Utilization of Next Generation Sequencing to detect and assign pathogenicity to balanced rearrangements identified by conventional cytogenetics. U. Aypar1, K. Pearce1, E. Thorland1, J. Evans2, V. Sarangi2, C. Wang2, Y. Asmann3, N. Hoppman1 1) Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN; 2) Department of Health Sciences Research, Mayo Clinic, Rochester, MN; 3) Department of Health Sciences Research, Mayo Clinic, Jacksonville, FL.

   Balanced rearrangements are found in approximately 1 in 175 phenotypically normal individuals. However, balanced rearrangements can result in an abnormal phenotype due to cryptic deletions or duplications at the breakpoints, disruption of a gene, position effects resulting from the rearrangement, or uniparental disomy. When detected by karyotyping in a patient with abnormal phenotypic features, parental studies are typically performed in order to determine pathogenicity; however even when de novo, such rearrangements are thought to be pathogenic only 6.7% of the time. Chromosomal microarray (CMA) is useful in detecting cryptic imbalances in some cases; however, the majority have normal array results, and in these cases, no additional clinical testing is currently available to further assess pathogenicity. Therefore, we performed next generation sequencing in four patients, three with abnormal phenotypic features and one with a normal phenotype, in whom a de novo balanced rearrangement was detected by conventional chromosome analysis and normal CMA results were obtained. Our goal is to develop a reflex test for patients with de novo balanced rearrangements in order to provide a non-equivocal test result. Samples with a (1;9)(q44;p13)dn, t(2;3)(q23;q27)dn, t(2;8)(q35;q22.1)dn, and inv(6)(p12;q27)dn were sequenced by paired-end whole genome sequencing on the Illumina HiSeq2000. A variety of bioinformatics algorithms designed to detect structural variants were used, including Delly and CREST. In two of the three cases with abnormal phenotype, disruption of a gene with known pathogenic mutations was identified. The third sample had no genes disrupted by the rearrangement. The patient with a normal phenotype had disruption of two genes with no known clinical significance. Based on these findings, we conclude that the clinical utility of next generation sequencing in the context of cytogenetically balanced rearrangements is significant, and incorporation of this technology to routine cytogenetic testing is recommended.
RearrangementBreakpoint 1Breakpoint 2
46,XX,t(1;9)(q44;p13)dn1q44: AKT39p13: no gene
46,XX,t(2;3)(q23;q27)dn2q23: no gene3q27: TP63
46,XX,t(2;8)(q35;q22.1)dn2q35: MREG8q22.1: LINC00534
46,XX,inv(6)(p12;q27)dn6p12: no gene6q27: no gene

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