bims-novged Biomed News
on Non-viral vectors for gene delivery
Issue of 2022–12–04
eleven papers selected by
the Merkel lab, Ludwig-Maximilians University
  1. Synthesis and Application of Low Molecular Weight PEI-based Copolymers for siRNA Delivery with Smart Polymer Blends.
  2. Optimization of Lung Surfactant Coating of siRNA Polyplexes for Pulmonary Delivery.
  3. Single-component lipid nanoparticles for engineering SOCS1 gene-silenced dendritic cells to boost tumor immunotherapy.
  4. Hybrid core-shell particles for mRNA systemic delivery.
  5. Lipid nanoparticles for delivery of RNA therapeutics: Current status and the role of in vivo imaging.
  6. A Thermal and Enzymatic Dual-Stimuli Responsive DNA-Based Nanomachine for Controlled mRNA Delivery.
  7. Advances in Lipid-Based Co-delivery Systems for Cancer and Inflammatory Diseases.
  8. Species-agnostic polymeric formulations for inhalable messenger RNA delivery to the lung.
  9. Respiratory mucosal vaccination of peptide-poloxamine-DNA nanoparticles provides complete protection against lethal SARS-CoV-2 challenge.
  10. Multi-Layered PLGA-PEI Nanoparticles Functionalized with TKD Peptide for Targeted Delivery of Pep5 to Breast Tumor Cells and Spheroids.
  11. Current Research on Spray-Dried Chitosan Nanocomposite Microparticles for Pulmonary Drug Delivery.
  1. Macromol Biosci. 2022 Nov 29. e2200409
    Synthesis and Application of Low Molecular Weight PEI-based Copolymers for siRNA Delivery with Smart Polymer Blends.
    Yao Jin, Friederike Adams, Judith Möller, Lorenz Isert, Christoph M Zimmermann, David Keul, Olivia M Merkel.
      Polyethylenimine (PEI) is a commonly used cationic polymer for small-interfering RNA (siRNA) delivery due to its high transfection efficiency at low commercial cost. However, high molecular weight PEI is cytotoxic and thus, its practical application is limited. In this study, we investigated different formulations of low molecular weight PEI (LMW-PEI) based copolymers PEI-PCL (800 Da-40 kDa) and PEI-PCL-PEI (5 kDa-5 kDa-5 kDa) blended with or without PEG-PCL (5 kDa-4 kDa) to prepare nanoparticles via nanoprecipitation using a solvent displacement method with sizes around 100 nm. PEG-PCL can stabilize the nanoparticles, improve their biocompatibility, and extend their circulation time in vivo. The nanoparticles composed of PEI-PCL-PEI and PEG-PCL showed higher siRNA encapsulation efficiency than PEI-PCL/PEG-PCL based nanoparticles at low N/P ratios, higher cellular uptake, and a gene silencing efficiency of around 40% as a result of the higher molecular weight PEI blocks. These results suggested that the PEI-PCL-PEI/PEG-PCL nanoparticle system could be a promising vehicle for siRNA delivery at minimal synthetic effort. This article is protected by copyright. All rights reserved.
    Keywords:  PEG-PCL; low molecular weight PEI; nanoprecipitation; siRNA delivery
    DOI:  https://doi.org/10.1002/mabi.202200409
  2. Pharm Res. 2022 Nov 29.
    Optimization of Lung Surfactant Coating of siRNA Polyplexes for Pulmonary Delivery.
    Domizia Baldassi, Thi My Hanh Ngo, Olivia M Merkel.
       PURPOSE: The aim of this study was to understand how coating with a pulmonary surfactant, namely Alveofact, affects the physicochemical parameters as well as in vitro behavior of polyethylenimine (PEI) polyplexes for pulmonary siRNA delivery.
    METHODS: Alveofact-coated polyplexes were prepared at different Alveofact:PEI coating ratios and analyzed in terms of size, PDI and zeta potential as well as morphology by transmission electron microscopy. The biological behavior was evaluated in a lung epithelial cell line regarding cell viability, cellular uptake via flow cytometry and gene downregulation by qRT-PCR. Furthermore, a 3D ALI culture model was established to test the mucus diffusion and cellular uptake by confocal microscopy as well as gene silencing activity by qRT-PCR.
    RESULTS: After optimizing the coating process by testing different Alveofact:PEI coating ratios, a formulation with suitable parameters for lung delivery was obtained. In lung epithelial cells, Alveofact-coated polyplexes were well tolerated and internalized. Furthermore, the coating improved the siRNA-mediated gene silencing efficiency. Alveofact-coated polyplexes were then tested on a 3D air-liquid interface (ALI) culture model that, by expressing tight junctions and secreting mucus, resembles important traits of the lung epithelium. Here, we identified the optimal Alveofact:PEI coating ratio to achieve diffusion through the mucus layer while retaining gene silencing activity. Interestingly, the latter underlined the importance of establishing appropriate in vitro models to achieve more consistent results that better predict the in vivo activity.
    CONCLUSION: The addition of a coating with pulmonary surfactant to polymeric cationic polyplexes represents a valuable formulation strategy to improve local delivery of siRNA to the lungs.
    Keywords:  air-liquid interface; polyplexes; pulmonary delivery; pulmonary surfactant; siRNA delivery
    DOI:  https://doi.org/10.1007/s11095-022-03443-3
  3. Biomater Sci. 2022 Nov 28.
    Single-component lipid nanoparticles for engineering SOCS1 gene-silenced dendritic cells to boost tumor immunotherapy.
    Zexuan Yu, Mengtong Wu, Yingshuang Huang, Yishu Wang, Yijun Chen, Qiulin Long, Ziming Lin, Lingjing Xue, Caoyun Ju, Can Zhang.
      Dendritic cells (DCs) that can prime antitumor responses show great potential in tumor immunotherapy, whereas the unsatisfactory effect which can be ascribed in part to the high expression of inhibitory cytokines, such as the suppressor of cytokine signaling 1 (SOCS1), restricts their application. Thus, silencing these genes in DCs is essential for DC-based therapy. However, safe and effective delivery of siRNA to DCs still faces challenges. Herein, we designed single-component lipid nanoparticles comprising a solely cationic lipid (OA2) for introducing siRNA into mouse DCs in order to inhibit the immunosuppressive gene and boost the effector responses of DC-based therapy. Compared to other multi-component lipid nanoparticles, single-component lipid nanoparticles are theoretically easy-to-control and detective, which is beneficial for future translation. We showed that the application of OA2 lipid nanoparticles significantly downregulated the expression of SOCS1 in DCs over 50%, compared with the commercial lipofectine2000. Besides, the treatment of OA2 lipid nanoparticles had no influence on the antigen capture of DCs. Thus, we fabricated a SOCS1-downregulated DC vaccine pulsed with Ova antigen and demonstrated that the antigen presentation and pro-inflammatory factor secretion ability of DCs were improved due to the SOCS1 downregulation, leading to an ameliorated immunosuppressive tumor microenvironment and finally exhibiting potent tumor prevention and suppression in B16-Ova tumor-bearing mice. Single-component lipid nanoparticles, which provide an available vector platform for siRNA delivery to primary DCs, appear to be a potent tool to engineer DCs and in turn boost DC-based tumor immunotherapy.
    DOI:  https://doi.org/10.1039/d2bm01549h
  4. J Control Release. 2022 Nov 25. pii: S0168-3659(22)00787-8. [Epub ahead of print]
    Hybrid core-shell particles for mRNA systemic delivery.
    Valentina Andretto, Mathieu Repellin, Marine Pujol, Eyad Almouazen, Jacqueline Sidi-Boumedine, Thierry Granjon, Heyang Zhang, Katrien Remaut, Lars Petter Jordheim, Stéphanie Briançon, Isabel Sofia Keil, Fulvia Vascotto, Kerstin C Walzer, Ugur Sahin, Heinrich Haas, David Kryza, Giovanna Lollo.
      mRNA based infectious disease vaccines have opened the venue for development of novel nucleic acid-based therapeutics. For all mRNA therapeutics dedicated delivery systems are required, where different functionalities and targeting abilities need to be optimized for the respective applications. One option for advanced formulations with tailored properties are lipid-polymer hybrid nanoparticles with complex nanostructure, which allow to combine features of several already well described nucleic acid delivery systems. Here, we explored hyaluronic acid (HA) as coating of liposome-mRNA complexes (LRCs) to investigate effects of the coating on surface charge, physicochemical characteristics and biological activity. HA was electrostatically attached to positively charged complexes, forming hybrid LRCs (HLRCs). At different N/P ratios, physico-chemical characterization of the two sets of particles showed similarity in size (around 200 nm) and mRNA binding abilities, while the presence of the HA shell conferred a negative surface charge to otherwise positive complexes. High transfection efficiency of LRCs and HLRCs in vitro has been obtained in THP-1 and human monocytes derived from PBMC, an interesting target cell population for cancer and immune related pathologies. In mice, quantitative biodistribution of radiolabeled LRC and HLRC particles, coupled with bioluminescence studies to detect the protein translation sites, hinted towards both particles' accumulation in the hepatic reticuloendothelial system (RES). mRNA translated proteins though was found mainly in the spleen, a major source for immune cells, with preference for expression in macrophages. The results showed that surface modifications of liposome-mRNA complexes can be used to fine-tune nanoparticle physico-chemical characteristics. This provides a tool for assembly of stable and optimized nanoparticles, which are prerequisite for future therapeutic intervention using mRNA-based nanomedicines.
    Keywords:  Core-shell nanoparticles; Hyaluronic acid; Hybrid lipid-polymer nanoparticles; IVT mRNA; In vivo biodistribution; Lipoplexes
    DOI:  https://doi.org/10.1016/j.jconrel.2022.11.042
  5. Theranostics. 2022 ;12(17): 7509-7531
    Lipid nanoparticles for delivery of RNA therapeutics: Current status and the role of in vivo imaging.
    Han Na Jung, Seok-Yong Lee, Somin Lee, Hyewon Youn, Hyung-Jun Im.
      Lipid nanoparticles (LNPs) have been one of the most successful nano-delivery vehicles that enable efficient delivery of cytotoxic chemotherapy agents, antibiotics, and nucleic acid therapeutics. During the coronavirus disease (COVID-19) pandemic, LNP-based COVID-19 messenger RNA (mRNA) vaccines from Pfizer/BioNTech and Moderna have been successfully developed, resulting in global sales of $37 billion and $17.7 billion, respectively, in 2021. Based on this success, the development of multiple LNP-based RNA therapeutics is gaining momentum due to its potential in vaccines and therapeutics for various genetic diseases and cancers. Furthermore, imaging techniques can be utilized to evaluate the pharmacokinetics and pharmacodynamics (PK/PD) effects, which helps target discovery and accelerates the development of LNP-based mRNA therapies. A thorough introduction and explanation of the components of LNPs and its functions along with various production methods of formulating LNPs are provided in this review. Furthermore, recent advances in LNP-based RNA therapeutics in clinics and clinical trials are explored. Additionally, the evaluation of PK/PD of LNPs for RNA delivery and the current and potential roles in developing LNP-based mRNA pharmaceutics through imaging techniques will be discussed.
    Keywords:  Drug delivery system; In vivo imaging; Lipid nanoparticles; RNA therapeutics; Vaccines
    DOI:  https://doi.org/10.7150/thno.77259
  6. Adv Sci (Weinh). 2022 Dec 03. e2204905
    A Thermal and Enzymatic Dual-Stimuli Responsive DNA-Based Nanomachine for Controlled mRNA Delivery.
    Feng Li, Xiaolei Sun, Jing Yang, Jin Ren, Mengxue Huang, Shengqi Wang, Dayong Yang.
      The extreme instability of mRNA makes the practical application of mRNA-based vaccines heavily rely on efficient delivery system and cold chain transportation. Herein, a DNA-based nanomachine, which achieves programmed capture, long-term storage without cryopreservation, and efficient delivery of mRNA in cells, is developed. The polythymidine acid (Poly-T) functionalized poly(N-isopropylacrylamide) (DNA-PNIPAM) is synthesized and assembled as the central compartment of the nanomachine. The DNA-PNIPAM nano-assembly exhibits reversible thermal-responsive dynamic property: when lower than the low critical solution temperature (LCST, ≈32 °C) of PNIPAM, the DNA-PNIPAM transforms into extension state to expose the poly-T, facilitating the hybridization with polyadenylic acid (Poly-A) tail of mRNA; when higher than LCST, DNA-PNIPAM re-assembles and achieves an efficient encapsulation of mRNA. It is remarkable that the DNA-PNIPAM nano-assembly realizes long-term storage of mRNA (≈7 days) at 37 °C. Biodegradable 2-hydroxypropyltrimethyl ammonium chloride chitosan is assembled on the outside of DNA-PNIPAM to facilitate the endocytosis of mRNA, RNase-H mediating mRNA release occurs in cytoplasm, and efficient mRNA translation is achieved. This work provides a new disign principle of nanosystem for mRNA delivery.
    Keywords:  DNA nanotechnology; mRNA delivery; nanomedicine; self-assembly
    DOI:  https://doi.org/10.1002/advs.202204905
  7. Adv Healthc Mater. 2022 Dec 01. e2202400
    Advances in Lipid-Based Co-delivery Systems for Cancer and Inflammatory Diseases.
    Chinmay M Jogdeo, Sudipta Panja, Shrey Kanvinde, Ekta Kapoor, Kasturi Siddhanta, David Oupický.
      Combination therapy targeting multiple therapeutic targets is a favorable strategy to achieve better therapeutic outcomes in cancer and inflammatory diseases. Co-delivery is a subfield of drug delivery that aims to achieve combined delivery of diverse therapeutic cargoes within the same delivery system, thereby ensuring delivery to the same site and providing an opportunity to tailor the release kinetics as desired. Amongst the wide range of materials being investigated in the design of co-delivery systems, lipids have stood out on account of their low toxicity, biocompatibility, and ease of formulation scale-up. This review highlights the advances of the last decade in lipid-based co-delivery systems focusing on the co-delivery of drug-drug, drug-nucleic acid, nucleic acid-nucleic acid, and protein therapeutic based combinations for targeted therapy in cancer and inflammatory diseases. This article is protected by copyright. All rights reserved.
    Keywords:  Co-delivery; cancer; combination therapy; inflammatory bowel disease; lipid nanoparticles; psoriasis
    DOI:  https://doi.org/10.1002/adhm.202202400
  8. Nat Mater. 2022 Nov 28.
    Species-agnostic polymeric formulations for inhalable messenger RNA delivery to the lung.
    Laura Rotolo, Daryll Vanover, Nicholas C Bruno, Hannah E Peck, Chiara Zurla, Jackelyn Murray, Richard K Noel, Laura O'Farrell, Mariluz Araínga, Nichole Orr-Burks, Jae Yeon Joo, Lorena C S Chaves, Younghun Jung, Jared Beyersdorf, Sanjeev Gumber, Ricardo Guerrero-Ferreira, Santiago Cornejo, Merrilee Thoresen, Alicia K Olivier, Katie M Kuo, James C Gumbart, Amelia R Woolums, Francois Villinger, Eric R Lafontaine, Robert J Hogan, M G Finn, Philip J Santangelo.
      Messenger RNA has now been used to vaccinate millions of people. However, the diversity of pulmonary pathologies, including infections, genetic disorders, asthma and others, reveals the lung as an important organ to directly target for future RNA therapeutics and preventatives. Here we report the screening of 166 polymeric nanoparticle formulations for functional delivery to the lungs, obtained from a combinatorial synthesis approach combined with a low-dead-volume nose-only inhalation system for mice. We identify P76, a poly-β-amino-thio-ester polymer, that exhibits increased expression over formulations lacking the thiol component, delivery to different animal species with varying RNA cargos and low toxicity. P76 allows for dose sparing when delivering an mRNA-expressed Cas13a-mediated treatment in a SARS-CoV-2 challenge model, resulting in similar efficacy to a 20-fold higher dose of a neutralizing antibody. Overall, the combinatorial synthesis approach allowed for the discovery of promising polymeric formulations for future RNA pharmaceutical development for the lungs.
    DOI:  https://doi.org/10.1038/s41563-022-01404-0
  9. Biomaterials. 2022 Nov 18. pii: S0142-9612(22)00547-6. [Epub ahead of print]292 121907
    Respiratory mucosal vaccination of peptide-poloxamine-DNA nanoparticles provides complete protection against lethal SARS-CoV-2 challenge.
    Si Sun, Entao Li, Gan Zhao, Jie Tang, Qianfei Zuo, Larry Cai, Chuanfei Xu, Cheng Sui, Yangxue Ou, Chang Liu, Haibo Li, Yuan Ding, Chao Li, Dongshui Lu, Weijun Zhang, Ping Luo, Ping Cheng, Yuwei Gao, Changchun Tu, Bruno Pitard, Joseph Rosenecker, Bin Wang, Yan Liu, Quanming Zou, Shan Guan.
      The ongoing SARS-CoV-2 pandemic represents a brutal reminder of the continual threat of mucosal infectious diseases. Mucosal immunity may provide robust protection at the predominant sites of SARS-CoV-2 infection. However, it remains unclear whether respiratory mucosal administration of DNA vaccines could confer protective immune responses against SARS-CoV-2 challenge due to insurmountable barriers posed by the airway. Here, we applied self-assembled peptide-poloxamine nanoparticles with mucus-penetrating properties for pulmonary inoculation of a COVID-19 DNA vaccine (pSpike/PP-sNp). The pSpike/PP-sNp not only displays superior gene transfection and favorable biocompatibility in the mouse airway, but also promotes a tripartite immunity consisting of systemic, cellular, and mucosal immune responses that are characterized by mucosal IgA secretion, high levels of neutralizing antibodies, and resident memory phenotype T-cell responses in the lungs of mice. Most importantly, immunization with pSpike/PP-sNp completely eliminates SARS-CoV-2 infection in both upper and lower respiratory tracts and enables 100% survival rate of mice following lethal SARS-CoV-2 challenge. Our findings indicate PP-sNp is a promising platform in mediating DNA vaccines to elicit all-around mucosal immunity against SARS-CoV-2.
    Keywords:  DNA vaccine; Mucosal vaccine; Nonviral delivery system; Pulmonary delivery; SARS-CoV-2
    DOI:  https://doi.org/10.1016/j.biomaterials.2022.121907
  10. Int J Nanomedicine. 2022 ;17 5581-5600
    Multi-Layered PLGA-PEI Nanoparticles Functionalized with TKD Peptide for Targeted Delivery of Pep5 to Breast Tumor Cells and Spheroids.
    Akhil K Mohan, Minsa M, T R Santhosh Kumar, G S Vinod Kumar.
       Purpose: Peptide-based therapy is a promising strategy for cancer treatment because of its low drug resistance. However, the major challenge is their inability to target cancer cells specifically. So, a targeted nano-delivery system that could deliver therapeutic peptides selectively to cancer cells to stimulate their action is highly desirable. This study aims to deliver the antitumor peptide, Pep5, to breast tumor cells selectively using a targeting peptide functionalised multi-layered PLGA-PEI nanoparticles.
    Methods: In this study, Pep5 entrapped PLGA-PEI (Pep5-PPN) dual layered nanoparticles were developed. These nanoparticles were decorated with TKD (Pep5-TPPN) on their surface for site-specific delivery of Pep5 to breast tumor cells. The particles were then characterized using various instrumental analyses. In vitro cytotoxicity of the particles was evaluated in estrogen receptor positive (ER+ve) and triple negative breast cancer (TNBC) cells. An ex vivo tumor spheroid model was used to analyze the antitumor activity of the particles.
    Results: Uniformly round Pep5-TPPN particles were synthesized with an average diameter of 420.8 ± 14.72 nm. The conjugation of PEI over Pep5-PLGA nanoparticles shifted the zeta potential from -11.6 ± 2.16 mV to +20.01 ± 2.97 mV. In vitro cytotoxicity analysis proved that TKD conjugation to nanoparticles enhanced the antitumor activity of Pep5 in tested breast cancer cells. Pep5-TPPN induced cytoskeletal damage and apoptosis in the tested cells, which showed that the mechanism of action of Pep5 is conserved but potentiated. Active targeting of Pep5 suppressed the tumor growth in ex vivo spheroid models.
    Conclusion: A multi-layered nanoparticle functionalized with dual peptide was fabricated for active tumor targeting, which stimulated Pep5 activity to reduce the tumor growth in vitro and ex vivo.
    Keywords:  Pep 5; breast; cancer; peptide; tumor
    DOI:  https://doi.org/10.2147/IJN.S376358
  11. Pharm Nanotechnol. 2022 Nov 28.
    Current Research on Spray-Dried Chitosan Nanocomposite Microparticles for Pulmonary Drug Delivery.
    Saba Albetawi.
      Using the pulmonary route for systemic and local drug delivery is an attractive method of drug administration because it has a high alveolar surface area, abundant blood flow, a thin air-blood barrier, and low metabolic activity. In recent years, the evolution of inhalable chitosan nanocomposite microparticles formulations enabled researchers to develop new pulmonary drug delivery platforms that combine the advantages of microparticles and nanoparticles using a biocompatible, biodegradable polymer with polycationic nature and inherent immunogenicity that enhances cell targeting. Therefore, this review aims to offer an overview of the recent advances in inhalable chitosan nanocomposites microparticles formulated in the previous five years in terms of primary nanoparticles manufacturing methods; namely, ionic crosslinking of chitosan using tripolyphosphate, electrospinning/electrospraying, layer-by-layer deposition, and nanospray drying; final microparticles manufacturing techniques using spray drying, nano spray drying, and supercritical assisted spray drying; in addition to the process optimization of the previously mentioned manufacturing methods. Furthermore, this review highlights using chitosan and its derivatives in primary nanoparticles preparation and as a polysaccharide to distribute the prepared nanoparticles in microparticles. Finally, this review discusses the factors affecting yield, encapsulation efficiency, in vitro aerosolization properties, size, morphological characters, in vitro release, and in vivo evaluation of inhalable chitosan nanocomposite microparticles.
    Keywords:  Chitosan; Lung Deposition; Nanocomposite; Nanocomposite Microparticles; Pulmonary; Spray Drying
    DOI:  https://doi.org/10.2174/2211738511666221128093822
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