Clinical and Biological Characteristics According to the Burden of JAK2V617F Mutated Allele in BCR-ABL Negative MPNs
Authors: Zamora Lurdes , Cabezon Marta , Garcia Olga , Alonso Esther , Marce Silvia , Fernandez Cristalina , Xicoy Blanca , Vela Dolors , Miguel Luz San , Portal Imma , Xandri Marisol , Buch Joan , Rodriguez Lluis , Dominguez Diana , Lopez Natividad , Granada Isabel , Plensa Esther , Ruiz-Xivillé Neus , Navarro Jose-Tomas , Boqué Concha , Gallardo David , Ribera Josep-Maria , Feliu Evarist , Millá Fuensanta 
Published: October 2019

Abstract
Abstract 1437
JAK2 mutation testing and karyotye are routinely used for diagnosis of myeloproliferative neoplasms (MPNs) but they have not been incorporated into risk stratification. Although JAK2V617F mutation in MPNs has been one of the most seminal medical discoveries in recent years, it is not clear the importance of the amount of JAK2V617F allele. Some studies correlate the JAK2 allele burden with a higher hemoglobin level, leukocyte count, splenomegaly and thrombosis and more probability of transformation to MF or AML.
The aim of this study was to determine whether cytogenetic data, JAK2 mutation status and allele burden correlates with cytological subtypes, clinical complications or provide prognostic information.

Methods
A retrospective study was conducted with samples centralized in a unique laboratory since 2005. A total of 526 patients were included (median age 63; 243 males) with classic MPNs (348 ET, 135 PV, 43 PMF) fulfilling 2008 WHO criteria and in accordance with the Declaration of Helsinki.
Conventional cytogenetic was performed in bone marrow samples obtained at diagnosis (n=205) and at progression to MF or AML (n=46).
DNA was extracted from peripheral blood using QIAamp DNA mini kit (Qiagen). All samples were coded and assayed blindly in duplicate to detect JAK2V617F mutation with an allele-specific PCR using TaqMan allelic discrimination, with 2 specific probes to measure the respective fluorescence of each allele. Then, JAK2 MutaQuant assay (Ipsogen, Luminy Biotech) was used to detect the JAK2V617F quantity by real-time PCR, detecting fluorescent signals using double-dye hyrolysis oligonucleotide probes with calibration standards at 4 different concentrations. Homozygous (HOZ) ratio was considered when percentage was higher than 50.
Laboratory (hemoglobin, WBC and platelet counts) and clinical data (constitutional symptoms, splenomegaly, complications, OS and DFS) were collected. For continuous variables parametric and non parametric statistics were used. Survival analysis was performed using Kaplan-Meier estimate and log-rank tests were used for comparisons. The χ2 and Fisher's exact tests were used to analyze differences in the distribution of variables among patient subsets. p-value less than 0.05 were considered statistically significant.

Results
Aberrant karyotypes were seen in 15/205 (7%) cases at diagnosis (4% in ET and PV and 40% in PMF). At progression to MF or AML we have cytogenetic studies in 22 patients, and 10 (45%) harbor alterations.
A total of 283 patients (64%) were JAK2V617F, 61% ET (4% HOZ), 75% PV (28% HOZ) and 55% PMF (16% HOZ). The median value of JAK2V617F was 26% (range, 1–99.9%). No correlation was seen between JAK2 and karyotype at diagnosis, but 7/9 patients with aberrant karyotype at progression had JAK2V617F mutation.
JAK2 correlations with laboratory and clinical data are summarized in Table 1 and 2.

Conclusions
JAK2V617F is associated with a more pronounced myeloproliferative phenotype (higher hemoglobin level, platelet or leukocyte count).
Patients with JAK2V617F HOZ have a higher probability of splenomegaly. Hematological complications do not depend on JAK2 ratio but mutated patients have more probability of thrombosis or hemorrhage.
OS and PFS do not depend on JAK2 status, but patients with JAK2V617F heterozygous seem to have a slightly better outcome.

Disclosures:
No relevant conflicts of interest to declare.