Abstract Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by the loss of neuronal E3 ligase UBE3A with no available treatment. Restoring UBE3A levels via downregulation of the paternally cis-acting long non-coding antisense transcript (UBE3A-ATS) is a potential disease modifying. Developing molecules targeting human UBE3A-ATS is challenging because it is expressed only in neurons and lacks animal species sequence conservation. To overcome this, we performed a library screen of locked-nucleic acid (LNA)-modified antisense oligonucleotides (ASOs) on AS patient-derived neurons, identifying initial sequences. Further optimization led to the identification of the ASO, RO7248824, which selectively and potently reduces UBE3A-ATS, while concomitantly upregulating the UBE3A mRNA and protein. These properties held true in both human AS patient- and neurotypical-, as well as cynomolgus monkey-derived neurons. In vivo use of tool molecules in wild-type (WT) and AS Ube3am-/p+ mice, revealed a steep relationship between UBE3A-ATS knock-down and UBE3A mRNA/protein upregulation, whereby an almost 90% downregulation was needed to achieve a 50% upregulation, respectively. This relationship was confirmed in cynomolgus monkeys. Whereby, repeated lumbar intrathecal administrations of RO7248824 was well tolerated without adverse in-life effects or tissue pathology and produced a robust, long lasting (up to 3 months) paternal reactivation of UBE3A mRNA/protein across key monkey brain regions. Our results demonstrate that AS human pluripotent stem cell neurons serve as an excellent translational tool and furthermore LNA-modified ASOs exhibit excellent drug-like properties. Sustained efficacy translated to infrequent, intrathecal dosing and serves as the basis for the ongoing clinical development of RO7248824 for AS. Graphical Abstract Graphical abstract. From AS patient blood to a neuronal screen, identifies clinical ASO with excellent in vivo properties. ( 1 ) Patients were recruited. ( 2 ) Whereby blood was reprogrammed into hIPSC and subsequently differentiated into neurons. ( 3 ) ASOs were designed and screened on human neurons to downregulate the UBE3A-ATS likely via directed RNase H Cleavage of Nascent Transcripts. ( 4a ) RO7248824 was identified that potently and selective reduces UBE3A-ATS, concomitantly with upregulating the UBE3A sense transcript and protein which was used for in vitro pk/pd. ( 4b ) In parallel tool murine ASO were used demonstrate in vivo POC.(5) Pivotal nonhuman primate studies to monitor safety and predict the human dose. (6) RO7248824 is in AS clinical trial . One Sentence Summary From angelman syndrome human neuron screen to cynomolgus monkey proof of concept identifies the clinical molecule RO7248824

Angelman syndrome patient neuron screen identifies a potent and selective clinical ASO targeting <i>UBE3A-ATS</i> with long lasting effect in cynomolgus monkey

Angelman syndrome patient neuron screen identifies a potent and selective clinical ASO targeting UBE3A-ATS with long lasting effect in cynomolgus monkey