Genetic and functional characterization of inherited complex chromosomal rearrangements in a family with multisystem anomalies

Purpose: Complex chromosomal rearrangements (CCRs) are rare structural variants involving three or more chromosomal break- points. Most de novo reported CCRs pose challenges for diagnosis and management. They often require karyotyping, fluorescence in situ hybridization (FISH), and chromosomal microarray analysis (CMA) for clinical diagnosis because of the limitations of each method. Here we report an inherited exceptionally complex CCR involving 4 chromosomes and 11 breakpoints in a family with multisystem anomalies. Methods: We evaluated the CCRs using karyotyping, FISH, CMA, and two emerging genomic technologies: high-throughput chromosome conformation capture sequencing (Hi-C; aka genomic proximity mapping, GPM) and optical genome mapping (OGM). We also performed functional studies using transcriptome and methylome analyses. Results: The proband, who had intellectual disability and immune deficiency, shared CCRs with her unaffected mother involving chromosomes 1, 7, and 11 by karyotyping. However, CMA revealed a duplication and three deletions in the proband in contrast to her mother's balanced genome. Hi-C (GPM) and OGM detected the CCRs and copy number alterations but also uncovered additional breakpoints at high resolution, including an insertion in 4p and two cryptic rearrangements at 7p. Transcriptome and methylome analyses identified likely biological pathways associated with the proband's phenotypes. Conclusion: Combining cytogenetic and genomic methods provided comprehensive characterization and defined the breakpoints at high resolution in both proband and mother. This underscores the value of novel cytogenetic and genomic techniques in deciphering complex genome rearrangements and the significance of integrative genomic analysis and functional characterization in understanding clinical phenotypes.