I012 Novel conjugated nucleic acid therapeutics in iPSC-derived 2D and 3D models of Huntington’s disease

<h3>Background</h3> The diverse nucleic acid therapeutic (NAT) -based strategies under investigation for Huntington’s disease (HD), whether through lowering the burden of mutant Huntingtin (HTT) expression or targeting disease-modifying factors such as DNA mismatch repair (MMR) genes, all require productive, safe, stable and specific delivery of ASO to disease-relevant tissues. <h3>Aims</h3> The TransNAT Consortium aims to elucidate the mechanisms underlying productive NAT delivery through a multi-stage iterative approach to NAT development and testing. As part of TransNAT, we are systematically assessing target engagement and toxicity of HTT and MALAT1-lowering. ASOs conjugated to novel ligands with novel linker chemistries. These are first characterised in cell lines followed by human iPSC-derived models alongside mouse models to direct the design of future therapeutic NATs. <h3>Methods/Techniques</h3> We performed high-content screening using qPCR, immunocytochemistry and cell viability assays to identify NATs with promising characteristics by gymnotic uptake in 2D cortical and striatal neuron cultures differentiated from HD and control iPSCs. Promising candidate NATs are progressed to screening in organoid and cortico-striatal assembloid models. <h3>Results/Outcome</h3> We have developed and characterised human ex vivo models of HD in which we are able to interrogate NAT uptake, activity and toxicity. Using this we have screened a panel of novel ASO conjugates with varied linkers. <h3>Conclusions</h3> The advancement of our mechanistic understanding of NAT biology will support advancements not only in the development of HD-targeting nucleic acid-based therapeutics that will be delivered safely to disease-relevant tissues, but also promises to be more widely relevant for other neurodegenerative diseases.