Concomitant and noncanonical JAK2 and MPL mutations in JAK2V617F- and MPLW515 L-positive myelofibrosis.

Sequential genotyping for phenotype-driver mutations in JAK2 (exon 14), CALR (exon 9), and MPL (exon 10) is recommended in patients with myeloproliferative neoplasms. Yet, atypical JAK2- and MPL-mutations were described in some triple-negative patients. Whether noncanonical and/or concomitant JAK2- and MPL-mutations exist in myelofibrosis (MF) regardless of phenotype-driver mutations is not yet elucidated. For this, next-generation sequencing (NGS) was performed using blood genomic DNA from 128 MF patients (primary MF, n = 93; post-ET-MF, n = 18; post-PV-MF, n = 17). While no atypical JAK2- or MPL-mutations were seen in 24 CALR-positive samples, two JAK2-mutations [c.3323A > G, p.N1108S; c.3188G > A, p.R1063H] were detected in two of the 21 (9.5%) triple-negative patients. Twelve of the 82 (14.6%) JAK2V617F-positive cases had coexisting germline JAK2-mutations [JAK2R1063H, n = 6; JAK2R893T, n = 1; JAK2T525A, n = 1] or at least one somatic MPL-mutation [MPLY591D, n = 3; MPLW515 L, n = 2; MPLE335K, n = 1]. Overall, MPL-mutations always coexisted with JAK2V617F and/or other MPL-mutations. None of the JAK2V617F plus a second JAK2-mutation carried a TET2-mutation but all patients with JAK2V617F plus an MPL-mutation harbored a somatic TET2-mutation. Four genomic clusters could be identified in the JAK2V617F-positive cohort. Cluster-I (10%) (noncanonical JAK2mutated (mut)  + TET2wildtype (wt) ) were younger and had less proliferative disease compared with cluster-IV (5%) (TET2mut  + MPLmut ). In conclusion, recurrent concomitant classical and/or noncanonical JAK2- and MPL-mutations could be detected by NGS in 15.7% of JAK2V617F- and MPLW515-positive MF patients with genotype-phenotype associations. Many of the germline and/or somatic mutations might act as "Significantly Mutated Genes" contributing to the pathogenesis and phenotypic heterogeneity. A cost-effective NGS-based approach might be an important step towards patient-tailored medicine.