A Single Methylene Group in Oligoalkylamine‐Based Cationic Polymers and Lipids Promotes Enhanced mRNA Delivery

Jarzębińska, Anita; Pasewald, Tamara; Lambrecht, Jana; Mykhaylyk, Olga; Kümmerling, Linda; Beck, Philipp; Hasenpusch, Günther; Rudolph, Carsten; Plank, Christian; Dohmen, Christian

doi:10.1002/anie.201603648

Cited by 88 publications

(69 citation statements)

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“…This stands in stark contrast to previous studies on intravenous delivery of PBAE‐DNA complexes, in which jetPEI outperformed PBAEs by two orders of magnitude in the lungs . A recent study demonstrated in vivo mRNA expression in the lungs after pulmonary administration of cationic polymeric nanoparticles; however, to our knowledge, ours is the first example of biodegradable polymeric nanoparticles successfully delivering mRNA to the lungs after intravenous injection in mice.…”

Section: Figurecontrasting

confidence: 95%

“…The function of PEGylated DD90‐C12‐122 and C32‐C12‐118 nanoparticles demonstrated here warrants continued development and optimization to further improve potency. One approach to potentially access even more potent formulations would utilize design‐of‐experiments methodology to vary the formulation components, which has been shown to improve mRNA delivery efficacy in a separate liver‐targeting system . Importantly, these data demonstrate a correlation between serum stability and in vivo efficacy; the terpolymers that form serum‐stable nanoparticles when formulated with a low amount (7 mol %) of PEG‐lipid also led to the most potent in vivo mRNA delivery.…”

Section: Figurementioning

confidence: 99%

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Polymer–Lipid Nanoparticles for Systemic Delivery of mRNA to the Lungs

et al. 2016

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Therapeutic nucleic acids hold great promise for the treatment of disease but require vectors for safe and effective delivery. Synthetic nanoparticle vectors composed of poly(β-amino esters) (PBAEs) and nucleic acids have previously demonstrated potential utility for local delivery applications. To expand potential utility to include systemic delivery of mRNA, here we develop hybrid polymer-lipid nanoformulations for systemic delivery to the lung. Through co-formulation of PBAEs with lipid-polyethylene glycol (PEG), mRNA formulations were developed with increased serum stability and increased in vitro potency. The formulations were capable of functional delivery of mRNA to the lungs following intravenous administration in mice. To our knowledge, this is the first description of systemic administration of mRNA for delivery to the lungs using a degradable polymer-lipid nanoparticle. KeywordsNanoparticles; mRNA; Drug delivery; PBAE; Stability dgander@mit.edu. Supporting information for this article is given via a link at the end of the document.More detailed experimental procedures can be found in the supplementary information provided. Previous studies have suggested that serum stability may limit the efficacy of PBAEs for systemic administration. HHS Public Access[15] To this end, PBAE terpolymer copolymers containing internal alkyl tails were developed.[16] The alkyl amine was incorporated to enable formulation of terpolymers with other hydrophobic components in order to increase particle stability in physiological conditions. Here, we demonstrate that PBAE terpolymer nanoparticles can be formulated with a polyethylene glycol-modified lipid (PEG-lipid) and result in serum stabilized nanoparticles and improved efficacy. PBAEs that were found to be serum stabilized in vitro upon incorporation of PEG-lipid at 7 mol% were evaluated in vivo and shown to be capable of mRNA delivery to the lungs in mice following intravenous administration.The PBAEs selected for synthesis in this study were based on analysis of literature results from this class of materials for DNA delivery.[16] We synthesized 7 PBAEs that included 6 lead terpolymers and 1 lead non-terpolymer (i.e. not including the alkyl amine) from the monomers shown in Figure 1a. Synthetic methodology was adapted for synthesis in dimethylformamide (DMF) and purification in diethylether in order to reduce the possibility of toxicity arising from excess monomer (Scheme S1).[16] Polymer characterization was performed via GPC, 1H-NMR, and FTIR (Figures S1-3). Nanoparticles ( Figure 1b) were prepared by mixing each synthesized polymer with mRNA coding for firefly luciferase diluted in acidic (pH 5.2) buffer. Nanoparticles were formulated such that the ratio of amine groups in the polymer to phosphate groups in the nucleic acid (N/P) was 57; this is consistent with a weight ratio of approximately 50:1, similar to what has been used for successful PBAE transfections previously.[17]Stable nanoparticles were formed both with and without addition of 7 mol% PEG-lipid ...

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Section: Figurecontrasting

confidence: 95%

Section: Figurementioning

confidence: 99%

Polymer–Lipid Nanoparticles for Systemic Delivery of mRNA to the Lungs

et al. 2016

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“…For in vivo purposes, cmRNAs (AA and AAstop) were formulated using Ethris’ (Planegg, Germany) proprietary cationic lipid formulation LF132, which was based on Jarzebińska et al. 57 In order to examine their effectiveness in vitro , cells were transfected with 10, 50, and 100 ng/100 μL of AA-LF132 or AAstop-LF132. As vehicle control, 2% sucrose (diluted accordingly) was applied.…”

Section: Methodsmentioning

confidence: 99%

Exploring Cytotoxic mRNAs as a Novel Class of Anti-cancer Biotherapeutics

Hirschberger

Jarzębińska

Kessel

et al. 2018

Molecular Therapy - Methods & Clinical Development

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New treatments to overcome the obstacles of conventional anti-cancer therapy are a permanent subject of investigation. One promising approach is the application of toxins linked to cell-specific ligands, so-called immunotoxins. Another attractive option is the employment of toxin-encoding plasmids. However, immunotoxins cause hepatoxicity, and DNA therapeutics, among other disadvantages, bear the risk of insertional mutagenesis. As an alternative, this study examined chemically modified mRNAs coding for diphtheria toxin, subtilase cytotoxin, and abrin-a for their ability to reduce cancer cell growth both in vitro and in vivo. The plant toxin abrin-a was the most promising candidate among the three tested toxins and was further investigated. Its expression was demonstrated by western blot. Experiments with firefly luciferase in reticulocyte lysates and co-transfection experiments with EGFP demonstrated the capability of abrin-a to inhibit protein synthesis. Its cytotoxic effect was quantified employing viability assays and propidium iodide staining. By studying caspase-3/7 activation, Annexin V-binding, and chromatin condensation with Hoechst33258 staining, apoptotic cell death could be confirmed. In mice, repeated intratumoral injections of complexed abrin-a mRNA resulted in a significant reduction (89%) of KB tumor size compared to a non-translatable control mRNA.

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“…Oligoalkylamines grafted to an 8000 Da poly(acrylic acid) (PAA8k) scaffold complexed with chemically modified mRNA resulted in another kind of polyplexes with transfection capacity. Intravenous administration of PAA8k-luciferase mRNA systems in mice showed different luminescence signal in liver depending on their structure [125].…”

Section: Polymeric Systemsmentioning

confidence: 99%

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

et al. 2020

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The use of messenger RNA (mRNA) in gene therapy is increasing in recent years, due to its unique features compared to plasmid DNA: Transient expression, no need to enter into the nucleus and no risk of insertional mutagenesis. Nevertheless, the clinical application of mRNA as a therapeutic tool is limited by its instability and ability to activate immune responses; hence, mRNA chemical modifications together with the design of suitable vehicles result essential. This manuscript includes a revision of the strategies employed to enhance in vitro transcribed (IVT) mRNA functionality and efficacy, including the optimization of its stability and translational efficiency, as well as the regulation of its immunostimulatory properties. An overview of the nanosystems designed to protect the mRNA and to overcome the intra and extracellular barriers for successful delivery is also included. Finally, the present and future applications of mRNA nanomedicines for immunization against infectious diseases and cancer, protein replacement, gene editing, and regenerative medicine are highlighted.

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A Single Methylene Group in Oligoalkylamine‐Based Cationic Polymers and Lipids Promotes Enhanced mRNA Delivery

Cited by 88 publications

References 17 publications

Polymer–Lipid Nanoparticles for Systemic Delivery of mRNA to the Lungs

Polymer–Lipid Nanoparticles for Systemic Delivery of mRNA to the Lungs

Exploring Cytotoxic mRNAs as a Novel Class of Anti-cancer Biotherapeutics

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

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