Work from my group has demonstrated that proteins determine the fate of PS ASOs in all biological systems, identified the key proteins involved in cellular pharmacodynamics, pharmacokinetics and toxicology, characterized the binding of PS ASO to many of the critical proteins and demonstrated that most cytotoxic PS ASOs in all cells and organs studied are to interactions with paraspeckle proteins and RNAse H1 that delocalize these PS ASO protein aggregates to the nucleolus resulting in nucleolar toxicity and apoptosis. Importantly, we have shown that straightforward chemical modifications can alter protein binding and in the case of toxic PS ASOs, ablate or reduce the cytotoxicity with little effect on potency. In more recent work we have thoroughly evaluated the structure activity relationships associated with reduced cytotoxicity.
We have also shown that PS ASOs alter the fates of many of the proteins to which they bind and forma remarkable number of PS ASO protein-RNA-aggregates. In the current manuscript, we take the important step of characterizing the phase separation properties of toxic PS ASO protein aggregates. Remarkable, we show that solid phase aggregates of toxic PS ASOs and paraspeckle proteins are cytoprotective because some aggregates form micro-nucleolus-like aggregates that can transcribe and process pre-ribosomal RNA.