CALR mutant protein rescues the response of MPL p.R464G variant associated with CAMT to eltrombopag

Congenital amegakaryocytic thrombocytopenia (CAMT) is a severe inherited thrombocytopenia due to loss-of-function mutations affecting the thrombopoietin (TPO) receptor, MPL. Here, we report a new homozygous MPL variant responsible for CAMT in 1 consanguineous family. The propositus and her sister presented with severe thrombocytopenia associated with mild anemia. Next-generation sequencing revealed the presence of a homozygous MPL^R464G mutation resulting in a weak cell-surface expression of the receptor in platelets. In cell lines, we observed a defect in MPL^R464G maturation associated with its retention in the endoplasmic reticulum. The low cell-surface expression of MPL^R464G induced very limited signaling with TPO stimulation, leading to survival and reduced proliferation of cells. Overexpression of a myeloproliferative neoplasm–associated calreticulin (CALR) mutant did not rescue trafficking of MPL^R464G to the cell surface and did not induce constitutive signaling. However, it unexpectedly restored a normal response to eltrombopag (ELT), but not to TPO. This effect was only partially mimicked by the purified recombinant CALR mutant protein. Finally, the endogenous CALR mutant was able to restore the megakaryocyte differentiation of patient CD34⁺ cells carrying MPL^R464G in response to ELT.