case_images: PMC10570518_01
case_images: Results of SNP microarray analysis in PMF patient. Whole-genome analysis of SNVs and CN-LOH provided by ChAS software in DNA samples obtained at chronic phase (A) and leukemic conversion (B). Each dot corresponds to a single SNP in the array. The DNA copy number losses (deletions/microdeletions) at the level of chromosomes 7q, 8q, 10p, 11p, and Xp are associated with a loss of signal intensity corresponding to a decrease in Weighted Log2 ratio (black arrows). In sample B, the CN-LOH on chromosome 1 that was acquired at leukemic transformation is shown (black box). (C) Ideogram of chromosome 1 and the region of CN-LOH indicated to the right in purple; also, the affected cytobands are shown. (D) Magnified image of SNP array results of chromosome 1 before and after the occurrence of UPD. No changes in Weighted Log2 ratio are observed. Allele difference and B allele frequency (BAF) plots indicate the presence of two tracks (red boxes) instead of the normal three tracks (black boxes), consistent with the loss of heterozygosity. SNP, single-nucleotide polymorphism; SNVs, single-nucleotide variants; CN-LOH, copy neutral loss of heterozygosity; UPD, uniparental disomy.
case_images: fonc-13-1266996-g002.jpg
case_images: PMC10570518_01_fonc-13-1266996-g002.jpg
case_images: f2
case_images: In both samples, five clinically significant copy number losses according to Online Mendelian Inheritance in Man (OMIM) were detected at regions 7q22.1, 8q11.1-q11.21, 10p12.1-p11.22, 11p14.1-p11.2, and Xp11.4, revealing a complex karyotype ( Figures 2A, B ). Additionally, an acquired UPD (aUPD) at 1p36.3-1p12 (118 Mb), covering almost the entire length of the short arm of chromosome 1, was identified only in the sample obtained at the moment of leukemic conversion ( Figure 2C ). The CN-LOH observed at chromosome 1p is partial, being suggestive of a mosaic aUPD ( Figure 2D ).
case_images: Schematic representation of clonal evolution in PMF patient. CALR del52, NRAS Y64D, and PTPN11 S502 mutations were detected at diagnosis in peripheral blood granulocytes in a heterozygous state according to VAF. At leukemic stage, CALR del52 was detected with a similar VAF in CD34+ cells, NRAS Y64D mutation suffered a transition to a homozygous state, PTPN11 mutation was no longer found, and a new NRAS mutation (G12D) was acquired. The leukemic clone arose from the CALR-mutated ancestral clone. Each circle contains a population of cells; in each cell, mutant alleles are represented by colored points on vertical parallel lines to suggest heterozygous versus homozygous state. PMF, primary myelofibrosis; VAF, variant allele frequency.
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case_images: PMC10570518_01_fonc-13-1266996-g003.jpg
case_images: f3
case_images: B, natural killer cells, monocytes, and dendritic cells. Based on the similar VAF (approximately 50%) of CALR mutation detected in the paired DNA samples, indicating a heterozygous status, it may be assumed that the leukemic clone evolved from the CALR-mutated clone through acquisition of a segmental UPD at chromosome 1p that led to homozygosity of initially detected NRAS Y64D mutation and of an additional NRAS G12D mutation ( Figure 3 ).