Clinical and genomic characterization of corpus callosum abnormalities (CCA) in 107 Tunisian patients using a stepwise diagnostic approach

Background Corpus callosum abnormalities (CCA) represent a heterogeneous group of neurodevelopmental disorders resulting from disturbances in midline patterning, neuronal migration, and axonal guidance. Their genetic architecture includes chromosomal abnormalities, pathogenic copy-number variations (CNVs), and single-gene disorders; however, genotype–phenotype correlations remain incompletely defined, particularly in underrepresented populations. Methods We investigated the clinical and genomic characteristics of 107 Tunisian patients with radiologically confirmed CCA recruited over a 15-year period. A sequential genomic strategy was applied, including conventional karyotyping in all patients, fluorescence in situ hybridization (FISH) and multiplex ligation-dependent probe amplification (MLPA) when clinically indicated, chromosomal microarray analysis (array-CGH) in 54 selected patients, and whole-exome sequencing (WES) in six unresolved cases. CNVs were interpreted according to ACMG/ClinGen recommendations, and sequence variants were classified according to ACMG/AMP criteria. Results Most patients presented syndromic forms of CCA (95.3%) associated with additional cerebral and/or extracerebral manifestations. Clinically relevant genomic abnormalities supporting a molecular diagnosis were identified in 11/107 patients (10.3%). Conventional karyotyping identified chromosomal abnormalities in three patients (2.8%), while array-CGH detected pathogenic or likely pathogenic CNVs in ten of 54 tested patients (18.5%). These CNVs involved genomic regions previously implicated in neurodevelopmental disorders and CCA, including 14q12, 4p16.3–p16.1, 1q43–q44, 1p32.3p31.3 –p32, 5p14.3, 5q35, 16q23.1–q24.3, and 18pterp11.1. WES identified rare variants in WLS , ERCC8 , and VPS39 , while a MID1 variant was classified as likely benign. No sequence variant met ACMG/AMP criteria for pathogenicity, and none was considered diagnostic. Conclusion This study provides a comprehensive characterization of CCA in a Tunisian referral cohort and highlights the major contribution of pathogenic genomic imbalances to the molecular diagnosis of these disorders. The findings emphasize the complementary role of cytogenetic and sequencing approaches while illustrating the challenges of interpreting rare sequence variants in patients with complex neurodevelopmental phenotypes. Larger cohorts with systzbematic genomic testing, segregation analyses, and functional validation will be required to further refine genotype–phenotype correlations in CCA.