Abstract

Using NanoSIMS imaging our group has shown that it is possible to measure the absolute concentration of nucleic acid-based therapeutics in what is generally considered to be the endosomal space. In this field there is a consensus that essentially all (~99%) of the nucleic acid-based therapeutic that enters the cell is trapped in the endosomal space, thus this amount of material is fundamental to the interpretation of downstream events such as endosomal escape and gene silencing. Gene silencing in vitro is measured with respect to incubation concentration. We have shown that this value is fundamentally decoupled from cellular uptake therefore does not provide mechanistic insight. We propose that the intracellular concentration is a key parameter which needs to be considered when the efficacy of a nucleic acid-therapeutic being evaluated. It will serve to remove the ambiguity that arises from the convoluted relationship between exposure and cellular uptake. This presentation will discuss the concepts that enable intracellular quantification using NanoSIMS and highlight the process we have made in applying this technology in the industry setting.