Polycythaemia vera (PV), essential thrombocythaemia (ET), and primary myelofibrosis (PMF) are the three classical Philadelphia chromosome-negative myeloproliferative neoplasms (MPNs). More than 95% of PV patients and ~50% of ET and PMF patients have mutations in the JAK2 gene. Around 5% of patients with ET or PMF carry mutations in the thrombopoietin receptor gene MPL. In December 2013, two research groups reported mutations in the calreticulin gene (CALR) in ~25% of ET and 35% of PMF patients.1,2 Mutations in JAK2, MPL, or CALR were found to be mutually exclusive. ET and PMF patients who did not have mutations in any of the three genes were referred to as ‘triple-negative’.1 The two groups identified 361 and 192 different types of CALR mutations, two of which were common: a 52 base pair deletion (Type 1; c.1099_1150del) and a 5 base pair insertion (Type 2; c.1154_1155insTTGTC). These accounted for 53%1/45%2 and 32%1/41%2 of all the CALR mutations, respectively. In recent reports, non-Type 1/2 CALR mutations have been categorised together with Type 1 or 2 into ‘Type 1-like’ and ‘Type 2-like’ mutations.3,4 All mutations in CALR led to a shift of the reading frame and the expression of a novel, common C-terminal peptide in the CALR protein.1,2 CALR mutations were described as early events in the clonal development of the disease1,2 and were detected in a range of haematopoietic stem and progenitor cell (HSPC) fractions from patients.2
In ET, CALR mutations were associated with significantly lower age, lower haemoglobin levels, lower white blood cell counts, and higher platelet levels compared with JAK2 mutations.1,2,5-7 When stratifying ET patients according to genotype (mutant JAK2, MPL or CALR, or triple-negative), there was neither a significant difference in overall survival nor in the incidence of disease progression to myelofibrosis or acute myeloid leukaemia based on statistical models correcting for age.5-7 CALR mutant ET patients had a significantly lower incidence of thrombosis compared to patients with the JAK2 mutation.6 A stratification of ET patients according to Type 1-like and Type 2-like CALR mutations showed in one study that particularly Type 2-like mutations were associated with a lower incidence of thrombosis,3 while no differences were observed in another study.8 Patients with CALR Type 1-like mutations had a significantly higher incidence of myelofibrotic transformation compared to both, patients with JAK2 or CALR Type 2-like mutations.3
In PMF, patients with CALR mutations were significantly younger, had a higher platelet count,1,9,10 and a lower leukocyte count1,9 than those with JAK2 mutations. They were less likely to develop anaemia, thrombocytopenia, or leukocytosis.9 In contrast to ET, CALR mutations patients had a significantly better age-corrected overall survival than patients with JAK2 mutations or triple-negative patients.9,10 This difference is predominantly attributable to Type 1-like rather than Type 2-like mutations.4 In multivariable models correcting for age and variables of the International Prostate Symptom Score (IPSS), or Dynamic International Prognostic Scoring System for myelofibrosis (IPSS), or the Dynamic IPSS plus the mutational status had an independent prognostic effect.9,10 CALR mutant patients had a significantly lower age-adjusted cumulative incidence of thrombosis than JAK2 mutant patients with PMF.9
Biochemical studies in cell lines revealed that mutant CALR binds to the thrombopoietin receptor MPL and activates downstream signalling.11-13 In a retroviral mouse model, mice transplanted with mutant CALR-expressing HSPCs developed an MPN-like disease phenotype showing elevated platelet counts and progression to myelofibrosis.14 While in the stem cell compartment only CALR Type 1 mutant cells had a competitive advantage over wildtype cells, both Type 1 and Type 2 CALR mutations were associated with elevated numbers of megakaryocytes and platelets.14 Expression of MPL was required for the development of the disease phenotype in mice.14
In conclusion, mutations in CALR are novel markers in MPN that are strongly associated with thrombocytosis in patients. Activated MPL signalling is a central requirement for the disease pathogenesis in a mutant CALR context. Genotyping for CALR mutations aids diagnosis, and may have a role in prognosis and treatment decisions, although so far only retrospective studies have been published.