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Letters to the Editor
Table 1. Characteristics of patients with identified germline mutations or deletions.
Patient ID
Patient 1
Patient 2
Patient 3
Patient 4
Patient 5
Sex Age FAB WBC BM at subtype count blast Dx (x109/L) count
Cytogenetics
46,XX[30]
47,idem,+15,+20[21] (VAF=50%)
45,XY,-7 [5] GATA2 NM_032638.4:c.1008del (p.Lys336Asnfs*51)
Germline deletion/mutation
CEBPA NM_004364.3: c.65_125del (p.Pro22Leufs*118)
ACMG-AMP clinical significance (rules applied¥)
Somatic Ref. mutations
atAML diagnosis
F 3
F 13
M 16
F 9
F 14
M1 13.4
50
Pathogenic
Likely
pathogenic
Pathogenic
Pathogenic
Pathogenic
CEBPA NM_004364.3: c.934_936dup (p.Gln312dup)
WT1 NM_024426.3: c. 1357_1361delins AGTAG (p.Cys453_Lys454delinsSerArg) (VAF=26%)
None 10
JAK2 NM_004972.3:c. 1849G>T (p.Val617Phe) (VAF=44%) SETBP1 NM_015559.2:c.2602G>T (p.Asp868Tyr) (VAF=42%)
KRAS NM_NM_033360.2: c.38G>A (p.Gly13Asp)(VAF=19%)
None 9
M5 4 77
M0 105 NA
M6 3 18
M6 1.7 21
GATA2 NM_032638.4:c. 7,t(11;19)(q23;p13.3)[3]/ 1114G>A(p.Ala372Thr)
45,XX,-
47,XX,+8[14]/49,XX,+8,
+add (9)(q22),+21[9]
(VAF=48%)
RUNX1
NM_001754.4: c.601C>T
(p.Arg201*) (VAF=48%)
48.XXXc. RUNX1 NM_001754.4 deletion +21[22]/47,XXXc[3] (size:1.8 Mb, CN: 1.0,
mean log2 ratio: -0.54)
¥The interpretation of the pathogenicity of the identified germline alterations was based on the American College of Medical Genetics and Genomics and Association for Molecular Pathology guidelines and the adapted guidelines from the ClinGen Myeloid Malignancy Variant Curation Expert Panel for the RUNX1 gene.12,13 Chromosomal locations are annotated according to human reference assembly GRCh37/hg19. Rules applied to classify the variants are detailed in Online Supplementary Table S4. Dx: diagnosis; FAB: French-American-British classification; WBC: white blood cells; BM: bone marrow; ACMG/AMP: American College of Medical Genetics and Genomics/Association for Molecular Pathology; AML: acute myeloid leukemia; F: female; M: male; CN: copy number state. NA: not available.
ly described.10 Cytoscan HD array (ThermoFisher) was used, according to the manufacturer’s instructions, to screen for gene deletions. The data were analyzed using Chromosome Analysis Suite (ChAS) 3.1 software (ThermoFisher). Since germline specimen collection was not planned by the ELAM02 protocol, blood samples taken at the time of complete remission were used to assess the germline status of variants and deletions by direct Sanger sequencing and SNP-array, respectively. Complete remission was defined as cytological remis- sion associated with a level of WT1 expression below 2% in the bone marrow and below 0.1% in peripheral blood or a core-binding factor transcript expression level below 0.01% in core-binding factor AML. The patho- genic status of the suspected germline alterations was established as per American College of Medical Genetics and Genomics and Association for Molecular Pathology (ACMG-AMP) guidelines.12,13 The study was approved by the Ethics Committee of Saint-Antoine Paris University Hospital (Paris, France) and was conducted in accor- dance with the Declaration of Helsinki.
Investigations were restricted to patients with dele- tions, double pathogenic variants or unique pathogenic variants with a high VAF (a threshold of ≥30% was cho- sen to avoid being too restrictive) in one of the follow- ing genes: CEBPA, ETV6, GATA2, RUNX1 and TP53 at AML diagnosis. Of note, as the cis-trans allelic configu- ration for identified variants was not assessed for this
study, the term double mutation was preferred, when appropriate.
Fifty-two patients (13%) out of the 385 analyzed met at least one of these criteria (45 with pathogenic variants and 7 with targeted deletions) (Online Supplementary Table S2). Among these patients, 17 had CEBPA double pathogenic variants, including one patient with a homozygous pathogenic variant. RUNX1 pathogenic variants were found in 16 patients (4 with double muta- tions and 12 with a VAF ≥30%). GATA2, ETV6 and TP53 pathogenic variants with a VAF ≥30% were identified in 12, three, and two patients respectively. Targeted dele- tions encompassing RUNX1 or ETV6 were found in five and two patients, respectively (size range: 50 kb to 1.8 Mb) (Figure 1). Three of the 52 patients did not reach complete remission, including one patient who died dur- ing induction.
Thirty-eight patients had available DNA samples at the time of complete remission. Direct Sanger sequenc- ing and SNP-array analysis of these samples identified five patients with still detectable alterations comprising four with pathogenic/likely pathogenic variants (CEBPA, n=1; GATA2, n=2; RUNX1, n=1) and one with a patho- genic deletion (RUNX1; size: 1.8 Mb) (Table 1, Online Supplementary Table S3, Online Supplementary Figure S1).
Patient n. 1 was a 3-year old girl who carried a frameshift CEBPA pathogenic variant (NM_004364:c.65_125del (p.Pro22Leufs*118)) (Figure
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