A self-assembled, modular nucleic acid-based nanoscaffold for multivalent theranostic medicine

Abstract

Rationale : Within the field of personalized medicine there is an increasing focus on designing flexible, multifunctional drug delivery systems that combine high efficacy with minimal side effects, by tailoring treatment to the individual. Methods : We synthesized a chemically stabilized ~4 nm nucleic acid nanoscaffold, and characterized its assembly, stability and functional properties in vitro and in vivo . We tested its flexibility towards multifunctionalization by conjugating various biomolecules to the four modules of the system. The pharmacokinetics, targeting capability and bioimaging properties of the structure were investigated in mice. The role of avidity in targeted liver cell internalization was investigated by flow cytometry, confocal microscopy and in vivo by fluorescent scanning of the blood and organs of the animals. Results : We have developed a nanoscaffold that rapidly and with high efficiency can self-assemble four chemically conjugated functionalities into a stable, in vivo -applicable system with complete control of stoichiometry and site specificity. The circulation time of the nanoscaffold could be tuned by functionalization with various numbers of polyethylene glycol polymers or with albumin-binding fatty acids. Highly effective hepatocyte-specific internalization was achieved with increasing valencies of tri-antennary galactosamine (triGalNAc) in vitro and in vivo . Conclusion : With its facile functionalization, stoichiometric control, small size and high serum- and thermostability, the nanoscaffold presented here constitutes a novel and flexible platform technology for theranostics.

Improved Cancer Targeting by Multimerizing Aptamers on Nanoscaffolds

“…The locked nucleic acid (LNA)-based HJ scaffold 14 was used as a platform for attachment and multimerization of aptamers. It consists of four 12-nt oligonucleotides (Q1, Q2, Q3, and Q4) containing a mixture of 2ʹ-O-methyl (2ʹ-OMe) and LNA nucleotides (indicated in gray and red, respectively, in Figure 1A).…”

“…Q1, Q2, Q3, and Q4 were synthesized by the group of Jesper Wengel (University of Southern Denmark [SDU]), as previously described. 14 All DNA strands were purchased from Sigma-Aldrich.…”

“…We have recently developed a modular platform system termed the Holliday junction (HJ), which is capable of combining up to four different functionalities in a single, well-defined structure. 14 The HJ consists of four 12-nt-long, chemically modified nucleic acid modules with a chemical handle on each 5ʹ end, allowing simple click chemistry-based bioconjugation. This 3-to 4nm large scaffold is highly stable in vivo, pharmacokinetically tunable, and can be targeted to specific organs using small-molecule ligands.…”

“…This 3-to 4nm large scaffold is highly stable in vivo, pharmacokinetically tunable, and can be targeted to specific organs using small-molecule ligands. 14 In this study, we extend the use of the HJ to study the effect of aptamer multimerization for tumor targeting. By using simple copper-free click chemistry, we conjugated one to three aptamers covalently to the chemically modified HJ and demonstrated an increased homing to study the effect on targeting cancer cells in vitro and tumors in vivo.…”

A Wireframe DNA Cube: Antibody Conjugate for Targeted Delivery of Multiple Copies of Monomethyl Auristatin E

“…In its core, the programmability of oligonucleotides allows for precise design of constructs, where chosen modifiers, such as drugs, targeting units and imaging agents, can be precisely placed throughout the structure in a predefined ratio [16] . Multiple different systems have been developed, but currently most of these systems are used for either imaging or delivery of less potent drugs such as the DNA intercalator doxorubicin [17–21] . We envisioned that by combining the use of a DNA nanostructure and an antibody, we would be able to prepare an ADC with high loading, good control of DAR, and use highly potent drugs.…”