Smith-Magenis Syndrome in a Dish: A Model to Study Disease Mechanisms.

Smith-Magenis syndrome (SMS) is a rare neurodevelopmental disorder caused by RAI1 haploinsufficiency, either due to point mutations in the RAI1 gene or to chromosomal deletions on 17p11.2 that include this gene. Despite the identification of RAI1 as the main causal gene, the cellular mechanisms underlying SMS remain largely unclear. The main objective of this thesis was to explore how RAI1 haploinsufficiency affects cellular homeostasis and neural development, using patient-derived cell models as a platform to study disease mechanisms and identifying potential therapeutic targets and testing pharmacological approaches to rescue disease-related phenotypes. The first part of this work focused on fibroblast lines derived from six SMS patients (three carrying RAI1 mutations and three with 17p11.2 deletions). Part of this work has been published in 2022. Transcriptomic and functional analyses revealed that RAI1 haploinsufficiency alters lipid metabolism, autophagic flux, and lysosomal homeostasis, leading to an accumulation of lipid droplets and impaired degradation pathways. In the second part, induced pluripotent stem cells (hiPSCs) were generated from four patient fibroblast lines and extensively characterized to confirm their pluripotency, genomic stability, and differentiation potential. Finally, hiPSC-derived neural stem cells (hiNSCs) were obtained and differentiated into neural lineages — neurons, astrocytes, and oligodendrocytes — over a 31-day time course. Throughout differentiation, we monitored lipid droplet accumulation, autophagy, lysosomal acidification, and reactive oxygen species (ROS) levels. SMS-derived neural cells displayed aberrant neurogenesis, altered lysosomal function, and persistent metabolic stress, characterized by lipid accumulation and dysregulated ROS production. Together, these findings highlight a critical link between RAI1 deficiency, lysosomal dysfunction, and impaired lipid metabolism, which converge to disrupt neural development and cellular homeostasis in SMS. This integrative disease model provides a valuable platform for investigating disease mechanisms and testing potential therapeutic approaches.