Somatic CNVs and LOH in Acute Myelomonocitic Leukemia
Authors: Romano Alessandra , Barresi Vincenza , Musso Nicola , Palumbo Giuseppe A. , Di Raimondo Francesco , Condorelli Daniele 
Published: April 2020

Abstract
In acute myeloid leukemias (AML) chromosomal aberrations, detectable by conventional cytogenetics or targeted molecular techniques, provide the basis for a classification with prognostic relevance. However, cases with normal cytogenetics and undefined prognosis still constitute the single largest group. Recent advances in genome-wide analysis of submicroscopic DNA segment copy number variations (CNVs) may allow the identification of novel molecular tumor-associated abnormalities in the normal cytogenetics group (somatic CNVs). However, CNVs are also present physiologically in the normal population (germline CNVs) (Redon et al., 2006) and can represent potential predisposition factors in disease. Indeed, CNVs can have dramatic phenotypic consequences as a result of altering gene dosage, disrupting coding sequences, or perturbing long-range gene regulation. We used the last generation of Affymetrix single nucleotide polymorphism (SNP)/CNV microarrays (SNP Array 6.0) containing probes for the detection of CNVs and SNPs, with an inter-marker distance of 680 bases and a resolution power of 100 kb. SNP Array 6.0 Assay kit (Affymetrix, Santa Clara, CA) is able to assess copy number changes (CNVs) at a resolution comparable with data obtained using oligonucleotide-array-comparative genomic hybridization (aCGH) and provides also information on loss of heterozygosity (LOH) of the allelic imbalance and copy number neutral type. In the present communication we report preliminary results of a study aimed to test the ability of such arrays to distinguish tumor-associated somatic CNVs and LOHs from germ-line ones by comparing bone marrow samples from AML patients at diagnosis (&gt;90% blasts) and at the remission phase. So far, 8 M4–M5 FAB subtype AML patients have been studied, 6 females, 2 males (median age 38 years, range 25–51). At diagnosis 4 cases with normal karyotype, 2 cases with trisomies (respectively trisomy 13 in 25% and trisomy 22 in 80% of 20 metaphases), 1 inversion (inv (16) (p13q22)) and 1 balanced translocation (t (6;14) (q27;q23)) were detected by conventional cytogenetic analysis. We obtained arrays with quality control (QC) call rates in excess of 90% in all cases (&gt;95% in 8/13 cases) and MAPD &lt;0.4 (&lt;0.35 in 7/13 arrays), using Genotyping Console Version 2.1 for signal intensity analysis, as recommended by Affymetrix. To obtain copy number and LOH calls we used a predefined reference model file, obtained from 270 healthy individuals (HapMap collection). All samples that were regarded as normal karyotype by chromosomal banding had detectable submicroscopic abnormalities by the SNP/CNV array assay. Results obtained are reported in table 1. We found 13 somatic gains not in overlap with known CNVs deposited in the Toronto Database of Genomic Variants. The only recurrent somatic CNV (2/8 patients) was a gain of 109kb in 7q22.1, where genes MGC57359 and GATS map. Five recurrent germline CNVs have been detected, both at diagnosis and remission samples, which could represent regions determining susceptibility to AML. The trisomy 13 case showed a whole chromosome somatic LOH at chromosome 21. 3/8 patients had an interstitial somatic LOH in 19q13.12 in correspondence with adhesion molecules genes (CEACAM1, MEGF8, PSG 1-6-7, ZNF 526). Finally, we detected an interstitial germline LOH, common to all samples, in 16q22.1, where CBFB (core binding factor beta) gene maps, involved in FAB subtypes evaluated in this study. Although this is an ongoing study, with preliminary results, we think that such genome-wide characterization of sub-microscopic DNA alterations might contribute to the discovery of new markers and target genes, with diagnostic, prognostic or therapeutic relevance.
CNV or LOH per sample Normal karyotype Abnormal karyotype median range median range CNV (diagnosis) 38 16–52 79 18–163 ratio gain/loss at diagnosis 7 6–7 15 4–22 CNV (remission) 24 16–32 49 20–167 ratio gain/loss at remission 1 1–2 9 1–20 germline CNV 18 14–21 33 7–77 somatic CNV 16 2–30 51 3–86 LOH (diagnosis) 299 285–314 316 307–317 LOH (remission) 291 285–297 295 282–318 germline LOH 283 278–289 287 287–308 somatic LOH 16 12–19 20 8–30
Table 1