Characterization of ETV6-Related Thrombocytopenia (ETV6-RT): a new form of thrombocytopenia associated with a risk of developing hematologic neoplasms

ETV6-Related Thrombocytopenia (ETV6-RT) is a rare form of inherited autosomal dominant thrombocytopenia first identified in 2015 that, together with ANKRD26-Related Thrombocytopenia (ANKRD26-RT) and Familial Platelet Disorder (FDP/AML), belongs to the category of “myeloid neoplasms with germline predisposition and preexisting platelet disorders” according to the 2016 World Health Organization classification. ETV6-RT is caused by germline mutations in ETV6 gene that encodes a transcriptional repressor known for its role in hematopoiesis and megakaryopoiesis. ETV6 was initially identified as tumor suppressor frequently involved in somatic translocations responsible for leukemia development. Despite its role in tumor process is well characterized, ETV6 germline mutations and their involvement in megakaryopoiesis are still poorly described. Therefore, the main aim of this doctoral research was to give insights into the molecular characterization of novel ETV6 variants together with their patogenicity mechanism. Thanks to the consolidated collaboration with IRCCS S. Matteo (Pavia), all the three years have been constantly characterized by the screening on the ETV6 gene in probands belonging to a large cohort of probands affected by Inherited Thrombocytopenia (IT) but lacking of a molecular diagnosis. The screening highlighted so far a total of 7 new ETV6 missense variants in 11 families. After the in silico prediction, we set up and performed a functional study workflow (western blot, reporter assay and immunofluorescence) in order to evaluate the pathogenic role of each variant. Luciferase (reporter) assay showed the loss of repressive function in 5 out of 7 ETV6 variants and the immunofluorescence clarified that the pathogenic mechanism consists in an abnormal retention of mutated proteins in the cytoplasm and not to a defect in DNA binding, as expected by the position in which the mutations occurs. Moreover, we hypothesized that ETV6, like RUNX1, could control the expression level of ANKRD26 during megakaryopoiesis, severily impairing platelet production and promoting the neoplastic evolution under defective conditions.