Dharmacon’s Accell Delivery Technology Signals Increasing Focus on non-Endosomal siRNA Uptake Strategies

[Update May 8, 2008: Based on a recent patent application by Dharmacon, WO 2008/036825, it appears that Accell siRNAs are in fact very similar in principle to Alnylam's 2004 cholesterol-conjugated siRNA paper in Nature and for which the Australian patent office has now accepted a related patent application (Australian Patent Application No. 2004206255)].

Dharmacon (now part of Thermo Fisher Scientific), the company that rose to prominence as one of the first and trusted siRNA suppliers that understood to maintain their leading position by staying at the cutting-edge of RNAi research, just announced the introduction of Accell siRNAs.

Accell siRNAs may not be just yet another type of modified siRNA, but appear to be inherently cell membrane permeable allowing for straight-forward in vitro gene knockdown in all cell types tested, including the notoriously difficult-to-transfect primary cells, without the need for prior formulation with special transfection reagents. This is in contrast to conventional siRNAs that, due to charge and size, are not cell permeable and are therefore require formulation with delivery reagents such that they may gain access to the cytoplasm (the site of Ago2-mediated RNAi) following endosomal uptake and lysis of that compartment. This can be toxic, either because of the chemistry of the specific formulation or the triggering of cytokine responses by engaging TLR receptors in the endosomes.

It is also clear that although potent gene silencing may be achieved following endosomal uptake, typically only a fraction of the siRNA that had been delivered to the cell is actually active in gene silencing. In fact, the rate-limiting step in vivo in many instances may not be getting the siRNA to the target cell, but getting it into the target cells once in close proximity. In addition to better understanding the endosomal uptake-release mechanism, research into alternative technologies may therefore prove highly rewarding.

Although technical details were not disclosed, it is reasonable to assume that Accell siRNAs have been rendered more cell permeable either by the addition of a small lipophile to the siRNA duplex or by modifying the highly charged siRNA backbone without compromising siRNA silencing activity. Stanley Crooke, the CEO of ISIS, has e.g. noted before that it is the amphiphilic nature of single-stranded antisense nucleic acids facilitates their cellular uptake, so that it is conceivable that Accell siRNAs may have been engineered to be similarly amphiphilic.

The current Accell siRNA technology, however, has some drawbacks in that a special tissue culture media is required and therefore does not allow them to be directly translated to in vivo applications, although formulating them in ways that get them to their target cells should not represent an insurmountable challenge, with small conjugates such as cholesterol being likely candidates. The recommended concentration of 1 micromolar siRNA (about 50-fold higher of what is typically used in vitro) further indicates that the process needs to be optimized before Accell siRNAs will be widely used for large-scale and routine tissue culture experimentation. The immediate market potential should therefore largely come from work with primary cells.

Accell siRNAs reminds us that systemic RNAi delivery is a multi-step process, starting with extravasation of the siRNAs from the blood stream into the tissues to cell attachment and cellular entry, and the potential for improving each step is significant. It is fascinating to see how intensely each of these tasks are being addressed and the many creative solutions coming out of these efforts.