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

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 potentially disease modifying. Developing molecules targeting human UBE3A-ATS is challenging because its expression and function is restricted to neurons and lacks 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. Rounds of optimization led to the identification of RO7248824, which selectively and potently reduces UBE3A-ATS, while concomitantly upregulating UBE3A mRNA and protein. These properties held true in both human AS patient- and neurotypical-, as well as cynomolgus monkey-derived neurons. Tool molecules targeting murine UBE3A-ATS , revealed a steep relationship between UBE3A-ATS knock-down and UBE3A mRNA/protein upregulation in both wild-type (WT) and AS Ube3am-/p+ mice, whereby an almost 90% downregulation was required to achieve a 50% reactivation, respectively. This relationship was confirmed in cynomolgus monkeys. Repeated lumbar intrathecal administrations of RO7248824 was well tolerated without adverse in-life effects and produced a robust, long lasting (> 3 months) paternal UBE3A reactivation in multiple monkey brain regions. Our results demonstrate that AS induced pluripotent stem cell derived neurons serve as an excellent translational tool leading to the identification of LNA-modified ASOs with excellent drug-like properties translating to infrequent, intrathecal dosing and serves as the basis for the ongoing clinical development of RO7248824 for AS.