FISH and SNP-Array Karyotyping Improve the Detection of Recurrent Chromosomal Defects Including Del(5q), Monosomy 7, Del(7q), Trisomy 8, and Del(20q) in Myelodysplastic Syndromes.
Authors: Makishima Hideki , Rataul Manjot , Gondek Lukasz P. , Huh Jungwon , Cook James R. , Theil Karl S. , Sekeres Mikkael A , Kuczkowski Elizabeth , Maciejewski Jaroslaw P. 
Published: April 2020

Abstract
Cytogenetic aberrations identified by conventional metaphase cytogenetics (MC) have an important diagnostic and prognostic role in evaluating patients with myelodysplastic syndromes (MDS), and results can affect the choice of therapy. Fluorescence in situ hybridization (FISH) can complement MC by providing information derived from both interphase and metaphase nuclei. However, clinically practical FISH strategies are limited to detection of the most common lesions in MDS, including −5/del(5q), −7/del(7q), del(20q), and trisomy 8. The ability to obtain informative results from interphase nuclei and the relative ease of scoring greater numbers of cells are advantages of FISH as compared to MC. Still, the clinical relevance of small numbers of abnormal cells, apart from detection of residual disease, remains unclear. Single nucleotide polymorphism array (SNP-A)-based karyotyping can reveal genetically unbalanced defects with superior resolution compared to MC and FISH, as well as identify segmental uniparental disomy (UPD) that cannot be detected by either method. We sought to determine whether the overall diagnostic yield for detecting common recurring genetic defects associated with MDS could be improved using a strategy incorporating MC, FISH and SNP-A. Using a standardized approach, we focused our investigation on detection of −5/del(5q), −7/del(7q), trisomy 8, and del(20q). We studied 62 patients, including 42 MDS, 5 MDS/MPD, and 15 secondary AML, with standard MC, FISH probes for chromosomes 5, 7, 8, and 20, and SNP-A karyotyping using Affymetrix 250K and/or 6.0 SNP array platform. The detection rate for del(5q) was 35%, 35%, and 37% by MC, FISH, and SNP-A, respectively. No single method detected all of the defects, and detection rates improved when results of all methods were combined. For example, the rate for detection of del(5q) increased incrementally to 39% (MC+FISH), 44% (MC+SNP-A), 42% (FISH+SNP-A), and 44% when all 3 methods were applied. Similar findings were observed for −7/del(7q), trisomy 8, and −20/del(20q): after combining all methods the detection rates improved from 8% to 17%, from 10% to 17%, and from 8% to 10%, respectively, as compared with MC alone. Discrepant results among these methods were related to poor growth (N=2) and low percentage of positive metaphases (small clonal size; N=2). In addition, small somatic deletions (N=6) and UPD (N=2) were not detected by MC or FISH. Larger defects that were detected by SNP-A (e.g., from 5q14.2 to 5q23.1) did not overlap with either loci 5p15.2 (D5S630) or 5q31 (EGR1) used in the FISH probes. We conclude that metaphase cytogenetics, interphase FISH, and SNP-A are complementary techniques that, when applied and interpreted together, can improve the diagnostic yield for identifying genetic lesions in MDS. SNP-A allows for identification of topographically smaller defects and copy-neutral loss of heterozygosity without a requirement for successful cytogenetic analysis. While FISH affords the ability to quantitate the number of affected cells, it is only useful to screen for specific, known defects of a certain size. Whether novel defects as identified by FISH or SNP-A karyotyping will have prognostic impact or affect the results of therapy is the subject of ongoing investigation.