Optical genome mapping enables constitutional chromosomal aberration detection.

Chromosomal aberrations including structural variations (SVs) are a major cause of human genetic diseases. Their detection in clinical routine still relies on standard cytogenetics. Drawbacks of these tests are a very low resolution (karyotyping) and the inability to detect balanced SVs or indicate the genomic localization and orientation of duplicated segments or insertions (copy number variant [CNV] microarrays). Here, we investigated the ability of optical genome mapping (OGM) to detect known constitutional chromosomal aberrations. Ultra-high-molecular-weight DNA was isolated from 85 blood or cultured cells and processed via OGM. A de novo genome assembly was performed followed by structural variant and CNV calling and annotation, and results were compared to known aberrations from standard-of-care tests (karyotype, FISH, and/or CNV microarray). In total, we analyzed 99 chromosomal aberrations, including seven aneuploidies, 19 deletions, 20 duplications, 34 translocations, six inversions, two insertions, six isochromosomes, one ring chromosome, and four complex rearrangements. Several of these variants encompass complex regions of the human genome involved in repeat-mediated microdeletion/microduplication syndromes. High-resolution OGM reached 100% concordance compared to standard assays for all aberrations with non-centromeric breakpoints. This proof-of-principle study demonstrates the ability of OGM to detect nearly all types of chromosomal aberrations. We also suggest suited filtering strategies to prioritize clinically relevant aberrations and discuss future improvements. These results highlight the potential for OGM to provide a cost-effective and easy-to-use alternative that would allow comprehensive detection of chromosomal aberrations and structural variants, which could give rise to an era of "next-generation cytogenetics."

Next-generation cytogenetics: Comprehensive assessment of 52 hematological malignancy genomes by optical genome mapping.

Somatic structural variants (SVs) are important drivers of cancer development and progression. In a diagnostic set-up, especially for hematological malignancies, the comprehensive analysis of all SVs in a given sample still requires a combination of cytogenetic techniques, including karyotyping, FISH, and CNV microarrays. We hypothesize that the combination of these classical approaches could be replaced by optical genome mapping (OGM). Samples from 52 individuals with a clinical diagnosis of a hematological malignancy, divided into simple (<5 aberrations, n = 36) and complex (≥5 aberrations, n = 16) cases, were processed for OGM, reaching on average: 283-fold genome coverage. OGM called a total of 918 high-confidence SVs per sample, of which, on average, 13 were rare and >100 kb. In addition, on average, 73 CNVs were called per sample, of which six were >5 Mb. For the 36 simple cases, all clinically reported aberrations were detected, including deletions, insertions, inversions, aneuploidies, and translocations. For the 16 complex cases, results were largely concordant between standard-of-care and OGM, but OGM often revealed higher complexity than previously recognized. Detailed technical comparison with standard-of-care tests showed high analytical validity of OGM, resulting in a sensitivity of 100% and a positive predictive value of >80%. Importantly, OGM resulted in a more complete assessment than any previous single test and most likely reported the most accurate underlying genomic architecture (e.g., for complex translocations, chromoanagenesis, and marker chromosomes). In conclusion, the excellent concordance of OGM with diagnostic standard assays demonstrates its potential to replace classical cytogenetic tests as well as to rapidly map novel leukemia drivers.

HER2, chromosome 17 polysomy and DNA ploidy status in breast cancer; a translational study.

Breast cancer treatment depends on human epidermal growth factor receptor-2 (HER2) status, which is often determined using dual probe fluorescence in situ hybridisation (FISH). Hereby, also loss and gain of the centromere of chromosome 17 (CEP17) can be observed (HER2 is located on chromosome 17). CEP17 gain can lead to difficulty in interpretation of HER2 status, since this might represent true polysomy. With this study we investigated whether isolated polysomy is present and how this effects HER2 status in six breast cancer cell lines and 97 breast cancer cases, using HER2 FISH and immunohistochemistry, DNA ploidy assessment and multiplex ligation dependent probe amplification. We observed no isolated polysomy of chromosome 17 in any cell line. However, FISH analysis did show CEP17 gain in five of six cell lines, which reflected gains of the whole chromosome in metaphase spreads and aneuploidy with gain of multiple chromosomes in all these cases. In patients' samples, gain of CEP17 indeed correlated with aneuploidy of the tumour (91.1%; p < 0.001). Our results indicate that CEP17 gain is not due to isolated polysomy, but rather due to widespread aneuploidy with gain of multiple chromosomes. As aneuploidy is associated with poor clinical outcome, irrespective of tumour grade, this could improve future therapeutic decision making.