Primary myelofibrosis (PMF) is a clonal blood disorder characterized by mutually exclusive driver mutations in JAK2, CALR, or MPL genes. To explore the epigenetic impact of these mutations, we analyzed DNA methylation (DNAm) profiles from PMF patients. Notably, no differences were found in DNAm between JAK2 and CALR mutated cases, whereas MPL mutations displayed slightly distinct patterns. Furthermore, induced pluripotent stem cell (iPSC) models with JAK2 mutations indicated only a moderate association with PMF-related epigenetic changes, suggesting that these alterations may not be directly driven by the mutations themselves. Additionally, PMF-associated epigenetic changes showed minimal correlation with allele burden and were largely influenced by shifts in the cellular composition. PMF DNAm profiles compared with those from other myeloid malignancies - such as acute myeloid leukemia, juvenile myelomonocytic leukemia, and myelodysplastic syndrome – showed numerous overlapping changes, making it difficult to distinguish PMF based on individual CpGs. However, a PMF score created by combining five CpGs was able to discern PMF from other diseases in both training and validation datasets. These findings demonstrate that PMF driver mutations do not directly evoke epigenetic changes. While PMF shares certain epigenetic alterations with other myeloid malignancies, epigenetic signatures can distinguish between PMF and related diseases.

W.W. and V.T. are involved in the company Cygenia GmbH (www.cygenia.com) that can provide service for epigenetic analysis to other scientists. Apart from this the authors have no competing interests to declare.

This research was supported by funds from the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) within CRU344/417911533 Untangling and Targeting Mechanisms of Myelofibrosis in Myeloproliferative Neoplasms (W.W., S.K.), 363055819/GRK2415 (W.W.); WA 1706/12-2 (W.W.); WA1706/14-1 (W.W.); ZE 432/10-1 (M.Z.); KO2155/7-1 (S.K.), KO2155/7-2 (S.K.), KO2155/9-2 (S.K) and KO2155/6-1 (S.K); and the ForTra gGmbH fuer Forschungstransfer der Else Kroener-Fresenius-Stiftung (W.W.).

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