scholarly journals Tunable Composition of Dynamic Non-Viral Vectors over the DNA Polyplex Formation and Nucleic Acid Transfection

Polymers ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 1313 ◽  
Author(s):  
Lilia Clima ◽  
Bogdan Florin Craciun ◽  
Gabriela Gavril ◽  
Mariana Pinteala
Keyword(s):  
Molecular Weight ◽  
Nucleic Acid ◽  
Viral Vectors ◽  
Viral Vector ◽  
Transfection Efficiency ◽  
Binding Capacity ◽  
Nucleic Acid Delivery ◽  
Force Microscopy ◽  
Transmission Electron

Polyethylene glycol (PEG) functionalization of non-viral vectors represents a powerful tool through the formation of an overall surface charge shielding ability, which is fundamental for efficient nucleic acid delivery systems. The degree of non-viral vector PEGylation and the molecular weight of utilized PEG is crucial since the excessive use of PEG units may lead to a considerable reduction of the DNA-binding capacity and, subsequently, in a reduction of in vitro transfection efficiency. Herein, we report a detailed study on a series of dynamic combinatorial frameworks (DCFs) containing PEGylated squalene, poly-(ethyleneglycol)-bis(3-aminopropyl) of different lengths, and branched low molecular weight polyethylenimine components, reversibly connected in hyperbranched structures, as efficient dynamic non-viral vectors. The obtained frameworks were capable of forming distinct supramolecular amphiphilic architectures, shown by transmission electron microscopy (TEM) and dynamic light scattering (DLS), with sizes and stability depending on the length of PEG units. The interaction of PEGylated DCFs with nucleic acids was investigated by agarose gel retardation assay and atomic force microscopy (AFM), while their transfection efficiency (using pCS2+MT-Luc DNA as a reporter gene) and cytotoxicity were evaluated in HeLa cells. In addition, the data on the influence of the poly-(ethyleneglycol)-bis(3-aminopropyl) length in composition of designed frameworks over transfection efficiency and tolerance in human cells were analyzed and compared.

scholarly journals Graphene Oxide as 2D Platform for Complexation and Intracellular Delivery of siRNA

2018 ◽  
Author(s):  
Irene de Lázaro ◽  
Sandra Vranic ◽  
Domenico Marson ◽  
Artur Filipe Rodrigues ◽  
Maurizio Buggio ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Nucleic Acid ◽  
Gene Silencing ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Unmet Need ◽  
Sharp Decrease ◽  
Intracellular Delivery ◽  
Nucleic Acid Delivery ◽  
Primary Mouse

AbstractThe development of efficient and safe nucleic acid delivery vectors remains an unmet need holding back translation of gene therapy approaches into bedside. Graphene oxide (GO) could help bypass such bottleneck thanks to its large surface area, versatile chemistry and biocompatibility, which could overall enhance transfection efficiency while abolishing some of the limitations linked to the use of viral vectors. Here, we aimed to assess the capacity of bare GO, without any further surface modification, to complex a short double-stranded nucleic acid of biological relevance (siRNA) and mediate its intracellular delivery. GO formed stable complexes with siRNA at 10:1, 20:1 and 50:1 GO:siRNA mass ratios. Complexation was further corroborated by atomistic molecular dynamics simulations. GO:siRNA complexes were promptly internalized in a primary mouse cell culture, as early as 4 h after exposure. At this time point, intracellular siRNA levels were comparable to those provided by a lipid-based transfection reagent that achieved significant gene silencing. Time-lapse tracking of internalized GO and siRNA evidenced a sharp decrease of intracellular siRNA from 4 to 12 h, while GO was sequestered in large vesicles, which may explain the lack of biological effect (i.e. gene silencing) achieved by GO:siRNA complexes. This study underlines the potential of non-surface modified GO flakes to act as 2D siRNA delivery platforms, without the need for cationic functionalization, but warrants further vector optimization to allow effective release of the nucleic acid and achieve efficient gene silencing.

scholarly journals Synergistic Effect of Low Molecular Weight Polyethylenimine and Polyethylene Glycol Components in Dynamic Nonviral Vector Structure, Toxicity, and Transfection Efficiency

Molecules ◽  
2019 ◽  
Vol 24 (8) ◽  
pp. 1460 ◽  
Author(s):  
Bogdan Florin Craciun ◽  
Gabriela Gavril ◽  
Dragos Peptanariu ◽  
Laura Elena Ursu ◽  
Lilia Clima ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Nucleic Acid ◽  
Transfection Efficiency ◽  
Morphological Data ◽  
Low Molecular Weight ◽  
Nonviral Vectors ◽  
Main Challenge ◽  
Vector Structure ◽  
Imine Bond

When studying polyethylenimine derivatives as nonviral vectors for gene delivery, among the important issues to be addressed are high toxicity, low transfection efficiency, and nucleic acid polyplex condensation. The molecular weight of polyethylenimine, PEGylation, biocompatibility and, also, supramolecular structure of potential carrier can all influence the nucleic acid condensation behavior, polyplex size, and transfection efficiency. The main challenge in building an efficient carrier is to find a correlation between the constituent components, as well as the synergy between them, to transport and to release, in a specific manner, different molecules of interest. In the present study, we investigated the synergy between components in dynamic combinatorial frameworks formed by connecting PEGylated squalene, poly-(ethyleneglycol)-bis(3-aminopropyl) and low molecular weight polyethylenimine components to 1,3,5-benzenetrialdehyde, via reversible imine bond, applying a dynamic combinatorial chemistry approach. We report comparative structural and morphological data, DNA binding affinity, toxicity and transfection efficiency concerning the ratio of polyethylenimine and presence or absence of poly-(ethyleneglycol)-bis(3-aminopropyl) in composition of dynamic combinatorial frameworks. In vitro biological assessments have revealed the fact that nonviral vectors containing poly-(ethyleneglycol)-bis(3-aminopropyl) and the lowest amount of polyethylenimine have significant transfection efficiency at N/P 50 ratio and display insignificant cytotoxicity on the HeLa cell line.

Polymeric Vectors for Strategic Delivery of Nucleic Acids

Nano LIFE ◽  
2017 ◽  
Vol 07 (02) ◽  
pp. 1730003
Author(s):  
Andrew Dunn ◽  
Donglu Shi
Keyword(s):  
Molecular Weight ◽  
Transfection Efficiency ◽  
Cell Types ◽  
Primary Amines ◽  
Biological Research ◽  
Specific Cell ◽  
Nucleic Acid Delivery ◽  
Biological Studies

Genomic modification through nucleic acid delivery is a frequently applied method in fundamental biological studies and offers a potent therapeutic strategy for disease treatment and biological research. Delivery of nucleic sequences is therefore an attractive facet of biological nanotechnology as highly specific, efficient, and nonantagonistic delivery is necessary for in vivo and clinical use. Previous vectors have suffered from immunogenic responses, serum dependent inactivation, and cytotoxicity, hindering their translational applicability. Current research in polymeric-based nucleotide delivery strives to offer a highly biocompatible, broad use vector through the utilization of polypeptide and polyamine conjugation that can be easily tailored for specific targeting or wide dissemination. Cross-linking low molecular weight polyamines and lipophilic derivatization for amphiphile creation has lead to improved biocompatibility and transfection efficiency compared to higher molecular weight polyamines. Derivatization of hyperbranched and dendritic polyamido- and polyamines has allowed for the formation of efficient in vivo transfection vectors; ring opening synthesis of N-carboxyanhydride amino acids have led to controlled peptide architectures for improved transfection while simultaneously providing convenient primary amines useful in functionalization. Polymer libraries of poly(ß-amino esters) have provided insights into useful architectures for in vitro and in vivo gene delivery. Grafting small molecules to polyamines, such as folate and galactose, for enhanced interaction with cell surface receptors for selective targeting of specific cell types has proven to be encouraging and remains a prominent aspect in biological nanotechnology.

scholarly journals 1,28-Di[(cholest-5-en-3β-yl)disulfanyl]-4,25-dioxo-3,8,12,17,21,26-hexaazaoctacosane tetrahydrochloride

2018 ◽  
Author(s):  
Pavel A. Puchkov ◽  
Elena V. Shmendel ◽  
Valeria D. Andreeva ◽  
Nina G. Morozova ◽  
Marina A. Zenkova ◽  
...  
Keyword(s):  
Thin Film ◽  
Gene Therapy ◽  
Nucleic Acid ◽  
Transfection Efficiency ◽  
Cationic Liposomes ◽  
Hek293 Cells ◽  
Nucleic Acid Delivery ◽  
Promising Candidate ◽  
Effective Delivery

The absence of highly effective delivery systems is a major challenge for gene therapy. Our work was aimed at the development of novel cationic liposomes possessing high transfection efficiency. For this purpose, a novel disulfide polycationic amphiphile 2S4 was synthesized. Cationic liposomes based on 2S4 and a helper lipid DOPE were formed by the thin film hydration method and exhibited effective pDNA delivery into the HEK293 cells, with a maximal transfection activity superior to that of the commercial agent Lipofectamine® 2000. Our results suggest that the polycationic amphiphile 2S4 is a promising candidate for in vitro nucleic acid delivery.

scholarly journals Peptide-Based Nanoparticles for Therapeutic Nucleic Acid Delivery

Biomedicines ◽  
2021 ◽  
Vol 9 (5) ◽  
pp. 583
Author(s):  
Prisca Boisguérin ◽  
Karidia Konate ◽  
Emilie Josse ◽  
Eric Vivès ◽  
Sébastien Deshayes
Keyword(s):  
Nucleic Acid ◽  
Delivery System ◽  
Viral Vectors ◽  
Positive Charge ◽  
Cell Penetrating Peptides ◽  
Nucleic Acid Delivery ◽  
Universal Approach ◽  
Biological Aspects

Gene therapy offers the possibility to skip, repair, or silence faulty genes or to stimulate the immune system to fight against disease by delivering therapeutic nucleic acids (NAs) to a patient. Compared to other drugs or protein treatments, NA-based therapies have the advantage of being a more universal approach to designing therapies because of the versatility of NA design. NAs (siRNA, pDNA, or mRNA) have great potential for therapeutic applications for an immense number of indications. However, the delivery of these exogenous NAs is still challenging and requires a specific delivery system. In this context, beside other non-viral vectors, cell-penetrating peptides (CPPs) gain more and more interest as delivery systems by forming a variety of nanocomplexes depending on the formulation conditions and the properties of the used CPPs/NAs. In this review, we attempt to cover the most important biophysical and biological aspects of non-viral peptide-based nanoparticles (PBNs) for therapeutic nucleic acid formulations as a delivery system. The most relevant peptides or peptide families forming PBNs in the presence of NAs described since 2015 will be presented. All these PBNs able to deliver NAs in vitro and in vivo have common features, which are characterized by defined formulation conditions in order to obtain PBNs from 60 nm to 150 nm with a homogeneous dispersity (PdI lower than 0.3) and a positive charge between +10 mV and +40 mV.

Non-Viral Vectors for Gene Delivery

2018 ◽  
Vol 9 (1) ◽  
pp. 4-11 ◽  
Author(s):  
Aparna Bansal ◽  
Himanshu
Keyword(s):  
Gene Therapy ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
Expression System ◽  
Target Cells ◽  
Specific Expression ◽  
Naked Dna

Introduction: Gene therapy has emerged out as a promising therapeutic pave for the treatment of genetic and acquired diseases. Gene transfection into target cells using naked DNA is a simple and safe approach which has been further improved by combining vectors or gene carriers. Both viral and non-viral approaches have achieved a milestone to establish this technique, but non-viral approaches have attained a significant attention because of their favourable properties like less immunotoxicity and biosafety, easy to produce with versatile surface modifications, etc. Literature is rich in evidences which revealed that undoubtedly, non–viral vectors have acquired a unique place in gene therapy but still there are number of challenges which are to be overcome to increase their effectiveness and prove them ideal gene vectors. Conclusion: To date, tissue specific expression, long lasting gene expression system, enhanced gene transfection efficiency has been achieved with improvement in delivery methods using non-viral vectors. This review mainly summarizes the various physical and chemical methods for gene transfer in vitro and in vivo.

scholarly journals Synthetic Histidine-Rich Peptides Inhibit Candida Species and Other Fungi In Vitro: Role of Endocytosis and Treatment Implications

2006 ◽  
Vol 50 (8) ◽  
pp. 2797-2805 ◽  
Author(s):  
Jingsong Zhu ◽  
Paul W. Luther ◽  
Qixin Leng ◽  
A. James Mixson
Keyword(s):  
Nucleic Acid ◽  
Antifungal Activity ◽  
Antifungal Agents ◽  
Fungal Growth ◽  
Nucleic Acid Delivery ◽  
Histatin 5 ◽  
Ph Buffering ◽  
Endosomal Acidification

ABSTRACT A family of histidine-rich peptides, histatins, is secreted by the parotid gland in mammals and exhibits marked inhibitory activity against a number of Candida species. We were particularly interested in the mechanism by which histidine-rich peptides inhibit fungal growth, because our laboratory has synthesized a variety of such peptides for drug and nucleic acid delivery. In contrast to naturally occurring peptides that are linear, peptides made on synthesizers can be varied with respect to their degrees of branching. Using this technology, we explored whether histidine-lysine (HK) polymers of different complexities and degrees of branching affect the growth of several species of Candida. Polymers with higher degrees of branching were progressively more effective against Candida albicans, with the four-branched polymer, H2K4b, most effective. Furthermore, H2K4b accumulated efficiently in C. albicans, which may indicate its ability to transport other antifungal agents intracellularly. Although H2K4b had greater antifungal activity than histatin 5, their mechanisms were similar. Toxicity in C. albicans induced by histatin 5 or branched HK peptides was markedly reduced by 4,4′-diisothiocyanato-stilbene-2,2′-disulfonate, an inhibitor of anion channels. We also determined that bafilomycin A1, an inhibitor of endosomal acidification, significantly decreased the antifungal activity of H2K4b. This suggests that the pH-buffering and subsequent endosomal-disrupting properties of histidine-rich peptides have a role in their antifungal activity. Moreover, the ability of the histidine component of these peptides to disrupt endosomes, which allows their escape from the lysosomal pathway, may explain why these peptides are both effective antifungal agents and nucleic acid delivery carriers.

IMMU-53. STING-ING GLIOBLASTOMA WITH SPHERICAL NUCLEIC ACIDS

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi104-vi105
Author(s):  
Akanksha Mahajan ◽  
Lisa Hurley ◽  
Serena Tommasini-Ghelfi ◽  
Corey Dussold ◽  
Alexander Stegh ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
High Surface ◽  
Biological Properties ◽  
Innate Immune ◽  
Limiting Factors ◽  
Nucleic Acid Delivery ◽  
Sting Pathway ◽  
Spherical Nucleic Acids

Abstract The Stimulator of Interferon Genes (STING) pathway represents a major innate immune sensing mechanism for tumor-derived DNA. Modified cyclic dinucleotides (CDNs) that mimic the endogenous STING ligand cGAMP are currently being explored in patients with solid tumors that are amenable to intratumoral delivery. Inadequate bioavailability and insufficient lipophilicity are limiting factors for clinical CDN development, in particular when consideration is given to systemic administration approaches. We have shown that the formulation of oligonucleotides into Spherical Nucleic Acid (SNA) nanostructures, i.e.,the presentation of oligonucleotides at high density on the surface of nanoparticle cores, lead to biochemical and biological properties that are radically different from those of linear oligonucleotides. First-generation brain-penetrant siRNA-based SNAs (NCT03020017, recurrent GBM) have recently completed early clinical trials. Here, we report the development of a STING-agonistic immunotherapy by targeting cGAS, the sensor of cytosolic dsDNA upstream of STING, with SNAs presenting dsDNA at high surface density. The strategy of using SNAs exploits the ability of cGAS to raise STING responses by delivering dsDNA and inducing the catalytic production of endogenous CDNs. SNA nanostructures carrying a 45bp IFN-simulating dsDNA oligonucleotide, the most commonly used and widely characterized cGAS activator, potently activated the cGAS-STING pathway in vitro and in vivo. In a poorly immunogenic and highly aggressive syngeneic mouse glioma model, in which tumours were well-established, only one dose of intranasal treatment with STING-SNAs decelerated tumour growth, improved survival and importantly, was well-tolerated. Our use of SNAs addresses the challenges of nucleic acid delivery to intracranial tumor sites via intranasal route, exploits the binding of dsDNA molecules on the SNA surface to enhance the formation of a dimeric cGAS:DNA complex and establishes cGAS-agonistic SNAs as a novel class of immune-stimulatory modalities for triggering innate immune responses against tumor.

scholarly journals Layered Double Hydroxide as a Potent Non-viral Vector for Nucleic Acid Delivery Using Gene-Activated Scaffolds for Tissue Regeneration Applications

Pharmaceutics ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 1219
Author(s):  
Lara S. Costard ◽  
Domhnall C. Kelly ◽  
Rachael N. Power ◽  
Christopher Hobbs ◽  
Sonia Jaskaniec ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Layered Double Hydroxide ◽  
Viral Vectors ◽  
Viral Vector ◽  
Nucleic Acid Delivery ◽  
Safe Alternative ◽  
Transfection Efficacy ◽  
Mass Ratios ◽  
Low Cytotoxicity ◽  
Double Hydroxide

Nonviral vectors offer a safe alternative to viral vectors for gene therapy applications, albeit typically exhibiting lower transfection efficiencies. As a result, there remains a significant need for the development of a nonviral delivery system with low cytotoxicity and high transfection efficacy as a tool for safe and transient gene delivery. This study assesses MgAl-NO3 layered double hydroxide (LDH) as a nonviral vector to deliver nucleic acids (pDNA, miRNA and siRNA) to mesenchymal stromal cells (MSCs) in 2D culture and using a 3D tissue engineering scaffold approach. Nanoparticles were formulated by complexing LDH with pDNA, microRNA (miRNA) mimics and inhibitors, and siRNA at varying mass ratios of LDH:nucleic acid. In 2D monolayer, pDNA delivery demonstrated significant cytotoxicity issues, and low cellular transfection was deemed to be a result of the poor physicochemical properties of the LDH–pDNA nanoparticles. However, the lower mass ratios required to successfully complex with miRNA and siRNA cargo allowed for efficient delivery to MSCs. Furthermore, incorporation of LDH–miRNA nanoparticles into collagen-nanohydroxyapatite scaffolds resulted in successful overexpression of miRNA in MSCs, demonstrating the development of an efficacious miRNA delivery platform for gene therapy applications in regenerative medicine.

scholarly journals Role of Lipid-Based and Polymer-Based Non-Viral Vectors in Nucleic Acid Delivery for Next-Generation Gene Therapy

Molecules ◽  
2020 ◽  
Vol 25 (12) ◽  
pp. 2866 ◽  
Author(s):  
Aniket Wahane ◽  
Akaash Waghmode ◽  
Alexander Kapphahn ◽  
Karishma Dhuri ◽  
Anisha Gupta ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Nucleic Acid ◽  
Messenger Rna ◽  
Viral Vectors ◽  
Cationic Lipids ◽  
Nucleic Acid Delivery ◽  
Nucleic Acid Analogs ◽  
Organ Specific ◽  
Interfering Rna

The field of gene therapy has experienced an insurgence of attention for its widespread ability to regulate gene expression by targeting genomic DNA, messenger RNA, microRNA, and short-interfering RNA for treating malignant and non-malignant disorders. Numerous nucleic acid analogs have been developed to target coding or non-coding sequences of the human genome for gene regulation. However, broader clinical applications of nucleic acid analogs have been limited due to their poor cell or organ-specific delivery. To resolve these issues, non-viral vectors based on nanoparticles, liposomes, and polyplexes have been developed to date. This review is centered on non-viral vectors mainly comprising of cationic lipids and polymers for nucleic acid-based delivery for numerous gene therapy-based applications.

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