bims-novged 2021-05-02 papers

on Non-viral vectors for gene delivery

Issue of 2021–05–02
seven papers selected by
the Merkel lab, Ludwig-Maximilians University and Benjamin Winkeljann, Ludwig-Maximilians University

Caveolae-Mediated Endocytosis Drives Robust siRNA Delivery of Polymeric Nanoparticles to Macrophages.
Controllable assembly/disassembly of polyphenol-DNA nanocomplex for cascade-responsive drug release in cancer cells.
Single-cell analysis reveals effective siRNA delivery in brain tumors with microbubble-enhanced ultrasound and cationic nanoparticles.
Delivery of Oligonucleotides Using a Self-Degradable Lipid-Like Material.
Viscous Core Liposomes Increase siRNA Encapsulation and Provides Gene Inhibition When Slightly Positively Charged.
Lipid Nanoparticle-Mediated Lymphatic Delivery of Immunostimulatory Nucleic Acids.
Amino acids functionalized dendrimers with nucleus accumulation for efficient gene delivery.

ACS Nano. 2021 Apr 29.

Caveolae-Mediated Endocytosis Drives Robust siRNA Delivery of Polymeric Nanoparticles to Macrophages.

Yudong Song, Yanhua Wu, Lu Xu, Ting Jiang, Cui Tang, Chunhua Yin.

  Cytosolic delivery of small interfering RNA (siRNA) remains challenging, and a profound understanding of the cellular uptake and intracellular processing of siRNA delivery systems could greatly improve the development of siRNA-based therapeutics. Here, we show that caveolae-mediated endocytosis (CvME) accounts for the robust siRNA delivery of mannose-modified trimethyl chitosan-cysteine/tripolyphosphate nanoparticles (MTC/TPP NPs) to macrophages by circumventing lysosomes. We show that the Golgi complex and ER are key organelles required for the efficient delivery of siRNA to macrophages in which the siRNA accumulation positively correlates with its silencing efficiency (r = 0.94). We also identify syntaxin6 and Niemann-Pick type C1 (NPC1) as indispensable regulators for MTC/TPP NPs-delivered siRNA into macrophages both in vitro and in vivo. Syntaxin6 and NPC1 knockout substantially decrease the cellular uptake and gene silencing of the siRNA delivered in MTC/TPP NPs in macrophages, which result in poor therapeutic outcomes for mice bearing acute hepatic injury. Our results suggest that highly efficient siRNA delivery can be achieved via CvME, which would give ideas for designing optimal delivery vectors to facilitate the clinical translation of siRNA drugs.

Keywords:  CvME; NPC1; Syntaxin6; anti-inflammatory efficacy; cytosolic delivery; key organelles; siRNA drugs

DOI:  https://doi.org/10.1021/acsnano.0c08596
Biomaterials. 2021 Apr 23. pii: S0142-9612(21)00202-7. [Epub ahead of print]273 120846

Controllable assembly/disassembly of polyphenol-DNA nanocomplex for cascade-responsive drug release in cancer cells.

Jinpeng Han, Yuchen Cui, Zi Gu, Dayong Yang.

  Developing nanocarrier systems with sufficient drug loading ability and efficient drug release behavior in cells is a powerful strategy to maximize therapeutic efficacies and minimize side effects of administered drugs. However, the two aspects are usually contradictory in a single nanocarrier. Herein, polyphenol-DNA nanocomplex with controllable assembly/disassembly behaviors is developed for responsive and sequential drug release in cancer cells. Programmable assembly of branched-DNA achieves multiple-gene loading, afterwards tannic acid (TA), plant-derived polyphenols as drugs mediate assembly of branched-DNA to form nanocomplex. Intracellularly, two-step disassembly process of nanocomplex enables efficient gene/drug release. Lysosomal acidic microenvironment induces the disassembly of nanocomplex to release TA and branched-DNA. Glutathione and DNase I in cytoplasm trigger the precise release of genes from branched-DNA. The efficacy of multiple-gene/chemo-therapy is demonstrated using in vitro and in vivo models. This work provides a controllable assembly/disassembly route to resolve the conflict between sufficient drug loading and efficient drug release in cells for therapeutics.

Keywords:  DNA nanostructures; DNA nanotechnology; Gene therapy; Natural polyphenol; Self-assembly

DOI:  https://doi.org/10.1016/j.biomaterials.2021.120846
Sci Adv. 2021 Apr;pii: eabf7390. [Epub ahead of print]7(18):

Single-cell analysis reveals effective siRNA delivery in brain tumors with microbubble-enhanced ultrasound and cationic nanoparticles.

Yutong Guo, Hohyun Lee, Zhou Fang, Anastasia Velalopoulou, Jinhwan Kim, Midhun Ben Thomas, Jingbo Liu, Ryan G Abramowitz, YongTae Kim, Ahmet F Coskun, Daniel Pomeranz Krummel, Soma Sengupta, Tobey J MacDonald, Costas Arvanitis.

RNA-based therapies offer unique advantages for treating brain tumors. However, tumor penetrance and uptake are hampered by RNA therapeutic size, charge, and need to be "packaged" in large carriers to improve bioavailability. Here, we have examined delivery of siRNA, packaged in 50-nm cationic lipid-polymer hybrid nanoparticles (LPHs:siRNA), combined with microbubble-enhanced focused ultrasound (MB-FUS) in pediatric and adult preclinical brain tumor models. Using single-cell image analysis, we show that MB-FUS in combination with LPHs:siRNA leads to more than 10-fold improvement in siRNA delivery into brain tumor microenvironments of the two models. MB-FUS delivery of Smoothened (SMO) targeting siRNAs reduces SMO protein production and markedly increases tumor cell death in the SMO-activated medulloblastoma model. Moreover, our analysis reveals that MB-FUS and nanoparticle properties can be optimized to maximize delivery in the brain tumor microenvironment, thereby serving as a platform for developing next-generation tunable delivery systems for RNA-based therapy in brain tumors.

DOI: https://doi.org/10.1126/sciadv.abf7390
Pharmaceutics. 2021 Apr 13. pii: 544. [Epub ahead of print]13(4):

Delivery of Oligonucleotides Using a Self-Degradable Lipid-Like Material.

Hiroki Tanaka, Nae Takata, Yu Sakurai, Tokuyuki Yoshida, Takao Inoue, Shinya Tamagawa, Yuta Nakai, Kota Tange, Hiroki Yoshioka, Masatoshi Maeki, Manabu Tokeshi, Hidetaka Akita.

  The world-first success of lipid nanoparticle (LNP)-based siRNA therapeutics (ONPATTRO®) promises to accelerate developments in siRNA therapeutics/gene therapy using LNP-type drug delivery systems (DDS). In this study, we explore the optimal composition of an LNP containing a self-degradable material (ssPalmO-Phe) for the delivery of oligonucleotides. siRNA or antisense oligonucleotides (ASO) were encapsulated in LNP with different lipid compositions. The hepatic knockdown efficiency of the target genes and liver toxicity were evaluated. The optimal compositions for the siRNA were different from those for ASO, and different from those for mRNA that were reported in a previous study. Extracellular stability, endosomal escape and cellular uptake appear to be the key processes for the successful delivery of mRNA, siRNA and ASO, respectively. Moreover, the compositions of the LNPs likely contribute to their toxicity. The lipid composition of the LNP needs to be optimized depending on the type of nucleic acids under consideration if the applications of LNPs are to be further expanded.

Keywords:  antisense oligonucleotide; lipid nanoparticle; siRNA

DOI:  https://doi.org/10.3390/pharmaceutics13040544
Pharmaceutics. 2021 Apr 01. pii: 479. [Epub ahead of print]13(4):

Viscous Core Liposomes Increase siRNA Encapsulation and Provides Gene Inhibition When Slightly Positively Charged.

Shayan Ahmed, Hugo Salmon, Nicholas Distasio, Hai Doan Do, Daniel Scherman, Khair Alhareth, Maryam Tabrizian, Nathalie Mignet.

  Since its discovery, evidence that siRNA was able to act as an RNA interference effector, led to its acceptation as a novel medicine. The siRNA approach is very effective, due to its catalytic mechanism, but still the limitations of its cellular delivery should be addressed. One promising form of non-viral gene delivery system is liposomes. The variable and versatile nature of the lipids keeps the possibility to upgrade the liposomal structure, which makes them suitable for encapsulation and delivery of drugs. However, to avoid the limitation of fast release for the hydrophilic drug, we previously designed viscous core liposomes. We aimed in this work to evaluate if these viscous core liposomes (NvcLs) could be of interest for siRNA encapsulation. Then, we sought to add a limited amount of positive charges to provide cell interaction and transfection. Cationic lipid dimyristoylaminopropylaminopropyl or the polymer poly(ethylenimine) were incorporated in NvcL to produce positively charged viscous core liposomes (PvcL) by a customized microfluidic device. We found that NvcLs increased the encapsulation efficiency and loading content with regards to the neutral liposome. Both PvcLPEI and PvcLDMAPAP exhibited transfection and GFP knock-down (≈40%) in both 2D and 3D cell cultures. Finally, the addition of slight positive charges did not induce cell toxicity.

Keywords:  3D cell cultures; dimyristoylaminopropylaminopropyl; encapsulation efficiency; gene delivery; loading content; microfluidic device; neutral viscous core liposomes; poly(ethylenimine); positively charged viscous core liposomes; siRNA

DOI:  https://doi.org/10.3390/pharmaceutics13040479
Pharmaceutics. 2021 Apr 03. pii: 490. [Epub ahead of print]13(4):

Lipid Nanoparticle-Mediated Lymphatic Delivery of Immunostimulatory Nucleic Acids.

Dongyoon Kim, Yina Wu, Gayong Shim, Yu-Kyoung Oh.

  Lymphatic delivery of a vaccine can be achieved using a dendritic cell (DC)-targeted delivery system that can cause DC to migrate to lymph nodes upon activation by an adjuvant. Here, we designed a mannose-modified cationic lipid nanoparticle (M-NP) to deliver the nucleic acid adjuvant, polyinosinic:polycytidylic acid (PIC). PIC-loaded M-NP (PIC/M-NP) showed stable lipoplexes regardless of the ligand ratio and negligible cytotoxicity in bone marrow-derived DC. DC uptake of PIC/M-NP was demonstrated, and an increased mannose ligand ratio improved DC uptake efficiency. PIC/M-NP significantly promoted the maturation of bone marrow-derived DC, and local injection of PIC/M-NP to mice facilitated lymphatic delivery and activation (upon NP uptake) of DC. Our results support the potential of PIC/M-NP in delivering a nucleic acid adjuvant for the vaccination of antigens.

Keywords:  dendritic cell-targeted delivery; lipid nanoparticle; lymphatic delivery; nucleic acid adjuvant

DOI:  https://doi.org/10.3390/pharmaceutics13040490
Int J Pharm. 2021 Apr 23. pii: S0378-5173(21)00446-4. [Epub ahead of print] 120641

Amino acids functionalized dendrimers with nucleus accumulation for efficient gene delivery.

Guoxin Tan, Jiayang Li, Dandan Liu, Hao Pan, Renfang Zhu, Yue Yang, Weisan Pan.

  Gene therapy is a promising approach to many diseases, however, the barriers in the gene delivery restrict its application. Therefore, in the present study, an efficient non-viral gene vector (PRHF/N/D) for overcoming the barriers in gene delivery was prepared. The synthesized PRHF integrated the advantages of PAMAM and amino acids, which could improve the cellular uptake, enhance the endosomal escape ability and minimize cytotoxicity. To further enhance nuclear entry of carrier, the nuclear localization signal (NLS) peptide was selected to add in the PRHF/D polyplexes. The PRHF/N/D polyplexes demonstrated good condensation capacity, wonderful pDNA protection and low toxicity. Moreover, the PRHF/N/D polyplexes showed the excellent transfection efficiency than P/D. PRHF/N/D further improve transfection capability than PRHF/D in the presence of NLS. After 4h of incubation, the mean fluorescence intensity of PRHF/N/D was also higher than the P/D and PRHF/D complexes. We then investigated the intracellular dissociation, the DNA is able to disassemble from PRHF/N/D gene carriers. Taken together, we exhibited that this PRHF/N/D polyplexes has the potential for use in the gene delivery.

Keywords:  dendrimer; endosomal escape; gene delivery; nucleus accumulation

DOI:  https://doi.org/10.1016/j.ijpharm.2021.120641