Targeted sequencing of 61 neurodegeneration genes in 186 Sicilian patients identified pathogenic/likely pathogenic variants in ~21% (diagnostic yield 16.7%), with GBA showing the highest number of pathogenic hits and broad phenotypic associations across AD, PD, FTD and parkinsonism.

Highlights clinically actionable genetic stratification—particularly enrichment of GBA (lysosomal) variants—supporting patient selection, biomarker development, and translational relevance for GCase/lysosome-targeted Parkinson's therapeutics and diagnostics.

Despite the technological advancements in modern genetic diagnosis, customized genetic panels are still frequently employed for diagnostic purposes due to their rapid, efficient, and cost-effective ability to detect genetic variants. In this study, we utilized a customized genetic panel designed to identify genetic variants associated with neurodegenerative disorders. The panel consisted of 61 genes and was applied to a cohort of 186 unrelated individuals diagnosed with different degenerative cognitive and movement disorders. The identified variants were filtered for a minor allele frequency of less than 1% and classified according to the American College of Medical Genetics (ACMG) guidelines. Our results showed that 20.97% of individuals carried at least one likely pathogenic or pathogenic variant, with 35% of those individuals diagnosed with Alzheimer's disease (AD). The positive diagnostic yield of the panel was 16.67%, calculated based on variant zygosity, inheritance pattern, and concordance with the clinical phenotype. Furthermore, 34.41% of the individuals carried variants of uncertain significance (VUS), and 44.62% carried benign variants. Variants have been found only in 58 genes. Among these, 24.14% showed benign variants, 48.28% had VUS, and 27.59% carried pathogenic or likely pathogenic variants. Principal component analysis analysis based on the variables "age at onset" in addition to the phenotypic scores "MMSE" (global cognitive screening test), "IADL" (instrumental activities of daily living), and "ADL" (basic activities of daily living) distributed the individuals associated with the specific disease and variant in the plot. Notably, the individuals showed AD exhibited an average age at onset of 68 ± 12.5 years and were differentiated in the plot. GBA gene exhibited the highest number of pathogenic variants (9) linked to AD, Parkinson's disease, early onset parkinsonism with epilepsy, fronto-temporal dementia, and mild cognitive impairment. The results highlighted a broad phenotypic heterogeneity associated with genes previously linked to only a limited number of neurodegenerative conditions, underscoring the value of the genetic testing performed. Translationally, although clinical exome sequencing has enabled novel gene discovery in neurodegenerative disorders, targeted genetic panels remain a cost-effective and clinically valuable approach for routine diagnostics. In this context, our study highlights the "real-world" utility and clinical impact of a focused panel-based strategy.