# Single-dose administration of therapeutic divalent siRNA targeting MECP2 prevents lethality for one year in an MECP2 duplication mouse model

## TL;DR

A single dose of siRNA targeting MECP2 gene expression prevents death and improves symptoms in a mouse model of MECP2 duplication syndrome for up to a year.

## Contribution

A novel siRNA strategy is introduced for MECP2 silencing with long-term efficacy and potential safety in treating MECP2 duplication syndrome.

## Key findings

- Six lead siRNA candidates achieved 25-75% MECP2 expression reduction for at least four months after a single dose.
- A single siRNA dose fully rescued early mortality and behavioral impairments in a severe MECP2 duplication mouse model.
- Isoform-selective siRNA targeting may prevent Rett Syndrome in mild MDS cases.

## Abstract

MECP2 duplication syndrome (MDS) is a rare X-linked neurodevelopmental disorder caused by duplications of the dosage-sensitive methyl-CpG-binding protein 2 (MECP2) gene. Developing effective therapies for MDS is particularly challenging due to the variability in MECP2 expression among patients and the potential risk of inducing Rett syndrome through excessive pharmacological intervention. Reducing dosage to optimize silencing levels often compromises durability and necessitates increased dosing frequency. We present here a series of fully chemically modified small interfering RNAs (siRNAs) designed for both isoform-selective and total MECP2 silencing. Among these, we identify six lead siRNA candidates across two distinct chemical scaffolds, achieving targeted total MECP2 expression reductions ranging from 25% to 75%, sustained for at least four months following a single administration. The efficacy and safety of human ortholog silencing were evaluated using two mouse models with distinct levels of human MECP2 transgene expression. In the severe duplication model, a single dose of the total isoform-silencing siRNA fully rescued early mortality and behavioral impairments. Additionally, we show that the isoform-selective targeting strategy may be safer in mild cases of MDS where exaggerated pharmacology may lead to Rett Syndrome. Overall, this study introduces a series of preclinical candidates with the capacity to address the varying levels of MECP2 duplication encountered in clinical settings. Furthermore, it establishes a target selection strategy that may be applied to other dosage-sensitive gene imbalances.

Therapeutic siRNAs provide safe and durable modulation of MECP2 for the treatment of mild and severe MECP2 Duplication Syndrome.