Authors :
Presenting Author: Janine Erler, PhD – NEUmiRNA Therapeutics
Lluís Riera Ponsati, PhD – NEUmiRNA Therapeutics
Celia Paloma Tamayo, MSc – NEUmiRNA Therapeutics
Jordan Higgins, MSc – RCSI
Omar Mamad, PhD – RCSI
Petra Bencúrová, PhD – RCSI
Albert Sanfeliu, PhD – RCSI
Jaideep Kesavan, PhD – RCSI
Sakari Kauppinen, PhD – Aalborg University
Mark McLaughlin, PhD – University of Glasgow
Rodrigo Gutierrez Qunitana, PhD – University of Glasgow
David Henshall, PhD – Royal College of Surgeons Ireland
Henrik Klitgaard, PhD – NEUmiRNA Therapeutics
Rationale:
We have shown that inhibition of microRNA-134 (miR-134) with a proprietary antisense oligo (ASO) developed by NEUmiRNA Therapeutics, NMT.001, results in disease-modification and disease reversal in an NIH-validated mouse model of drug-resistant epilepsy (DRE), in the absence of tachyphylaxis and adverse effects. Interim findings from the CURE-funded METriC large animal veterinary trial indicate similar results when miR-134 is inhibited in dogs with naturally occurring DRE. Additionally, inhibition of miR-134 in brain slices from DRE mice revealed potent antiepileptogenic effects. Here, we investigate the mechanism of action of miR-134 inhibition in achieving this unprecedented disease modification and reversal in DRE.
Methods:
We performed RNA sequencing of neurons treated with NMT.001 to explore potential on/off-target effects and to elucidate the molecular pathways altered. We then validated key molecules in mouse brain samples from the DRE model at the mRNA and protein level and compared findings to miR-134 target genes identified in the hippocampus of dogs and humans with DRE using cross-linked immunoprecipitation of Argonaute-bound RNAs. Finally, we analyzed electrophysiologic activity in brain slices from antimiR-treated animals and human iPSC-derived neurons.
Results:
RNA sequencing confirmed no-off target effects of NMT.001 and revealed the most significantly altered pathways associated with NMT.001 to be involved in the regulation of the neuronal network structure and directly linked with epilepsy. These pathways were found to be conserved among mRNA-134 targets identified in the DRE dog and human samples. Brain slices from DRE mice treated with antimiR-134 displayed reduced hyperexcitability. Under loose-patch recordings, we observed antimiR-134 decreases burst frequency and increases inter-burst intervals in human iPSC-derived neuronal networks while sparing various baseline neurophysiologic parameters.
Conclusions:
The molecular and electrophysiologic results from across 3 different species provide mechanistic insight into how inhibition of miR-134 achieves disease modification, and how this mechanism of action is conserved across species, providing evidence of the high degree of translatability to DRE and human epilepsy.
Funding: Innovation Fund Denmark (JE)
CURE Catalyst Award (RGQ)