Nanocages developed to deliver small interfering RNAs


Small interfering RNAs (siRNAs) are novel therapeutics that can be used to treat a wide range of diseases. This has led to a growing demand for selective, efficient, and safe ways of delivering siRNA in cells. 

Now, in a cooperation between the universities of Amsterdam and Leiden in the Netherlands, researchers have developed dedicated molecular nanocages for siRNA delivery. In a paper in the journal Chem, they present nanocages that are easy to prepare and display tunable siRNA delivery characteristics.

The nanocages were developed in the research group for Homogeneous, Supramolecular and Bio-inspired catalysis of Joost Reek and Bas de Bruin at the University of Amsterdam’s Van ‘t Hoff Institute for Molecular Sciences, with further studies in the group led by Alexander Kros at the Leiden Institute of Chemistry. 

The researchers were motivated by the potential of siRNA in gene therapy, which requires the need for effective delivery systems. They set out to develop nanocages with functional groups at the outside, making the cages capable of binding siRNA strands. As the binding is based on reversible bonds, the siRNA can, in principle, be released in a cellular environment. To explore the delivery characteristics of their nanocages, the researchers performed a laboratory study using various human cancer cells.

A range of nanocages

The nanocages are constructions of small molecular building blocks, so-called ditopic ligands, which are connected using metal atoms. A typical cage consists of 12 metal atoms and 24 ligands, hence the abbreviation M12L24. The researchers designed and synthesized five different ligands to form molecular cages with different siRNA binding affinities. They then prepared a range of siRNA binding nanocages using platinum or palladium as the connecting metal. The palladium nanocages are less stable in a cellular environment, and decomposition is one of the siRNA releasing mechanisms.

Schematic representation of the use of nanocages for siRNA delivery. When binding siRNA (arrow A), the nanocages with a typical diameter of 5-6 nanometre cluster into cage-RNA nanoparticles with a diameter of some 150 nanometres. This is small enough to enter a cell (arrow B) and deliver the siRNA. Image taken from the paper/Universiteit van Amsterdam

After screening nanocage characteristics such as stability and siRNA binding capability, the delivery characteristics were put to the test in assays based on siRNA-mediated green fluorescent protein (GFP) silencing. The cages were used to deliver siRNA to human GFP-expressing cells, so that fluorescence measurements could establish successful siRNA delivery. Two types of human cell lines were used: HeLa and U2OS.

Cage composition determines siRNA delivery

The researchers said they were surprised that they could not only demonstrate satisfying siRNA delivery, but also discovered a differentiation depending on the metal used in the nanocage. Where a platinum-based Pt12L24 nanocage showed highly effective siRNA delivery to U2OS cells, it showed little efficiency for HeLa. By contrast, the palladium-based Pd12L24 nanocage, derived from the same ligand building block, delivered siRNA to HeLa but not to U2OS. Such differentiation could not be observed in experiments where a commercially applied delivery system (lipofectamine) was used. The M12L24 nanocages thus introduce the possibility of tuning siRNA delivery characteristics by tuning the nanocage composition.

Four Pt12L24 nanocage structures investigated in the study, using platinum and a variety of ligands. The first nanocage was also synthesised in a variant using palladium instead of platinum. Image taken from the paper/Universiteit van Amsterdam

In the paper, the researchers consider this unique cell selectivity feature of the nanoparticles a promising addition to the field of targeted RNA gene material delivery, whose full potential is yet to be uncovered. 

Even though the current results were obtained in highly controlled laboratory research, they expect that the tunable RNA delivery of their nanocages will lead to future developments of desirable selective RNA nanomedicines.

siRNA delivery

Several companies working in the siRNA space made progress last year. SiSaf’s lead in house program, SIS-101-ADO, is an ADO2-specific siRNA combined with SiSaf’s Bio-Courier platform, which aims to suppress the expression of the mutant CLCN7 gene and thereby rescue bone mass and quality to nearly normal levels. 

Also, Eleven Therapeutics raised a total of $22 million in seed funding to develop a first-of-its-kind platform that designs siRNAs with ultra-long durability by utilizing high throughput combinatorial chemistry and artificial intelligence.

Eleven’s platform aims to decipher the structure-activity relationship (SAR) of siRNAs.