P216 | COEXISTENCE OF LOW BURDEN ALLELE <i>JAK2</i>V617F MUTATION AND <i>CALR</i> OR <i>/MPL<i> MUTATIONS IN A LARGE COHORT OF PATIENTS WITH CHRONIC MYELOPROLIFERATIVE NEOPLASMS

Chronic myeloproliferative neoplasms (MPNs) are clonal hematological disorders originating from altered myeloid cell proliferation in the bone marrow (BM) and peripheral blood (PB). These include polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). PV is characterized by an increase in red blood cells and mutations in the JAK2 gene; ET is characterized by an increase in platelets, with JAK2 mutations in 60%, CALR in 15-20% and MPL in 3-5%; PMF is characterized by medullary fibrosis, with JAK2 mutations in 60%, CALR in 20-25% and MPL in 5-8%. The diagnostic workflow for PV, ET and PMF includes testing for the V617F mutation of JAK2. If positive, further testing for CALR or MPL mutations is not necessary. In PV, negative JAK2 V617F results require testing for exon 12 mutations in JAK2, while for suspected ET or PMF testing for CALR and MPL mutations is required according to WHO. Although JAK2, CALR and MPL mutations are generally mutually exclusive, rare cases of co-existing JAK2V617F with CALR or MPL mutations have been reported. This study aimed to determine the percentage of patients affected by Ph negative MPNs with co-existing JAK2, CALR and MPL mutations. From 2020 to 2024, DNA from patients with suspected MPNs was extracted from BM or PB and analyzed by Next Generation Sequencing using the "Myeloid Solution" panel from Sophia Genetics. JAK2 V617F positive patients (VAF <5%) were confirmed by qPCR using the Ipsogen JAK2 MutaQuant kit, MPL mutations were confirmed by qPCR using the Ipsogen MPL MutaScreen kit and CALR mutations by the Ipsogen CALR RGQ PCR kit. Among 797 JAK2 V617F positive patients (VAF <5%), 1.13% (9 patients) had an additional MPL mutation and 0.63% (5 patients) had an additional CALR mutation. Within patients with mutated MPL, 6 had the c.1544G>T, 1 had the c.1543T>A and 2 had the c.1537_1539del. According to the American College of Medical Genetics (ACMG) the first two are classified as pathogenic, while the latter is classified as a variant of uncertain significance. In patients with CALR mutations, 3 had CALR Type 1 (c.1099_1150del), 1 had CALR Type 2 (c.1154delinsTTTGTTC) both classified as pathogenic and 1 had c.1121del, classified as likely pathogenic according to ACMG. MPNs are heterogeneous disorders characterized by mutations in JAK2, CALR, and MPL. Identification of low burden JAK2 V617F mutation does not exclude the presence of CALR or MPL mutations, which could affect clinical management.