Effect of Terminal Groups of Dendrimers in the Complexation with Antisense Oligonucleotides and Cell Uptake

Abstract

Poly(amidoamine) dendrimers are the most recognized class of dendrimer. Amino-terminated (PAMAM-NH2) and hydroxyl-terminated (PAMAM-OH) dendrimers of generation 4 are widely used, since they are commercially available. Both have different properties, mainly based on their different overall charges at physiological pH. Currently, an important function of dendrimers as carriers of short single-stranded DNA has been applied. These molecules, known as antisense oligonucleotides (asODNs), are able to inhibit the expression of a target mRNA. Whereas PAMAM-NH2 dendrimers have shown to be able to transfect plasmid DNA, PAMAM-OH dendrimers have not shown the same successful results. However, little is known about their interaction with shorter and more flexible molecules such as asODNs. Due to several initiatives, the use of these neutral dendrimers as a scaffold to introduce other functional groups has been proposed. Because of its low cytotoxicity, it is relevant to understand the molecular phenomena involving these types of dendrimers. In this work, we studied the behavior of an antisense oligonucleotide in presence of both types of dendrimers using molecular dynamics simulations, in order to elucidate if they are able to form stable complexes. In this manner, we demonstrated at atomic level that PAMAM-NH2, unlike PAMAM-OH, could form a well-compacted complex with asODN, albeit PAMAM-OH can also establish stable interactions with the oligonucleotide. The biological activity of asODN in complex with PAMAM-NH2 dendrimer was also shown. Finally, we revealed that in contact with PAMAM-OH, asODN remains outside the cells as TIRF microscopy results showed, due to its poor interaction with this dendrimer and cell membranes.

The complex of PAMAM-OH dendrimer with Angiotensin (1–7) prevented the disuse-induced skeletal muscle atrophy in mice

“…), as previously suggested. 37 The anti-atrophic effects of Ang-(1-7) on three typical features of disuse-induced muscle atrophy were recently described, specifically with regard to recovering muscle strength and protecting against the loss of myofibrillar proteins and UPP activation. Interestingly, these effects were reported when Ang-(1-7) was administered through osmotic pumps.…”

PAMAM Dendrimers Cross the Blood–Brain Barrier When Administered through the Carotid Artery in C57BL/6J Mice

“…They were toxic in a dosage-dependent manner, causing holes on the lipid membranes of the cells, ultimately leading to cell death [13,18,19,20,21]. Fortunately, PAMAM dendrimers can easily be designed to reduce or eliminate potential toxic effects by reducing the surface amine groups and replacing them with a more biocompatible group, such as the hydroxyl (–OH) moiety.…”

“…In the present study, both G1 and G4 amine surface dendrimers as well as mixed surface dendrimers, were used for in vitro uptake experiments by primary cortical neurons and glial cells. However, in order to avoid toxicity of purely amine surface dendrimers, as described in various studies [18,19,20,21], only the G4-mixed surface dendrimers were used for in vivo experiments that assessed the ability of the dendrimers to cross the BBB of C57BL/6J mice when administered through the carotid artery. Moreover, the G1 dendrimer (both the 100% amine surface and G1 90/10) has a 2-dimensional planar structure [6] which makes them incapable of carrying any cargo and were used only in in vitro experiments.…”

Bolaamphiphile-based nanocomplex delivery of phosphorothioate gapmer antisense oligonucleotides as a treatment for <em>Clostridium difficile</em>

“…50 The fluorescent single-stranded nucleic acid dye, Oligreen (Thermo Fisher Scientific, Wilmington, DE, USA), does not effectively bind to single-stranded phosphorothioate bases following cationic complexation, leading to a large decrease in fluorescence emission intensity compared with free oligonucleotide. Binding was measured by exclusion of the dye from oligonucleotide binding through a stepwise mixing of bolasomes into triplicate MES-buffered 2 µg/mL gapmer ASO solutions in black 96-well optical bottom microplates (Nunc; Thermo Fisher Scientific, Wilmington, DE, USA).…”