ACS Nano 12, 994−1005 (2018)
The lack of tissue and cell specificity of CRISPR–Cas9 delivery systems represents a major obstacle to the in vivo application of this gene editing technology for therapeutic purposes. The design of nanoparticles for targeted delivery has partially solved the problem, but residual mistargeting raises concerns of side effects arising from the DNA cleaving activity of the Cas9 nuclease in non-targeted cells.
The CRISPR–Cas9 system can typically be delivered in the form of a plasmid encoding the components of the editing machinery and a promoter that drives cellular expression of Cas9. Now, Luo and co-workers substitute the classical universal promoter with a cell-specific promoter that induces activation of CRISPR–Cas9 only in macrophages and monocytes. The system is encapsulated in cationic lipid-assisted polymeric nanoparticles. In vitro, different cells are able to take up the loaded nanoparticles, but only macrophages and monocytes can efficiently express Cas9. As a consequence, gene editing is also restricted to these cell types, as observed in the presence of a guide RNA directed against Nnt1. This gene, which encodes the protein netrin-1, is a potential therapeutic target in macrophages for type 2 diabetes.
Considering the in vitro cell specificity, the authors also perform experiments to investigate whether macrophage-specific CRISPR–Cas9 activation shows promise for therapeutic gene editing in animal models. Indeed, diabetic mice treated with the developed gene editing system directed against Nnt1 show a macrophage-specific decrease of netrin-1 concentration with negligible off-target effects, recapitulating the in vitro results. Moreover, the animals displayed normalized blood glucose levels and improved insulin sensitivity.