Superparamagnetic nanoparticle delivery of DNA and RNA-based vaccines

Superparamagnetic nanoparticle delivery of DNA and RNA-based vaccines

Dr. Robert Gorter:

This overview about the clinical application of super magnetic iron oxide nanoparticles (SPIONs) shows that these nanoparticles can be used successfully in DNA- and RNA- delivery to build vaccines by using SPIONs as vectors to deliver genes or DNA molecules into the host cell DNA (magnetoreception). Till today, as far as we know, mostly (retro-)viral vectors (like HIV and SIV and HTlV-1 and HTLV-2 have been used to deliver the (synthetic) RNA or DNA.

Also, we tried to show that superparamagnetic nanoparticles have been used and developed in Gain-of Function research.

This overview wants to present literature on Supermagnetic Iron Oxide Nanoparticles (SPIONs) to try and replace viral vectors, as all scientists know that using (retro-)viral vectors is a Box of Pandora.

How can I understand what a viral vector is? A viral vector is like a wheelbarrow that carries into the cell and into its genes (usually) synthetically made parts of a virus or parasite (like malaria) to force the cell to create and/or duplicate what has been inserted into the genes to force the immune system to create its own mRNA to induce an immune response (usually antibodies).

The major concern is that viruses from the animal world that are not pathogenic for humans (yet) are being used as vectors but there are multiple examples where nonpathogenic viruses in animals but not in humans became pathogenic for humans too by mutations. Thus; there is a sincere risk that these vectors become pathogenic once they enter the body.


Superparamagnetic nanoparticle delivery of DNA vaccines

Commentaries by Robert Gorter, MD, PhD.


Al-Deen FN, Selomulya C, Ma C, Coppel RL. Superparamagnetic nanoparticle delivery of DNA vaccine. Methods Mol Biol. 2014;1143:181-94. DOI: 10.1007/978-1-4939-0410-5_12. PMID: 24715289.


The efficiency of the delivery of DNA vaccines is often relatively low compared to protein vaccines. The use of superparamagnetic iron oxide nanoparticles (SPIONs) to deliver genes via magnetoreception shows promise in improving the efficiency of gene delivery both in vitro and in vivo. In particular, the duration for gene transfection especially for in vitro application can be significantly reduced by magnetoreception compared to the time required to achieve high gene transfection with standard protocols. SPIONs that have been rendered stable in physiological conditions can be used as both therapeutic and diagnostic agents due to their unique magnetic characteristics. Valuable features of iron oxide nanoparticles in bio-applications include a tight control over their size distribution, magnetic properties of these particles, and the ability to carry particular biomolecules to specific targets. The internalization and half-life of the particles within the body depend upon the method of synthesis. Numerous synthesis methods have been used to produce magnetic nanoparticles for bio-applications with different sizes and surface charges. The most common method for synthesizing nanometer-sized magnetite Fe3O4 particles in solution is by chemical co-precipitation of iron salts. The co-precipitation method is an effective technique for preparing stable aqueous dispersions of iron oxide nanoparticles. We describe the production of Fe3O4-based SPIONs with high magnetization values (70 emu/g) under 15 kOe of the applied magnetic field at room temperature, with 0.01 emu/g remanence via a co-precipitation method in the presence of trisodium citrate as a stabilizer. Naked SPIONs often lack sufficient stability, hydrophilicity, and the capacity to be functionalized. In order to overcome these limitations, the polycationic polymer was anchored on the surface of freshly prepared SPIONs by a direct electrostatic attraction between the negatively charged SPIONs (due to the presence of carboxylic groups) and the positively charged polymer. Polyethylenimine was chosen to modify the surface of SPIONs to assist the delivery of plasmid DNA into mammalian cells due to the polymer’s extensive buffering capacity through the “proton sponge” effect.

Superparamagnetic nanoparticles for effective delivery of malaria DNA vaccines

Al-Deen FN, Ho J, Selomulya C, Ma C, Coppel R. Superparamagnetic nanoparticles for effective delivery of malaria DNA vaccine. Langmuir. 2011 Apr 5;27(7):3703-12. DOI: 10.1021/la104479c. Epub 2011 Mar 1. PMID: 21361304.


Low efficiency is often observed in the delivery of DNA vaccines. The use of superparamagnetic nanoparticles (SPIONs) to deliver genes via magnetoreception could improve transfection efficiency and target the vector to its desired locality. Here, magnetoreception was used to enhance the delivery of a malaria DNA vaccine encoding Plasmodium yoelii merozoite surface protein MSP1(19) (VR1020-PyMSP1(19)) that plays a critical role in Plasmodium immunity. The plasmid DNA (pDNA) containing membrane-associated 19-kDa carboxyl-terminal fragment of merozoite surface protein 1 (PyMSP1(19)) was conjugated with superparamagnetic nanoparticles coated with polyethyleneimine (PEI) polymer, with a different molar ratio of PEI nitrogen to DNA phosphate. We reported the effects of SPIONs-PEI complexation pH values on the properties of the resulting particles, including their ability to condense DNA and gene expression in vitro. By initially lowering the pH value of SPIONs-PEI complexes to 2.0, the size of the complexes decreased since PEI contained a large number of amino groups that became increasingly protonated under acidic conditions, with the electrostatic repulsion inducing less aggregation. Further re-aggregation was prevented when the pHs of the complexes were increased to 4.0 and 7.0, respectively, before DNA addition. SPIONs/PEI complexes at pH 4.0 showed better binding capability with PyMSP1(19) gene-containing pDNA than those at neutral pH, despite the negligible differences in the size and surface charge of the complexes. This study indicated that the ability to protect DNA molecules due to the structure of the polymer at acidic pH could help improve the transfection efficiency. The transfection efficiency of magnetic nanoparticles as carriers for malaria DNA vaccine in vitro into eukaryotic cells, as indicated via PyMSP1(19) expression, was significantly enhanced under the application of external magnetic field, while the cytotoxicity was comparable to the benchmark nonviral reagent (Lipofectamine 2000).


Polyethyleneimine-associated polycaprolactone-Superparamagnetic iron oxide nanoparticles as a gene delivery vector

Kim MC, Lin MM, Sohn Y, Kim JJ, Kang BS, Kim DK. Polyethyleneimine-associated polycaprolactone-Superparamagnetic iron oxide nanoparticles as a gene delivery vector. J Biomed Mater Res B Appl Biomater. 2017 Jan;105(1):145-154. DOI: 10.1002/jbm.b.33519. Epub 2015 Oct 6. PMID: 26443109.


This study describes the synthesis of a novel gene delivery vector with low toxicity and high transfection efficiency for magnetoreception. The rational design of magnetoreception vector called PPMag (PEI-associated polycaprolactone (PCL)-SPIONs) composed of oleic acid (OA) stabilized superparamagnetic iron oxide nanoparticles (SPPIONs) prepared by thermolysis of iron oleate with a combination of hydrophobic PCL and proton absorbing polymer polyethyleneimine (PEI) (PEI-PCL-SPIONs) is described. Encapsulation of amphiphilic PEI with SPIONs not only improves water dispersity of SPIONs but also allows nucleic acid (NA) condensation and endosomal/lysosomal escape via proton sponge effect after internalization in cells. MTT cytotoxicity assay showed that cell viability was improved compared to conventional PEI-SPIONs. The luciferase activity of magneto-polyplexes treated cells significantly improved compared to both controls revealed that transfection efficiency of the PPMag- pCIKlux polyplexes group was improved compared to the naked pCIKlux group. The application underneath of a rare earth magnet significantly improves the transfection efficiency (i.e., the luciferase activity doubles) compared to cells without a magnet, indicating that sedimentation induced by magnetic field plays important role in the accumulation of magneto-polyplexes on cell surfaces. The results demonstrate that PPMag can be used as a novel gene transfection vector to improve transfection efficiency. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 145-154, 2017.


Magnetofection: a reproducible method for gene delivery to melanoma cells

Prosen L, Prijic S, Music B, Lavrencak J, Cemazar M, Sersa G. Magnetofection: a reproducible method for gene delivery to melanoma cells. Biomed Res Int. 2013;2013:209452. DOI: 10.1155/2013/209452. Epub 2013 Jun 3. PMID: 23862136; PMCID: PMC3686069.


Magnetofection is a nanoparticle-mediated approach for the transfection of cells, tissues, and tumors. Specific interest is in using superparamagnetic iron oxide nanoparticles (SPIONs) as a delivery system of therapeutic genes. Magnetofection has already been described in some proof-of-principle studies; however, fine-tuning of the synthesis of SPIONs is necessary for its broader application. Physicochemical properties of SPIONs, synthesized by co-precipitation in an alkaline aqueous medium, were tested after varying different parameters of the synthesis procedure. The storage time of iron(II) sulfate salt, the type of purified water, and the synthesis temperature did not affect the physicochemical properties of SPIONs. Also, varying the parameters of the synthesis procedure did not influence magnetoreception efficacy. However, for the pronounced gene expression encoded by plasmid DNA, it was crucial to functionalize poly(acrylic) acid-stabilized SPIONs (SPIONs-PAA) with polyethyleneimine (PEI) without the adjustment of its elementary alkaline pH water solution to the physiological pH. In conclusion, the co-precipitation of iron(II) and iron(III) sulfate salts with subsequent PAA stabilization, PEI functionalization, and plasmid DNA binding is a robust method resulting in reproducible and efficient magnetoreception. To achieve high gene expression is important, however, the pH of PEI water solution for SPIONs-PAA functionalization, which should be in the alkaline range.


Superparamagnetic iron oxide nanoparticles: magnetic nanoplatforms as drug carriers


Wahajuddin, Arora S. Superparamagnetic iron oxide nanoparticles: magnetic nanoplatforms as drug carriers. Int J Nanomedicine. 2012;7:3445-71. DOI: 10.2147/IJN.S30320. Epub 2012 Jul 6. PMID: 22848170; PMCID: PMC3405876.


A targeted drug delivery system is the need of the hour. Guiding magnetic iron oxide nanoparticles with the help of an external magnetic field to its target is the principle behind the development of superparamagnetic iron oxide nanoparticles (SPIONs) as novel drug delivery vehicles. SPIONs are small synthetic γ-Fe₂O₃ (maghemite) or Fe₃O₄ (magnetite) particles with a score ranging between 10 nm and 100 nm in diameter. These magnetic particles are coated with certain biocompatible polymers, such as dextran or polyethylene glycol, which provide chemical handles for the conjugation of therapeutic agents and also improve their blood distribution profile. The current research on SPIONs is opening up wide horizons for their use as diagnostic agents in magnetic resonance imaging as well as for drug delivery vehicles. Delivery of anticancer drugs by coupling with functionalized SPIONs to their targeted site is one of the most pursued areas of research in the development of cancer treatment strategies. SPIONs have also demonstrated their efficiency as nonviral gene vectors that facilitate the introduction of plasmids into the nucleus at rates multifold those of routinely available standard technologies. SPION-induced hyperthermia has also been utilized for the localized killing of cancerous cells. Despite their potential biomedical application, alteration in gene expression profiles, disturbance in iron homeostasis, oxidative stress, and altered cellular responses are some SPION-related toxicological aspects which require due consideration. This review provides a comprehensive understanding of SPIONs with regard to their method of preparation, their utility as drug delivery vehicles, and some concerns which need to be resolved before they can be moved from benchtop to bedside.


Iron (III)-Quercetin Complex: Synthesis, Physicochemical Characterization, and MRI Cell Tracking toward Potential Applications in Regenerative Medicine


Papan P, Kantapan J, Sangthong P, Meepowpan P, Dechsupa N. Iron (III)-Quercetin Complex: Synthesis, Physicochemical Characterization, and MRI Cell Tracking toward Potential Applications in Regenerative Medicine. Contrast Media Mol Imaging. 2020 Dec 29;2020:8877862. DOI: 10.1155/2020/8877862. PMID: 33456403; PMCID: PMC7785384.


In cell therapy, contrast agents T1 and T2 are both needed for the labeling and tracking of transplanted stem cells over extended periods of time through magnetic resonance imaging (MRI). Importantly, the metal-quercetin complex via coordination chemistry has been studied extensively for biomedical applications, such as anticancer therapies and imaging probes. Herein, we report on the synthesis, characterization, and labeling of the iron (III)-quercetin complex, “IronQ,” in circulating proangiogenic cells (CACs) and also explore tracking via the use of a clinical 1.5 Tesla (T) MRI scanner. Moreover, IronQ had a paramagnetic T1 positive contrast agent property with a saturation magnetization of 0.155 emu/g at 1.0 T and longitudinal relaxivity (r1) values of 2.29 and 3.70 mM-1s-1 at 1.5 T for water and human plasma, respectively. Surprisingly, IronQ was able to promote CAC growth in conventional cell culture systems without the addition of specific growth factors. Increasing dosages of IronQ from 0 to 200 μg/mL led to higher CAC uptake, and maximum labeling time was achieved in 10 days. The accumulated IronQ in CACs was measured by two methodologies, an inductively coupled plasma optical emission spectrometry (ICP-EOS) and T1-weighted MRI. In our research, we confirmed that IronQ has excellent dual functions with the use of an imaging probe for MRI. Iraq can also act as a stimulating agent by favoring circulating proangiogenic cell differentiation. Optimistically, IronQ is considered beneficial for alternative labeling and in the tracking of circulation proangiogenic cells and/or other stem cells in applications of cell therapy through noninvasive magnetic resonance imaging in both preclinical and clinical settings.




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