Targeted cancer exome sequencing reveals recurrent mutations in myeloproliferative neoplasms

Abstract

With the intent of dissecting the molecular complexity of Philadelphia-negative myeloproliferative neoplasms (MPN), we designed a target enrichment panel to explore, using next-generation sequencing (NGS), the mutational status of an extensive list of 2000 cancer-associated genes and microRNAs. The genomic DNA of granulocytes and in vitro-expanded CD3+T-lymphocytes, as a germline control, was target-enriched and sequenced in a learning cohort of 20 MPN patients using Roche 454 technology. We identified 141 genuine somatic mutations, most of which were not previously described. To test the frequency of the identified variants, a larger validation cohort of 189 MPN patients was additionally screened for these mutations using Ion Torrent AmpliSeq NGS. Excluding the genes already described in MPN, for 8 genes (SCRIB, MIR662, BARD1, TCF12, FAT4, DAP3, POLG and NRAS), we demonstrated a mutation frequency between 3 and 8%. We also found that mutations at codon 12 of NRAS (NRASG12V and NRASG12D) were significantly associated, for primary myelofibrosis (PMF), with highest dynamic international prognostic scoring system (DIPSS)-plus score categories. This association was then confirmed in 66 additional PMF patients composing a final dataset of 168 PMF showing a NRAS mutation frequency of 4.7%, which was associated with a worse outcome, as defined by the DIPSS plus score.

Targeted next-generation sequencing identifies a subset of idiopathic hypereosinophilic syndrome with features similar to chronic eosinophilic leukemia, not otherwise specified

“…In myeloproliferative neoplasms, discoveries include CALR mutations in JAK2 V617-negative classic myeloproliferative neoplasms, 5,6 as well as a number of recurrent mutations that correlate with the clinical features, prognosis, and treatment responses. [7][8][9] Mutational analysis in general can help to differentiate a clonal hematopoietic neoplasm from a reactive process in diagnostically challenging cases. However, this approach has been complicated by reports of frequent somatic mutations in healthy populations of older individuals.…”

Clonal evolution and clinical correlates of somatic mutations in myeloproliferative neoplasms

“…14, 15 Mutations in TP53, TET2, SH2B3, and IDH1 are more frequently observed in leukemic blasts from transformed MPN patients, suggesting a role for these gene mutations in leukemic transformation. [16][17][18][19] However, so far only mutations in ASXL1 and NRAS have been shown to be of prognostic value in patients with PMF.…”

“…[16][17][18][19] However, so far only mutations in ASXL1 and NRAS have been shown to be of prognostic value in patients with PMF. 15,20 Using targeted NGS to search for mutations in 104 cancer-related genes, we have defined the mutational profile of a cohort of 197 MPN patients and dissected the temporal order of acquisition and clonal architecture of mutational events. We further analyzed the impact of the somatic mutations on clinical outcome.…”

Correlation of mutation profile and response in patients with myelofibrosis treated with ruxolitinib

“…In recent years, with the explosion of whole genome sequencing efforts, a number of rare, but recurrent, mutations have been identified in myeloproliferative neoplasms (MPNs), suggesting that there may be several paths to the development of MPNs, which have heterogeneous clinical presentations and treatment responses. [3][4][5] To detect genes that may be correlated with response to ruxolitinib and other clinicopathologic features, we performed a comprehensive mutation profile of 29 genes recurrently mutated in primarily myeloid malignancies 6 in a cohort of 95 patients with MF who were treated with ruxolitinib in a phase 1/2 study. 7 …”