ANKRD26-related thrombocytopenia (THC2) is a rare inherited platelet disorder caused by germline variants in the 5′ untranslated region (UTR) of ANKRD26. While prior studies using in vitro models or isolated case reports have suggested impaired megakaryopoiesis as a central mechanism, detailed insights have remained elusive-primarily due to the rarity, fragility, and heterogeneity of megakaryocytes. Here, we present a comprehensive, cross-validated analysis of bone marrow samples from four independent THC2 patients, integrating single-cell transcriptomics and ex vivo functional profiling. Across all patients, we analyzed CD34⁺ hematopoietic stem and progenitor cells (HSPCs) (47,281 THC2-HSPCs vs. 51,907 control cells) and primary megakaryocytes (pMKs) (7,309 THC2-pMKs vs. 5,077 controls), uncovering a consistent pattern of megakaryocyte progenitors (MkP) expansion and a marked reduction in polyploid megakaryocytes-indicating a conserved pathophysiologic phenotype. In our index patient, we identified the 5′UTR single-nucleotide variant in ANKRD26 that led to significantly elevated expression across four megakaryocyte-lineage subsets-spanning multipotent progenitors, common myeloid progenitors, megakaryocyte-erythroid progenitors, and MkPs-as well as in terminally enriched pMKs. Spatial transcriptomics and confocal imaging localized ANKRD26 to the centrosome, implicating it in mitotic regulation during megakaryocyte maturation. Mechanistically, we discovered that elevated ANKRD26 induces apoptosis in polyploid megakaryocytes via JUNB-mediated transcriptional activation of CDKN1A (p21)-operating independently of the canonical p53-PIDDosome axis. This multi-patient study provides the most comprehensive cellular and molecular portrait of ANKRD26-driven thrombocytopenia to date, offering novel insights into defective megakaryopoiesis and identifying candidate therapeutic targets to restore platelet production.
Chen et al. (Thu,) studied this question.