Apparent ‘JAK2-negative’ polycythaemia vera due to compound mutations in exon 14

A somatic activating mutation in exon 14 of JAK2 involving the pseudokinase domain (c.1849G>T; p.V617F), is found in ~95% of patients with polycythaemia vera (PV), and is a major diagnostic criterion in the 2008 World Health Organization (WHO) classification (Swerdlow et al, 2008) and its proposed forthcoming revision (Barbui et al, 2015). A further 2–3% of PV patients have mutations in exon 12 of JAK2, so that very few PV patients are ‘JAK2‐negative’, lacking a somatic mutation in that gene. As there are many alternative causes of absolute or relative erythrocytosis the diagnosis sometimes remains in doubt when no JAK2 mutation is identified. Here we report a case of WHO‐defined PV with no JAK2 mutation detected on routine testing using a single nucleotide primer extension assay for JAK2 V617F and direct sequencing of exon 12. Targeted next generation sequencing (NGS) of the entire coding region of JAK2 revealed a novel mutation in exon 14 (c.1849_1853GTCTG>TTTCT; p.V617F/C618L). This mutation resulted in failure of the JAK2 V617F mutation‐specific primer to bind to the target, causing a false negative result with amplification only of the wild type allele. Direct sequencing of exon 14 in 7 additional cases of ‘JAK2‐negative’ polycythaemia without a conclusive diagnosis, identified one other compound mutation, demonstrating the diagnostic utility of broader sequencing of JAK2 in such cases.