A germline exon12 mutation and a late somatic mutation in a patient with essential thrombocythemia.

Background: It has been discovered that Janus kinase 2 () exon12 mutations lead to the polycythemia vera (PV) phenotype, while somatic mutations of calreticulin () are associated with essential thrombocythemia (ET) or primary myelofibrosis. In this article, we report a case of ET with coexistence of exon12 and mutations. The objective of this study was to elucidate the pathogenicity mechanism of a exon12 mutation (N533S) and the role of the coexistence of mutations on the hematological phenotype.

Methods: We designed a colony analysis of tumor cells obtained from this patient, and attempted to identify mutant genes using DNA from hair follicles. Mutation impairment prediction and conservative analysis were conducted to predict the mutation impairment and structure of N533S. In addition, we conducted a functional analysis of N533S by constructing Ba/F3 cell models.

Results: Three distinct tumor subclones, namely N533S/type1 , N533S/ , and N533S/type1 , were identified from the 17 selected erythroid and 21 selected granulocyte colonies. The analysis of hair follicles yielded positive results for N533S. According to the bioinformatics analysis, N533S may exert only a minor effect on protein function. Functional studies showed that N533S did not have a significant effect on the proliferation of Ba/F3 cells in the absence of interleukin-3 (IL-3), similar to wild-type . Notably, there were no increased phosphorylation levels of -downstream signaling proteins, including signal transducer and activator of transcription 3 (STAT3) and STAT5, in Ba/F3 cells harboring the N533S.

Conclusion: Our study revealed that the N533S/type1 subclone was linked to a significant expansion advantage in this patient, indicating that it may contribute to the development of the ET phenotype. We further demonstrated that N533S, as a noncanonical exon12 mutation, is a germline mutation that may not exert an effect on cell proliferation and protein function. These results and the present body of available data imply that certain noncanonical mutations are not gain-of-function mutations leading to the development of myeloproliferative neoplasms.