-siRNA nanoparticles as a therapeutic strategy to inhibit lung tumor growth.
Efficient gene transfection to lung cancer cells via Folate-PEI-Sorbitol gene transporter.
Zn-Dipicolylamine-based Reactive Oxygen Species-Responsive Lipids for siRNA Delivery and in vivo Colitis Treatment.
Targeted delivery to macrophages and dendritic cells by chemically modified mannose ligand-conjugated siRNA.
Study on lipid nanomicelles targeting placenta for the treatment of preeclampsia.
A Lysosome-activated Tetrahedral Nanobox for Encapsulated siRNA Delivery.
Dual Inhibitions on Glucose/Glutamine Metabolisms for Nontoxic Pancreatic Cancer Therapy.
Surface-decorated nanoparticles clicked into nanoparticle clusters for oligonucleotide encapsulation.
Synchronous Disintegration of Ferroptosis Defense Axis via Engineered Exosome-Conjugated Magnetic Nanoparticles for Glioblastoma Therapy.
Cholesterol-Mediated Seeding of Protein Corona on DNA Nanostructures for Targeted Delivery of Oligonucleotide Therapeutics to Treat Liver Fibrosis.

RSC Adv. 2020 Dec 17. 10(73): 45059-45066

pH-sensitive zwitterionic polycarboxybetaine as a potential non-viral vector for small interfering RNA delivery.

Huan Peng, Weihong Ji, Ruichen Zhao, Zhiguo Lu, Jun Yang, Yan Li, Xin Zhang.

Small interfering RNA (siRNA) has great potential for the treatment of various diseases. However, its intrinsic deficiencies seriously limit its application. Herein, pH-sensitive zwitterionic polymer polycarboxybetaine (PCB) was developed as a non-viral vector for siRNA. The PCB could be protonated in an acidic environment and become positively charged from a cancer site. After protonation, PCB could complex siRNA via electrostatic interaction, and its loading ability was enhanced with a decrease of pH value. Compared with the PEI 10k, PCB50 with a similar molecular weight had comparable siRNA loading ability and lower cytotoxicity. Besides, siRNA loaded by PCB50 could escape from endosomes and reduce the loss of drugs, and based on the excellent uptake and obvious apoptotic effect on HeLa cells, the pH-sensitive PCB with low cytotoxicity could be used as a non-viral vector for safe siRNA delivery for cancer treatment.

DOI: https://doi.org/10.1039/d0ra09359a
RSC Adv. 2021 Oct 04. 11(52): 32549-32558

Regulation of microglia polarization via mannose receptor-mediated delivery of siRNA by ligand-functionalized DoGo LNP.

Hai Xiao, Shuqin Han, Huricha Baigude.

The pro-inflammatory polarization of microglia after stroke is one of the major causes of secondary brain injury. Downregulation of the gene involved in canonical inflammatory pathways in glial cells can exert neuroprotective effects via inhibiting the release of pro-inflammatory factors. In this study, we functionalized DoGo lipids with mannose, the ligand of the mannose receptor (MR) that is expressed in microglia, and evaluated the MR-mediated cellular internalization of DoGo lipid nanoparticles (denote M3) carrying siRNA against TLR4 in BV2 cells in vitro. We confirmed that siTLR4/M3 complexes were specifically internalized by BV2 cells in a MR-dependent manner, and the treatment of oxygen glucose deprivation (OGD)-treated BV2 cells with siTLR4/M3 complexes resulted in remarkable silencing of TLR4, and induced downregulated M1 polarization and upregulated M2 polarization markers. Collectively, our data suggest that the M3 lipoplex is a promising microglia-targeting siRNA delivery agent.

DOI: https://doi.org/10.1039/d1ra04293a
Biomacromolecules. 2022 May 02.

Linear Poly(ethylenimine-propylenimine) Random Copolymers for Gene Delivery: From Polymer Synthesis to Efficient Transfection with High Serum Tolerance.

M Rachèl Elzes, Ine Mertens, Ondrej Sedlacek, Bart Verbraeken, Aniek C A Doensen, Maarten A Mees, Mathias Glassner, Somdeb Jana, Jos M J Paulusse, Richard Hoogenboom.

Naturally occurring oligoamines, such as spermine, spermidine, and putrescine, are well-known regulators of gene expression. These oligoamines frequently have short alkyl spacers with varying lengths between the amines. Linear polyethylenimine (PEI) is a polyamine that has been widely applied as a gene vector, with various formulations currently in clinical trials. In order to emulate natural oligoamine gene regulators, linear random copolymers containing both PEI and polypropylenimine (PPI) repeat units were designed as novel gene delivery agents. In general, statistical copolymerization of 2-oxazolines and 2-oxazines leads to the formation of gradient copolymers. In this study, however, we describe for the first time the synthesis of near-ideal random 2-oxazoline/2-oxazine copolymers through careful tuning of the monomer structures and reactivity as well as polymerization conditions. These copolymers were then transformed into near-random PEI-PPI copolymers by controlled side-chain hydrolysis. The prepared PEI-PPI copolymers formed stable polyplexes with GFP-encoding plasmid DNA, as validated by dynamic light scattering. Furthermore, the cytotoxicity and transfection efficiency of polyplexes were evaluated in C2C12 mouse myoblasts. While the polymer chain length did not significantly increase the toxicity, a higher PPI content was associated with increased toxicity and also lowered the amount of polymers needed to achieve efficient transfection. The transfection efficiency was significantly influenced by the degree of polymerization of PEI-PPI, whereby longer polymers resulted in more transfected cells. Copolymers with 60% or lower PPI content exhibited a good balance between high plasmid-DNA transfection efficiency and low toxicity. Interestingly, these novel PEI-PPI copolymers revealed exceptional serum tolerance, whereby transfection efficiencies of up to 53% of transfected cells were achieved even under 50% serum conditions. These copolymers, especially PEI-PPI with DP500 and a 1:1 PEI/PPI ratio, were identified as promising transfection agents for plasmid DNA.

DOI: https://doi.org/10.1021/acs.biomac.2c00210
Biomaterials. 2022 Apr 23. pii: S0142-9612(22)00179-X. [Epub ahead of print]285 121539

Aerosol delivery of star polymer-siRNA nanoparticles as a therapeutic strategy to inhibit lung tumor growth.

Z Ma, S W Wong, H Forgham, L Esser, M Lai, M N Leiske, K Kempe, G Sharbeen, J Youkhana, F Mansfeld, J F Quinn, P A Phillips, T P Davis, M Kavallaris, J A McCarroll.

  Lung cancer is a major contributor to cancer-related death worldwide. siRNA nanomedicines are powerful tools for cancer therapeutics. However, there are challenges to overcome to increase siRNA delivery to solid tumors, including penetration of nanoparticles into a complex microenvironment following systemic delivery while avoiding rapid clearance by the reticuloendothelial system, and limited siRNA release from endosomes once inside the cell. Here we characterized cell uptake, intracellular trafficking, and gene silencing activity of miktoarm star polymer (PDMAEMA-POEGMA) nanoparticles (star nanoparticles) complexed to siRNA in lung cancer cells. We investigated the potential of nebulized star-siRNA nanoparticles to accumulate into orthotopic mouse lung tumors to inhibit expression of two genes [βIII-tubulin, Polo-Like Kinase 1 (PLK1)] which: 1) are upregulated in lung cancer cells; 2) promote tumor growth; and 3) are difficult to inhibit using chemical drugs. Star-siRNA nanoparticles internalized into lung cancer cells and escaped the endo-lysosomal pathway to inhibit target gene expression in lung cancer cells in vitro. Nebulized star-siRNA nanoparticles accumulated into lungs and silenced the expression of βIII-tubulin and PLK1 in mouse lung tumors, delaying aggressive tumor growth. These results demonstrate a proof-of-concept for aerosol delivery of star-siRNA nanoparticles as a novel therapeutic strategy to inhibit lung tumor growth.

Keywords:  Aerosol drug delivery; Lung cancer; PLK1; Star polymers; siRNA; βIII-tubulin

DOI:  https://doi.org/10.1016/j.biomaterials.2022.121539
PLoS One. 2022 ;17(5): e0266181

Efficient gene transfection to lung cancer cells via Folate-PEI-Sorbitol gene transporter.

Kye Soo Cho, Sanghwa Kim, Hyung Bin Chun, Jae Hee Cheon, Myung-Haing Cho, Ah Young Lee, Rohidas B Arote.

Lung cancer is known to be one of the fatal diseases in the world and is experiencing treatment difficulties. Many treatments have been discovered and implemented, but death rate of patients with lung cancer continues to remain high. Current treatments for cancer such as chemotherapy, immunotherapy, and radiotherapy have shown considerable results, yet they are accompanied by side effects. One effective method for reducing the cytotoxicity of these treatments is via the use of a nanoparticle-mediated siRNA delivery strategy with selective silencing effects and non-viral vectors. In this study, a folate (FA) moiety ligand-conjugated poly(sorbitol-co-PEI)-based gene transporter was designed by combining low-molecular weight polyethyleneimine (LMW PEI) and D-sorbitol with FA to form FPS. Since folate receptors are commonly overexpressed in various cancer cells, folate-conjugated nanoparticles may be more effectively delivered to selective cancer cells. Additionally, siOPA1 was used to induce apoptosis through mitochondrial fusion. The OPA1 protein stability level is important for maintaining normal mitochondrial cristae structure and function, conserving the inner membrane structure, and protecting cells from apoptosis. Consequently, when FPS/siOPA1 was used for lung cancer in-vitro and in-vivo, it improved cell viability and cellular uptake.

DOI: https://doi.org/10.1371/journal.pone.0266181
Acta Biomater. 2022 Apr 27. pii: S1742-7061(22)00242-2. [Epub ahead of print]

Zn-Dipicolylamine-based Reactive Oxygen Species-Responsive Lipids for siRNA Delivery and in vivo Colitis Treatment.

Yu Guo, Xi He, Rui-Mo Zhao, Hui-Zhen Yang, Zheng Huang, Ji Zhang, Xiao-Qi Yu.

  The reduction of ROS and inflammatory factor levels plays important role in the treatment of colitis. A series of ROS-responsive lipids (ZnDPA-R) based on the thioketal structure were designed and synthesized. It was hoped that the lipidic materials could combine ROS consumption and siRNA delivery capacity to achieve synergistic treatment of colitis. The target liposomes could combine with the phosphate backbone of siRNA to form lipoplexes with the size of ∼100 nm, and could deliver siRNA cargo into the cell. The results of in vitro anti-inflammatory experiments showed that the lipids may effectively consume ROS in cells. Meanwhile, the lipoplexes significantly reduced the expression levels of TNF-α mRNA and related inflammatory factors in macrophages. After PEGylation, the lipoplex was used for the treatment of mouse colitis, and biodistribution results proved that the lipoplexes effectively aggregated in the intestine. The delivery system could not only response to the high ROS level at colitis via thioketal breaking, but also could assist in the treatment of inflammation by ROS consumption. The treatment results revealed that the levels of TNF-α mRNA and related inflammatory factors at the colon lesion were largely reduced, and the inflammatory symptoms were significantly relieved. Hematology test results indicated that the treatment was safe and induced no obvious side effects on mice. This study may shed light on the synergistic treatment for colitis via anti-inflammatory siRNA delivery and ROS depletion strategies. STATEMENT OF SIGNIFICANCE: Downregulation of inflammatory factors and reactive oxygen species (ROS) levels is critical in treating colitis. In the present study, a series of ROS-responsive lipid molecules based on the Zn-DPA headgroup and thioketal linkage were synthesized for delivering TNF-α siRNA and for treating colitis. In addition to silencing the expression of TNF-α mRNA and the related inflammatory factors, the material also achieved synergistic treatment by simultaneous consumption of ROS in the colon lesion. In vitro cell experiments and in vivo colitis treatment in mice showed that the lipoplex exerted a satisfactory therapeutic effect on colitis, and the symptoms of colitis in mice were significantly alleviated. The present study may shed light on the synergistic treatment for colitis through anti-inflammatory siRNA delivery and ROS depletion strategies.

Keywords:  Cationic lipids; ROS responsiveness; colitis; siRNA delivery; synergistic therapy

DOI:  https://doi.org/10.1016/j.actbio.2022.04.033
Nucleic Acids Res. 2022 May 07. pii: gkac308. [Epub ahead of print]

Targeted delivery to macrophages and dendritic cells by chemically modified mannose ligand-conjugated siRNA.

Keiji Uehara, Toshimasa Harumoto, Asana Makino, Yasuo Koda, Junko Iwano, Yasuhiro Suzuki, Mari Tanigawa, Hiroto Iwai, Kana Asano, Kana Kurihara, Akinori Hamaguchi, Hiroshi Kodaira, Toshiyuki Atsumi, Yoji Yamada, Kazuma Tomizuka.

Extrahepatic delivery of small interfering RNAs (siRNAs) may have applications in the development of novel therapeutic approaches. However, reports on such approaches are limited, and the scarcity of reports concerning the systemically targeted delivery of siRNAs with effective gene silencing activity presents a challenge. We herein report for the first time the targeted delivery of CD206-targetable chemically modified mannose-siRNA (CMM-siRNA) conjugates to macrophages and dendritic cells (DCs). CMM-siRNA exhibited a strong binding ability to CD206 and selectively delivered contents to CD206-expressing macrophages and DCs. Furthermore, the conjugates demonstrated strong gene silencing ability with long-lasting effects and protein downregulation in CD206-expressing cells in vivo. These findings could broaden the use of siRNA technology, provide additional therapeutic opportunities, and establish a basis for further innovative approaches for the targeted delivery of siRNAs to not only macrophages and DCs but also other cell types.

DOI: https://doi.org/10.1093/nar/gkac308
J Drug Target. 2022 May 03. 1-36

Study on lipid nanomicelles targeting placenta for the treatment of preeclampsia.

Yang Liu, Qimeng Zhang, Xingli Gao, Tong Wang.

  In view of the serious clinical harm of preeclampsia and the lack of effective treatment methods, a PEG-modified lipid hybrid micelle was designed with folic acid molecule on the surface, containing siRNA, targeted delivery to the placenta, interfering the expression of sFlt-1 and treating preeclampsia. In this paper, the preparation and characterization of lipid hybrid micelles were investigated in detail, the cytology in vitro and in vivo distribution, pharmacodynamics, safety and action mechanism of the preparation were studied, which laid a foundation for gene therapy of preeclampsia.

Keywords:  micelle; placenta; preeclampsia; sFlt-1; siRNA

DOI:  https://doi.org/10.1080/1061186X.2022.2068558
Adv Mater. 2022 May 05. e2201731

A Lysosome-activated Tetrahedral Nanobox for Encapsulated siRNA Delivery.

Yang Gao, Xingyu Chen, Taoran Tian, Tao Zhang, Shaojingya Gao, Xiaolin Zhang, Yangxue Yao, Yunfeng Lin, Xiaoxiao Cai.

  Tetrahedral framework nucleic acids (tFNAs) have attracted extensive attention as drug nanocarriers because of their excellent cellular uptake. However, for oligonucleotide cargos, tFNA mainly acts as a static delivery platform generated via sticky-ended ligation. Here, inspired by the original stable space inside the tetrahedral scaffold, we fabricate a dynamic lysosome-activated tFNA nanobox for completely encapsulating a short interfering RNA (siRNA) of interest. The closed tetrahedral structure endows cargo siRNA with greater resistance against RNase and serum and enables solid integration with the vehicle during delivery. Moreover, the pH-responsive switch of nanobox allows the controlled release of siRNA upon entry into lysosomes at cell culture temperature. Based on protective loading and active unloading, an excellent silencing effect on the target TNFα gene was achieved in vitro and in vivo experiments. Conclusively, nanobox offers a dynamic pH-sensitive confinement delivery system for siRNA and can be an extendable strategy for other small RNA. This article is protected by copyright. All rights reserved.

Keywords:  DNA nanotechnology; framework nucleic acids; lysosome activating; pH responsiveness; siRNA delivery

DOI:  https://doi.org/10.1002/adma.202201731
ACS Appl Mater Interfaces. 2022 May 05.

Dual Inhibitions on Glucose/Glutamine Metabolisms for Nontoxic Pancreatic Cancer Therapy.

Yingying Xu, Zhiguo Yu, Hao Fu, Yuedong Guo, Ping Hu, Jianlin Shi.

  Glucose and glutamine are two principal nutrients in mammalian cells that provide energy and biomass for cell growth and proliferation. Especially in cancer cells, glutamine could be a main alternative for energy and biomass supply once glucose metabolism is suppressed. Therefore, single inhibition of enzymes in either glucose metabolism or glutaminolysis, though maybe efficient in vitro, is far from being satisfactory for efficient in vivo cancer therapy. Here, we proposed a new strategy for dual inhibitions on both glucose and glutamine metabolisms concurrently by silencing mutated gene Kras and glutaminase 1 (GLS1) via nanomaterial-based siKras and siGLS1 delivery, rather than conventional highly toxic chemodrugs. Such a combination therapy could overcome the challenge that glucose and glutamine are alternatives to each other in the biosynthesis and energy production for cancer cells, resulting in much elevated treatment efficacy. In addition, layered double hydroxide (LDH), the siRNA carrier, enables an enhanced gene delivery efficiency compared to the commercial transfection agent Lipofectamine 2000. Briefly, Mg-Al LDH nanosheets, loaded with siKras and siGLS1 onto their surfaces by electrostatic adsorption, could release siRNA from lysosomes into the cytoplasm via the proton sponge effect of LDH, favoring the siRNA stability and gene silencing efficiency enhancements. The thus released siRNA could downregulate the expressions of Kras, GLS1, and other enzymes involved in glucose metabolism, resulting in the downregulations of ATP and other metabolites. Such a biosafe LDH/siRNA nanomedicine is able to efficiently suppress the growth of xenografts through cancer cell proliferation suppression, displaying its great potential as a simultaneous glucose/glutamine metabolism coinhibitor for treating pancreatic cancer.

Keywords:  GLS1; Kras; glucose metabolism inhibition; glutamine metabolism inhibition; layered double hydroxide nanoparticles; pancreatic cancer; siRNA

DOI:  https://doi.org/10.1021/acsami.2c00111
RSC Adv. 2020 Oct 07. 10(61): 37040-37049

Surface-decorated nanoparticles clicked into nanoparticle clusters for oligonucleotide encapsulation.

Wei Mao, Song Rae Kim, Hyuk Sang Yoo.

Gold nanoparticles (AuNPs) are the predominant and representative metal nano-carriers used for the tumor-targeted delivery of therapeutics because they possess advantages such as biocompatibility, high drug loading efficiency, and enhanced accumulation at tumor sites via the size-dependent enhanced permeability and retention (EPR) effect. In this study, we designed an AuNP functionalized with block polymers comprising polyethylenimine and azide group-functionalized poly(ethyl glycol) for the electrostatic incorporation of cytosine-guanine oligonucleotide (CpG ODN) on the surface. The ODN-incorporated AuNPs were cross-linked to gold nanoparticle clusters (AuNCs) via click chemistry using a matrix metalloproteinase (MMP)-2 cleavable peptide linker modified with alkyne groups at both ends. In the presence of Cu(i), azide groups and alkyne groups spontaneously cyclize to form a triazole ring with high fidelity and efficiency, and therefore allow single AuNPs to stack to larger AuNCs for increased EPR effect-mediated tumor targeting. 1H-NMR and Fourier transform infrared spectroscopy revealed the successful synthesis of an azide-PEG-grafted branched polyethylenimine, and the size and morphology of AuNPs fabricated by the synthesized polymer were confirmed to be 4.02 ± 0.45 nm by field emission-transmission electron microscopy. Raman spectroscopy characterization demonstrated the introduction of azide groups on the surface of the synthesized AuNPs. Zeta-potential and gel-retardation analysis of CpG-loaded AuNPs indicated complete CpG sequestration by AuNPs when the CpG : AuNP weight ratio was higher than 1 : 2.5. The clustering process of the CpG-loaded AuNPs was monitored and was demonstrated to be dependent on the alkyne linker-to-AuNP ratio. Thus, the clicked AuNC can be tailored as a gene carrier where a high accumulation of therapeutics is required.

DOI: https://doi.org/10.1039/d0ra06622b
Adv Sci (Weinh). 2022 May 04. e2105451

Synchronous Disintegration of Ferroptosis Defense Axis via Engineered Exosome-Conjugated Magnetic Nanoparticles for Glioblastoma Therapy.

Boyan Li, Xin Chen, Wei Qiu, Rongrong Zhao, Jiazhi Duan, Shouji Zhang, Ziwen Pan, Shulin Zhao, Qindong Guo, Yanhua Qi, Wenhan Wang, Lin Deng, Shilei Ni, Yuanhua Sang, Hao Xue, Hong Liu, Gang Li.

  Glioblastoma (GBM) is one of the most fatal central nervous system tumors and lacks effective or sufficient therapies. Ferroptosis is a newly discovered method of programmed cell death and opens a new direction for GBM treatment. However, poor blood-brain barrier (BBB) penetration, reduced tumor targeting ability, and potential compensatory mechanisms hinder the effectiveness of ferroptosis agents during GBM treatment. Here, a novel composite therapeutic platform combining the magnetic targeting features and drug delivery properties of magnetic nanoparticles with the BBB penetration abilities and siRNA encapsulation properties of engineered exosomes for GBM therapy is presented. This platform can be enriched in the brain under local magnetic localization and angiopep-2 peptide-modified engineered exosomes can trigger transcytosis, allowing the particles to cross the BBB and target GBM cells by recognizing the LRP-1 receptor. Synergistic ferroptosis therapy of GBM is achieved by the combined triple actions of the disintegration of dihydroorotate dehydrogenase and the glutathione peroxidase 4 ferroptosis defense axis with Fe3 O4 nanoparticle-mediated Fe2+ release. Thus, the present findings show that this system can serve as a promising platform for the treatment of glioblastoma.

Keywords:  blood-brain barrier; exosomes; ferroptosis; glioblastoma; magnetic nanoparticles

DOI:  https://doi.org/10.1002/advs.202105451
ACS Nano. 2022 May 02.

Cholesterol-Mediated Seeding of Protein Corona on DNA Nanostructures for Targeted Delivery of Oligonucleotide Therapeutics to Treat Liver Fibrosis.

Kyoung-Ran Kim, Junghyun Kim, Ji Hyun Back, Ji Eun Lee, Dae-Ro Ahn.

  The protein corona is a protein layer formed on the surface of nanoparticles administered in vivo and considerably affects the in vivo fate of nanoparticles. Although it is challenging to control protein adsorption on nanoparticles precisely, the protein corona may be harnessed to develop a targeted drug delivery system if the nanoparticles are decorated with a ligand with enhanced affinity to target tissue- and cell-homing proteins. Here, we prepared a DNA tetrahedron with trivalent cholesterol conjugation (Chol3-Td) that can induce enhanced interaction with lipoproteins in serum, which in situ generates the lipoprotein-associated protein corona on a DNA nanostructure favorable for cells abundantly expressing lipoprotein receptors in the liver, such as hepatocytes in healthy mice and myofibroblasts in fibrotic mice. Chol3-Td was further adopted for liver delivery of antisense oligonucleotide (ASO) targeting TGF-β1 mRNA to treat liver fibrosis in a mouse model. The potency of ASO@Chol3-Td was comparable to that of ASO conjugated with the clinically approved liver-targeting ligand, trivalent N-acetylgalactosamine (GalNAc3), demonstrating the potential of Chol3-Td as a targeted delivery system for oligonucleotide therapeutics. This study suggests that controlled seeding of the protein corona on nanomaterials can provide a way to steer nanoparticles into the target area.

Keywords:  DNA tetrahedron; cholesterol conjugation; liver delivery; oligonucleotide therapeutics; protein corona; targeted delivery

DOI:  https://doi.org/10.1021/acsnano.1c08508