NGS analysis of the mutational profile of patients with Ph-negative myeloproliferative neoplasms

Introduction . The identification of driver mutations in the JAK2, CALR, and MPL genes is a gold standard approach in the molecular diagnosis of patients with Ph-negative myeloproliferative neoplasms (Ph-MPNs). However, such patients are characterized by a heterogenous genomic landscape. Standard molecular genetic methods cannot be used to identify most somatic mutations, thus failing to provide a comprehensive understanding of the course and prognosis of Ph-MPNs and to confirm the clonality of the disease in patients with triple-negative status. The next generation sequencing (NGS) technology allows simultaneous analysis of an extensive panel of genes and identification of both pathogenic and driver mutations. Aim. To evaluate the possibility of using NGS to study the mutational status of patients with Ph-negative MPNs and to analyze the effect of identified pathogenic mutations on patient survival. Materials and methods . The study included 83 patients with polycythemia vera, essential thrombocythemia, and primary myelofibrosis aged from 19 to 85 years (the median onset age of 51 years). For all patients, sequencing was performed using a myeloid panel of 118 genes with an average reading depth of 1000x on MiSeq (Illumina, USA). The clinical significance of the mutations was determined using the COSMIC and Franklin databases. The survival rate was analyzed using the Kaplan–Meyer method followed by assessment of statistical significance using the Cox-Mantel test in the GraphPad Prism 8 environment. Results. Pathogenic mutations in 23 genes were detected in 39 (46%) patients out of the total cohort of patients. The most frequent mutations were detected in the ASXL1 gene in 25% of patients, which reduced event-free survival by 50.3% (Me = 7.83 years vs 15.75 years). The pathogenic mutations identified in other genes combined with mutations in driver genes also decreased event-free survival compared to patients with isolated driver mutations. Two or more pathogenic mutations significantly reduced event-free survival compared to patients with only one pathogenic mutation. The NGS method was also capable of identifying pathogenic mutations in 8 out of 10 triple-negative patients studied, thus confirming the clonality of the disease. Conclusions. The next-generation sequencing (NGS) method using a panel of 118 genes is an effective tool in identifying predictively significant mutations important for selecting the most effective personalized therapy to achieve hematologic response.