Single Center Experience with Screening for JAK2 V617F, cMPL W515L, cMPL W515K, and JAK2 L611V Mutations in MPN. Rarity of JAK2 L611V Does Not Justify Its Routine Screening

Abstract 5167

The myeloproliferative neoplasms (MPNs) polycythemia vera (PV), essential thrombocytosis (ET), and primary myelofibrosis (PMF) are caused by an unregulated clonal proliferation of cells derived from a pluripotent stem cell. The exon 14 JAK2 V617F somatic mutation and cMPL mutations in codon 515 of exon 10 have provided excellent diagnostic tools to distinguish MPNs from other diseases with similar clinical features. JAK2 V617F mutations are present in ∼95% of PV patients and ∼50% of ET or PMF patients. Other somatic mutations have also been found in JAK2 exon 12 in patients with PV. Recently, a new somatic mutation of JAK2 in exon 14, L611V, was reported in 3 out of 168 PV patients in cis with JAK2 V617F (Cleyrat Leukemia 2010).

Two cMPL mutations, cMPL W515L and cMPL W515K, have been identified in MPNs. The cMPL W515L mutation is reported to be present in ET and PMF patients, while cMPL W515K in some PMF patients. We designed and validated a new rapid and sensitive allele-specific quantitative PCR assay (AS-qPCR) for determination of the JAK2 L611V mutation. In our design, we included a locked nucleic acid and a mismatch in the allelic specific primers in order to enhance allelic discrimination in the PCR reaction. The assay was sensitive to mutant frequencies of >2% in the background of wild-type alleles. Allelic discrimination of the assay is 14 cycles for wild type and 11 cycles for mutant alleles.

We studied 584 patients referred with clinical diagnoses of MPN belonging to two different groups: 1) 390 patients, who, based on clinical and laboratory evaluations, had diagnoses of PV (n=74), ET (n=60), MF (n=30), other unspecified MPNs (n=189), and other non-PV patients with erythrocytosis (n=37); 2) DNA from patients referred to ARUP Laboratories with otherwise unspecified clinical diagnoses of MPN who were found to be 196 JAK2 V617F-positive but appropriate clinical and other testing was not available for classification. The first group was tested for the JAK2 V617F, JAK2 L611V, cMPL W515L and W515K mutations. Out of 390 patients, 37% (148/390) were JAK2 V617F positive (over >90% of PV patients), 9 (ET or PMF) patients carried the cMPL W515L mutation, and 1 PMF patient had 93% allelic burden for the cMPL W515K mutation. While cMPL W515L mutation was not previously reported in PV, one PV patient was positive for both the cMPL W515L and JAK2 V617F mutations. Of the 10 cMPL W515L mutations, two MF patients carried JAK2 V617F mutations concurrently. No JAK2 L611V mutation was detected in this cohort. The additional 196 JAK2 V617F-positive samples from the patients with unspecified MPNs were also screened for the JAK2 L611V mutation and none were positive.

We conclude that the rarity of the JAK2 L611V mutation does not justify its routine screening. We suggest that those patients who fulfill the WHO criteria for PV should be further evaluated by more productive laboratory studies such as screening for JAK2 exon 12 mutations, clonality studies in females and if negative assays for germline mutations such as EPOR and VHL gene mutations.