Functional classification of RUNX1 variants in familial platelet disorder with associated myeloid malignancies

Decker, Melanie; Lammens, Tim; Ferster, Alina; Erlacher, Miriam; Yoshimi, Ayami; Niemeyer, Charlotte M.; Ernst, Martijn P. T.; Raaijmakers, Marc H.G.P.; Duployez, Nicolas; Flaum, Andreas; Steinemann, Doris; Schlegelberger, Brigitte; Illig, Thomas; Ripperger, Tim

doi:10.1038/s41375-021-01200-w

Cited by 12 publications

(13 citation statements)

References 15 publications

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“…We tried to mitigate these issues by including WT RUNX1 as control and our assays indeed confirmed the effects of known pathogenic variants (Figure 2A, B, and E, p.R204Q). Non-hematopoietic cells such as HEK293T and HeLa have been used extensively to study RUNX1 function (24,50,51), but one could easily argue against their tissue relevance in the context of RUNX1-related leukemia, which is why we performed extensive experiments to confirm RUNX1 function in T-ALL cell lines and also in human cord blood CD34+ cells. Recent development of gene editing techniques in induced pluripotent stem cell (iPSC) provides an exciting system for this type of work (42,52).…”

Section: Runx1 and Jak3 Mutation Induced Etp T-all In Micementioning

confidence: 99%

Germline RUNX1 variation and predisposition to childhood acute lymphoblastic leukemia

Yang

Devidas

et al. 2021

Journal of Clinical Investigation

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Genetic alterations in the RUNX1 gene are associated with benign and malignant blood disorders, particularly of megakaryocyte and myeloid lineages. The role of RUNX1 in acute lymphoblastic leukemia (ALL) is less clear, particularly how germline genetic variation influences the predisposition to this type of leukemia. Sequencing 4,836 children with B-ALL and 1,354 cases of T-ALL, we identified 31 and 18 germline RUNX1 variants, respectively. RUNX1 variants in B-ALL consistently showed minimal damaging effects. By contrast, 6 T-ALL-related variants result in drastic loss of RUNX1 activity as a transcription activator in vitro. Ectopic expression of dominantnegative RUNX1 variants in human CD34+ cells repressed differentiation into erythroid, megakaryocytes, and T cells, while promoting myeloid cell development. Chromatin immunoprecipitation sequencing of T-ALL models showed distinctive patterns of RUNX1 binding by variant proteins. Further whole genome sequencing identified JAK3 mutation as the most frequent somatic genomic abnormality in T-ALL with germline RUNX1 variants. Co-introduction of RUNX1 variant and JAK3 mutation in hematopoietic stem and progenitor cells in mice gave rise to T-ALL with early T-cell precursor phenotype. Taken together, these results indicated that RUNX1 is an important predisposition gene for T-ALL and pointed to novel biology of RUNX1mediated leukemogenesis in the lymphoid lineages.

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Section: Runx1 and Jak3 Mutation Induced Etp T-all In Micementioning

confidence: 99%

Germline RUNX1 variation and predisposition to childhood acute lymphoblastic leukemia

Yang

Devidas

et al. 2021

Journal of Clinical Investigation

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“…In summary, we validated the clinical utility of our previously reported TAs 10 by studying additional RUNX1 variants and discussing them in their clinical context. Limitations of TAs regarding variants C-terminal of the codon 308 have to be considered, but, in the context of RUNX1-FPD, this applies only to a minority of observed RUNX1 missense variants.…”

Section: Resultsmentioning

confidence: 57%

“…We analyzed the ability of variants to activate transcription of RUNX1 target genes CSF1R , ETV1 , and MYL9 by applying luciferase reporter assays in nephrogenic HEK293T and hematopoietic HEL cells. 10 As controls, wild-type (WT) RUNX1b, the pathogenic variant Arg139Gln, and the benign variant Leu29Ser were included. The ability to activate transcription was scored into categories according to the MM-VCEP recommendations: (1) <20% (PS3 moderate); (2) 20% to 80%; (3) >80% to 115% (BS3 supporting); and (4) >115% (PS3 supporting) of WT activity.…”

Section: Methodsmentioning

confidence: 99%

“…RUNX1 missense variants must frequently be classified as variants of uncertain significance (VUS) not allowing clinical conclusions because only variants considered as (likely) pathogenic confirm diagnoses. As demonstrated, 10 functional analyses of VUS can contribute to their reclassification to (likely) pathogenic. Independent of patients’ phenotype and family history, transactivation assays (TAs) address RUNX1’s major function and are, therefore, suitable for first-line screening.…”

Section: Introductionmentioning

confidence: 97%

“…Independent of patients’ phenotype and family history, transactivation assays (TAs) address RUNX1’s major function and are, therefore, suitable for first-line screening. 10 In the present study, we evaluate applicability of TAs 10 for different regions of RUNX1 and applied them to 11 RUNX1 variants for independent validation of their clinical utility.…”

Section: Introductionmentioning

confidence: 99%

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Validation and clinical application of transactivation assays forRUNX1variant classification

et al. 2022

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Familial platelet disorder with associated myeloid malignancies (RUNX1-FPD) is caused by heterozygous pathogenic germline variants of RUNX1. In the present study, we evaluate the applicability of transactivation assays to investigate RUNX1 variants in different regions of the protein. We studied 11 variants to independently validate transactivation assays supporting variant classification following the ClinGen Myeloid Malignancies variant curation expert panel guidelines. Variant classification is key for the translation of genetic findings. We showed that new assays need to be developed to assess C-terminal RUNX1 variants. Two variants of uncertain significance (VUS) were reclassified to likely pathogenic. Additionally, our analyses supported the (likely) pathogenic classification of two other variants. We demonstrated functionality of four VUS, but reclassification to (likely) benign was challenging and suggested the need to reevaluate current classification guidelines. Finally, clinical utility of our assays was illustrated in the context of seven families. Our data confirmed RUNX1-FPD suspicion in three families with RUNX1-FPD-specific family history. Whereas for three variants identified in non RUNX1-FPD-typical families, no functional defect was detected. Applying functional assays to support RUNX1 variant classification can be essential for adequate care of index patients and their relatives at risk. It facilitates translation of genetic data into personalized medicine.

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Disorders of platelets and coagulation, thrombosis and blood transfusion

2022

Neonatal Haematology

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Functional classification of RUNX1 variants in familial platelet disorder with associated myeloid malignancies

Cited by 12 publications

References 15 publications

Germline RUNX1 variation and predisposition to childhood acute lymphoblastic leukemia

Germline RUNX1 variation and predisposition to childhood acute lymphoblastic leukemia

Validation and clinical application of transactivation assays forRUNX1variant classification

Disorders of platelets and coagulation, thrombosis and blood transfusion

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