Two pathogenetic intronic variants in SPG4/SPAST and expansion of the clinical presentation

• Detection in one Brazilian and in one unrelated Japanese family of two splice-site donor variants in SPAST, with disagreeing classification of pathogenicity. • Functional characterization of the two intronic variants demonstrated that both variants lead to disrupted protein domains and premature stop codons. • In both pedigrees, the identified variants co-segregate symptoms consistent with anorexia nervosa, reporting a formerly unrecognized association between SPAST mutations and disordered eating behaviours. Hereditary Spastic Paraplegia (HSP) is a group of inherited neurodegenerative disorders primarily characterized by progressive lower limb spasticity and weakness. Among the various genetic causes of HSP, pathogenetic variants in SPG4/ SPAST are the most frequently identified, making them the leading molecular cause of autosomal dominant HSP. The SPAST gene encodes spastin, a protein involved in microtubule dynamics. In this study, we focused on the functional characterization of two specific intronic variants in SPAST absent or present at a very low frequency in GnomAD database and with conflicting classification of pathogenicity, c.1245 + 5G > A and c.1493 + 2_1493 + 5del respectively. These variants were identified in two independent families, one of Brazilian origin and the other of Japanese descent. Our data shows that the splicing variants impact splicing. Furthermore, through segregation analysis and clinical assessments, we provided a detailed description of the affected individuals, emphasizing the clinical presentation associated with these genetic changes. Notably, in both families, the identified variants co-segregated symptoms consistent with anorexia nervosa, suggesting a potential, previously unrecognized association between SPAST pathogenic variants and disordered eating behaviors. Our findings contributed to the expanding clinical spectrum of SPG4-associated HSP and highlighted the importance of characterizing intronic SPAST variants. The characterization of intronic pathogenetic variants enhanced our understanding of their potential pathogenic mechanisms, which may have implications for both genetic diagnosis and the broader clinical management of HSP.