bims-novged 2023-07-16 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2023–07–16
nine papers selected by
the Merkel lab, Ludwig-Maximilians University

One-pot synthesis of dynamically cross-linked polymers for serum-resistant nucleic acid delivery.
Enhanced immunogenicity induced by mRNA vaccines with various lipid nanoparticles as carriers for SARS-CoV-2 infection.
Lipid nanoparticles (LNPs) for in vivo RNA delivery and their breakthrough technology for future applications.
Harnessing the potential of lipid nanoparticles for the delivery of chemically modified siRNA to combat hepatic adenovirus infection.
Overcoming the blood-brain barrier? - prediction of blood-brain permeability of hydrophobically modified polyethylenimine polyplexes for siRNA delivery into the brain with in vitro and in vivo models.
Genome editing mRNA nanotherapies inhibit cervical cancer progression and regulate the immunosuppressive microenvironment for adoptive T-cell therapy.
Non-equilibrium transport of nanoparticles across the lipid membrane.
Automated Parameterization of Coarse-Grained Polyethylenimine under a Martini Framework.
Rational nanoparticle design: Optimization using insights from experiments and mathematical models.

Biomater Sci. 2023 Jul 11.

One-pot synthesis of dynamically cross-linked polymers for serum-resistant nucleic acid delivery.

Zhisong Sun, Mengyao Ren, Bingchen Shan, Qiang Yang, Ziyin Zhao, Xun Liu, Lichen Yin.

Cationic polymers used for nucleic acid delivery often suffer from complicated syntheses, undesired intracellular cargo release and low serum stability. Herein, a series of ternary polymers were synthesized via facile green chemistry to achieve efficient plasmid DNA and mRNA delivery in serum. During the one-pot synthesis of the ternary polymer, acetylphenylboric acid (APBA), polyphenol and low-molecular weight polyethyleneimine (PEI 1.8k) were dynamically cross-linked with each other due to formation of an imine between PEI 1.8k and APBA and formation of a boronate ester between APBA and polyphenol. Series of polyphenols, including ellagic acid (EA), epigallocatechin gallate (EGCG), nordihydroguaiaretic acid (NDGA), rutin (RT) and rosmarinic acid (RA), and APBA molecules, including 2-acetylphenylboric acid (2-APBA), 3-acetylphenylboric acid (3-APBA) and 4-acetylphenylboric acid (4-APBA), were screened and the best-performing ternary polymer, 2-PEI-RT, constructed from RT and 2-APBA, was identified. The ternary polymer featured efficient DNA condensation to favor cellular internalization, and the acidic environment in endolysosomes triggered effective degradation of the polymer to promote cargo release. Thus, 2-PEI-RT showed robust plasmid DNA transfection efficiencies in various tumor cells in serum, outperforming the commercial reagent PEI 25k by 1-3 orders of magnitude. Moreover, 2-PEI-RT mediated efficient cytosolic delivery of Cas9-mRNA/sgRNA to enable pronounced CRISPR-Cas9 genome editing in vitro. Such a facile and robust platform holds great potential for non-viral nucleic acid delivery and gene therapy.

DOI: https://doi.org/10.1039/d3bm00685a
J Mater Chem B. 2023 Jul 14.

Enhanced immunogenicity induced by mRNA vaccines with various lipid nanoparticles as carriers for SARS-CoV-2 infection.

Yanhao Zhang, Ji Wang, Hanlei Xing, Chao Liu, Wenhui Zha, Shuo Dong, Yuhao Jiang, Xinsong Li.

mRNA vaccines have emerged as a highly promising approach for preventing cancer and infectious diseases, attributed to their superior immunogenicity, rapid development speed, and quality-controlled scale production. While homologous mRNA vaccine administration is currently the most prevalent method employed in clinical settings, heterologous administration is a promising avenue worth exploring. In this report, two types of mRNA vaccine formulations for SARS-CoV-2 infection were developed based on different lipid nanoparticle (LNP) delivery systems, and heterologous and homologous mRNA vaccinations were administered to explore the levels of immune responses comparatively. First, five novel H-series ionizable lipids were synthesized and confirmed by NMR and MS. Subsequently, six SARS-CoV-2 receptor-binding domain (RBD) mRNA-encapsulated LNP formulations were prepared using a microfluidic mixer based on H-series and MC3 lipids. These formulations exhibited spherical structures with an average diameter ranging from 90-140 nm, as characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM). The safety of these formulations was confirmed in vitro by the cytotoxicity assay. Moreover, transfection assay, lysosomal escape test, and western blot, and in vivo biodistribution analyses collectively demonstrated that lipids H03 and MC3 exhibited superior in vitro and in vivo delivery efficacy in comparison to other H-series lipids. Notably, H03-Fluc mRNA exhibited an approximately 2.2-fold higher in vivo bioluminescence signal intensity than MC3-Fluc mRNA. Additionally, evaluation of humoral immunity demonstrated that homologous H03-mRNA vaccination elicited an immune response that was approximately 3-fold higher than that of homologous MC3-mRNA vaccination. More significantly, the heterologous H03-mRNA/MC3-mRNA vaccination elicited an immune response that was approximately 2-3-fold higher than that of homologous H03-mRNA vaccination and 6-9-fold higher than that of homologous MC3-mRNA vaccination, without any observable adverse effects. These results suggest that heterologous mRNA vaccination is superior to homologous mRNA vaccination and may be attributed to differences in LNP carriers. Therefore, our research may inspire further exploration of different delivery systems to enhance mRNA-based therapeutics.

DOI: https://doi.org/10.1039/d3tb00303e
Adv Drug Deliv Rev. 2023 Jul 07. pii: S0169-409X(23)00305-8. [Epub ahead of print] 114990

Lipid nanoparticles (LNPs) for in vivo RNA delivery and their breakthrough technology for future applications.

Michaela Jeong, Yeji Lee, Jeongeun Park, Hyein Jung, Hyukjin Lee.

  RNA therapeutics show a significant breakthrough for the treatment of otherwise incurable diseases and genetic disorders by regulating disease-related gene expression. The successful development of COVID-19 mRNA vaccines further emphasizes the potential of RNA therapeutics in the prevention of infectious diseases as well as in the treatment of chronic diseases. However, the efficient delivery of RNA into cells remains a challenge, and nanoparticle delivery systems such as lipid nanoparticles (LNPs) are necessary to fully realize the potential of RNA therapeutics. While LNPs provide a highly efficient platform for the in vivo delivery of RNA by overcoming various biological barriers, several challenges remain to be resolved for further development and regulatory approval. These include a lack of targeted delivery to extrahepatic organs and a gradual loss of therapeutic potency with repeated doses. In this review, we highlight the fundamental aspects of LNPs and their uses in the development of novel RNA therapeutics. Recent advances in LNP-based therapeutics and preclinical/clinical studies are overviewed. Lastly, we discuss the current limitations of LNPs and introduce breakthrough technologies that might overcome these challenges in future applications.

Keywords:  RNA therapeutics; extrahepatic delivery; lipid nanoparticles (LNPs); repeated dose

DOI:  https://doi.org/10.1016/j.addr.2023.114990
Mol Ther Nucleic Acids. 2023 Sep 12. 33 44-46

Harnessing the potential of lipid nanoparticles for the delivery of chemically modified siRNA to combat hepatic adenovirus infection.

Quazi T H Shubhra, A K M Moshiul Alam.



Keywords:  drug delivery system; hepatic infection; human adenovirus; lipid nanoparticles; siRNA

DOI:  https://doi.org/10.1016/j.omtn.2023.06.005
J Control Release. 2023 Jul 10. pii: S0168-3659(23)00443-1. [Epub ahead of print]

Overcoming the blood-brain barrier? - prediction of blood-brain permeability of hydrophobically modified polyethylenimine polyplexes for siRNA delivery into the brain with in vitro and in vivo models.

Natascha Hartl, Bettina Gabold, Friederike Adams, Philipp Uhl, Sabrina Oerter, Sabine Gätzner, Marco Metzger, Ann-Christine König, Stefanie M Hauck, Antje Appelt-Menzel, Walter Mier, Gert Fricker, Olivia M Merkel.

  The blood-brain barrier (BBB) is a highly selective biological barrier that represents a major bottleneck in the treatment of all types of central nervous system (CNS) disorders. Small interfering RNA (siRNA) offers in principle a promising therapeutic approach, e.g., for brain tumors, by downregulating brain tumor-related genes and inhibiting tumor growth via RNA interference. In an effort to develop efficient siRNA nanocarriers for crossing the BBB, we utilized polyethyleneimine (PEI) polymers hydrophobically modified with either stearic-acid (SA) or dodecylacrylamide (DAA) subunits and evaluated their suitability for delivering siRNA across the BBB in in vitro and in vivo BBB models depending on their structure. Physicochemical characteristics of siRNA-polymer complexes (polyplexes (PXs)), e.g., particle size and surface charge, were measured by dynamic light scattering and laser Doppler anemometry, whereas siRNA condensation ability of polymers and polyplex stability was evaluated by spectrophotometric methods. The composition of the biomolecule corona that absorbs on polyplexes upon encountering physiological fluids was investigated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and by a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method. Cellular internalization abilities of PXs into brain endothelial cells (hCMEC/D3) was confirmed, and a BBB permeation assay using a human induced pluripotent stem cell (hiPSC)-derived BBB model revealed similar abilities to cross the BBB for all formulations under physiological conditions. However, biodistribution studies of radiolabeled PXs in mice were inconsistent with in vitro results as the detected amount of radiolabeled siRNA in the brain delivered with PEI PXs was higher compared to PEI-SA PXs. Taken together, PEI PXs were shown to be a suitable nanocarrier to deliver small amounts of siRNA across the BBB into the brain but more sophisticated human BBB models that better represent physiological conditions and biodistribution are required to provide highly predictive in vitro data for human CNS drug development in the future.

Keywords:  Blood-brain barrier; Hydrophobically modified cationic polymers; Polyplexes; Protein corona; hiPSC-derived BBB model; siRNA delivery

DOI:  https://doi.org/10.1016/j.jconrel.2023.07.019
J Control Release. 2023 Jul 12. pii: S0168-3659(23)00431-5. [Epub ahead of print]360 496-513

Genome editing mRNA nanotherapies inhibit cervical cancer progression and regulate the immunosuppressive microenvironment for adoptive T-cell therapy.

Kaijian Ling, Yin Dou, Neng Yang, Li Deng, Yanzhou Wang, Yudi Li, Leiyan Yang, Cheng Chen, Lupin Jiang, Qingchun Deng, Chenwen Li, Zhiqing Liang, Jianxiang Zhang.

  CRISPR/Cas9-based genome editing is promising for therapy of cervical cancer by precisely targeting human papillomavirus (HPV). To develop CRISPR/Cas9-based genome editing nanotherapies, a pH-responsive hybrid nonviral nanovector was constructed for co-delivering Cas9 mRNA and guide RNAs (gRNAs) targeting E6 or E7 oncogenes. The pH-responsive nanovector was fabricated using an acetalated cyclic oligosaccharide (ACD), in combination with low molecular weight polyethyleneimine. Thus obtained hybrid ACD nanoparticles (defined as ACD NP) showed efficient loading for both Cas9 mRNA and E6 or E7 gRNA, giving rise to two pH-responsive genome editing nanotherapies E6/ACD NP and E7/ACD NP, respectively. Cellularly, ACD NP exhibited high transfection but low cytotoxicity in HeLa cervical carcinoma cells. Also, efficient genome editing of target genes was achieved in HeLa cells, with minimal off-target effects. In mice bearing HeLa xenografts, treatment with E6/ACD NP or E7/ACD NP afforded effective editing of target oncogenes and considerable antitumor activities. More importantly, treatment with E6/ACD NP or E7/ACD NP notably promoted CD8+ T cell survival by reversing the immunosuppressive microenvironment, thereby leading to synergistic antitumor effects by combination therapy using the gene editing nanotherapies and adoptive T-cell transfer. Consequently, our pH-responsive genome editing nanotherapies deserve further development for the treatment of HPV-associated cervical cancer, and they can also serve as promising nanotherapies to improve efficacies of other immune therapies against different advanced cancers by regulating the immunosuppressive tumor microenvironment.

Keywords:  CRISPR/Cas9; Cervical cancer; Genome editing; T-cell transfer therapy; mRNA nanotherapy; pH-responsive nanoparticles

DOI:  https://doi.org/10.1016/j.jconrel.2023.07.007
Nanoscale. 2023 Jul 11.

Non-equilibrium transport of nanoparticles across the lipid membrane.

Younghoon Oh, Qiang Cui.

Development of effective strategies for the internalization of nanoparticles is essential in many applications, such as drug delivery. Most, if not all, previous studies are based on equilibrium considerations. In this work, inspired by the recent development of a pro-drug delivery strategy based on reversible esterification, we consider a non-equilibrium transport mechanism for nanoparticles of a 6 nm diameter across the lipid membrane. We divide the transport process into insertion and ejection steps, which are studied with coarse-grained models using free energy and reactive Monte Carlo simulations, respectively. The simulations show that the non-equilibrium transport efficiency is relatively insensitive to the fraction of reactive surface ligands once a modest threshold is surpassed, while the distribution pattern of different (hydrophilic, reactive and permanent hydrophobic) ligands on the nanoparticle surface has a notable impact on both the insertion and ejection steps. Our study thus supports a novel avenue for designing nanoparticles that are able to be efficiently internalized and provides a set of relevant guidelines for surface functionalization.

DOI: https://doi.org/10.1039/d3nr00930k
J Chem Inf Model. 2023 Jul 09.

Automated Parameterization of Coarse-Grained Polyethylenimine under a Martini Framework.

Subhamoy Mahajan, Tian Tang.

As a versatile polymer in many applications, synthesized polyethylenimine (PEI) is polydisperse with diverse branched structures that attain pH-dependent protonation states. Understanding the structure-function relationship of PEI is necessary for enhancing its efficacy in various applications. Coarse-grained (CG) simulations can be performed at length and time scales directly comparable with experimental data while maintaining the molecular perspective. However, manually developing CG forcefields for complex PEI structures is time-consuming and prone to human errors. This article presents a fully automated algorithm that can coarse-grain any branched architecture of PEI from its all-atom (AA) simulation trajectories and topology. The algorithm is demonstrated by coarse-graining a branched 2 kDa PEI, which can replicate the AA diffusion coefficient, radius of gyration, and end-to-end distance of the longest linear chain. Commercially available 25 and 2 kDa Millipore-Sigma PEIs are used for experimental validation. Specifically, branched PEI architectures are proposed, coarse-grained using the automated algorithm, and then simulated at different mass concentrations. The CG PEIs can reproduce existing experimental data on PEI's diffusion coefficient and Stokes-Einstein radius at infinite dilution as well as its intrinsic viscosity. This suggests a strategy where probable chemical structures of synthetic PEIs can be inferred computationally using the developed algorithm. The coarse-graining methodology presented here can also be extended to other polymers.

DOI: https://doi.org/10.1021/acs.jcim.3c00103
J Control Release. 2023 Jul 11. pii: S0168-3659(23)00441-8. [Epub ahead of print]

Rational nanoparticle design: Optimization using insights from experiments and mathematical models.

Owen Richfield, Alexandra S Piotrowski-Daspit, Kwangsoo Shin, W Mark Saltzman.

  Polymeric nanoparticles are highly tunable drug delivery systems that show promise in targeting therapeutics to specific sites within the body. Rational nanoparticle design can make use of mathematical models to organize and extend experimental data, allowing for optimization of nanoparticles for particular drug delivery applications. While rational nanoparticle design is attractive from the standpoint of improving therapy and reducing unnecessary experiments, it has yet to be fully realized. The difficulty lies in the complexity of nanoparticle structure and behavior, which is added to the complexity of the physiological mechanisms involved in nanoparticle movement throughout the body. In this review, we discuss the most important aspects of rational design of polymeric nanoparticles. Ultimately, we conclude that many experimental datasets are required to fully model polymeric nanoparticle behavior at multiple scales. Further, we suggest ways to consider the limitations and uncertainty of experimental data in creating nanoparticle design optimization schema, which we call quantitative nanoparticle design frameworks.

Keywords:  Multiscale mathematical modeling; Nanoparticle pharmacokinetics; Physiologically based pharmacokinetics; Polymeric nanoparticles; Rational nanoparticle design

DOI:  https://doi.org/10.1016/j.jconrel.2023.07.018