Abstract
INTRODUCTION: JAK2 p.V617F mutations are common in Polycythemia Vera (>90%), Essential Thrombocythemia (50%; ET) and Primary Myelofibrosis (30-50%; PMF). Somatic mutations of codons 515 or 505 in MPL Exon 10 are present in 5% of non-JAK2-mutated ET and 1% of PMF. MPL p.W515L mutation has also been identified in patients with Refractory Anemia with Ring Sideroblasts associated with Thrombocytosis (RARS-T). MPL mutation in Exon 10 codon 505 (S505N) was originally identified as a germ line mutation associated with hereditary thrombocytopenia, but was subsequently reported in ET and PMF. Most current evidence suggests that JAK2 and MPL mutations are mutually exclusive. Pardanani et al had reported 6 cases out of 1182 patients with concurrent JAK2 and MPL mutations, using assays of different sensitivities. In the reported cases the more frequent allele mutated is MPL and less frequent allele is JAK2 as a possible second hit. Here, we report 2 cases with concurrent JAK2 and MPL mutations.
METHODS: DNA was extracted and subjected to library preparation using the 50 gene Ion AmpliSeq™ Cancer Hotspot Panel v2, clonal amplification and Ion Sphere™ Particle enrichment on the Ion One Touch™ 2 and One Touch™ ES, and Next Generation Sequencing (NGS) on the Ion Torrent PGM System™. Data was analyzed using Variant Caller Software v.4.0 and NextGENe software v.2.3.4. The original target region BED file for the Ion AmpliSeq™ Cancer Hotspot Panel v2 was modified to include only MPL exon 10 and JAK2 exon 14 amplicons. Assay coverage averaged approximately 3000X.
RESULTS: From February 2015, JAK2/MPL NGS was performed on 105 specimens to determine somatic mutations in possible MPN and MDS/MPN cases. 2/105 (1.9%) of the specimens harbored concurrent JAK2 and MPL mutations compared to 6/1182 patients reported previously (0.5%). One patient had RARS-T and one had PMF. Mean age (69 and 74 years) was 71 years, hemoglobin (8.3 and 10.3 gm/dL) 9.3 gm/dL, hematocrit (25 and 26%) 25%, Mean Corpuscular Volume (92 and 92 fL) 92 fL, platelet count (171 and 1320 x 10E3/cmm) 745 x 10E3/cmm, White blood count-WBC (6.2 and 10.3 x 10E3/UL) 8.25 x10E3/cmm, monocytes (3-5%) 4%, Basophils (1 and 2%) 2%, Neutrophils (61 and 66%) 64%, Bands (3and 15%) 9%, Eosinophils (0 and 2%) 1%, peripheral blood blasts (0 and 4%) 2%, bone marrow fibrosis grade of 2 in a grading scale of 1-3. Both were males. Spleen was not palpable at diagnosis in either patient. The case with RARS-T had 4.5% MPL p.W515L mutation allele frequency and 22.7% JAK2 p.V617F mutation allele frequency. The case with PMF had 3.3% MPL p.S505N mutation allele frequency and 2.4% JAK2 p.V617F mutation allele frequency (detected at the follow up 6 months bone marrow in May 2015). They were treated with EPO analogues/Anagrelide and Ruxolitinib respectively. None had thrombosis or bleeding complications. At a mean follow-up of 366 days (113 and 620 day), both patients are alive.
CONCLUSION: MPL (codons 505 and 515) and JAK2 p.V617F mutations are not always mutually exclusive. NGS panels with dual gene coverage and high sensitivity identify coexistent mutations that would not be identified by single gene assays. The clinical significance of concurrent mutations remains unclear.
No relevant conflicts of interest to declare.
Author notes
Asterisk with author names denotes non-ASH members.
