Recurrent activating STAT5B N642H mutation in myeloid neoplasms with eosinophilia

Abstract

Determining the underlying cause of persistent eosinophilia is important for effective clinical management but remains a diagnostic challenge in many cases. We identified STAT5B N642H, an established oncogenic mutation, in 27/1715 (1.6%) cases referred for investigation of eosinophilia. Of the 27 mutated cases, a working diagnosis of hypereosinophilic syndrome (HES; n = 7) or a myeloid neoplasm with eosinophilia (n = 20) had been made prior to the detection of STAT5B N642H. Myeloid panel analysis identified a median of 2 additional mutated genes (range 0-4) with 4 cases having STAT5B N642H as a sole abnormality. STAT5B N642H was absent in cultured T cells of 4/4 positive cases. Individuals with SF3B1 mutations (9/27; 33%) or STAT5B N642H as a sole abnormality had a markedly better overall survival compared to cases with other additional mutations (median 65 months vs. 14 months; hazard ratio = 8.1; P < 0.001). The overall survival of STAT5B-mutated HES cases was only 30 months, suggesting that these cases should be reclassified as chronic eosinophilic leukemia, not otherwise specified (CEL-NOS). The finding of STAT5B N642H as a recurrent mutation in myeloid neoplasia with eosinophilia provides a new diagnostic and prognostic marker as well as a potential target for therapy.

Fig. 1

a ARMS assay for STAT5B N642H. Heterozygotes have 3 bands—control band (243 bp), wild type (wt) specific band (166 bp) and mutant allele specific band (124 bp). Using serial dilutions of DNA from a mutated case we estimated that the assay can detect STAT5B N642H at a variant allele frequency of 10%. b Summary of mutations for all 27 STAT5B N642H mutated cases as determined by ARMS PCR, Sanger sequencing and myeloid panel analysis. Group 1 are cases with no additional mutated genes, group 2 have additional mutated genes that include SF3B1 and group 3 have additional mutated genes that do not include SF3B1. ARCH = genes strongly associated with age-related clonal hematopoiesis. Full details of additional mutations are given in Supplementary Table 1

Fig. 2

Peripheral blood film (a), bone marrow smear (b) and bone marrow trephine biopsy section at ×20 (c) and ×100 (d). The blood film showed 40% neutrophils, 28% eosinophils, 4% basophils, 12% lymphocytes and 7% monocytes. The eosinophils were morphologically close to normal with only a very minor degree of vacuolation and hypogranularity but there was an increase in non-lobated forms. Bone marrow cellularity and megakaryocyte numbers were increased. The increased cellularity was due to an increase in all three granulocyte lineages (neutrophils, basophils and eosinophils). The trephine biopsy sections show hypercellularity, disorganisation and an increase in cells of neutrophil and eosinophil lineages. Reticulin was not increased

Fig. 3

Kaplan–Meier plots showing overall survival (OS) estimates for the STAT5B N642H mutated cases who had eosinophilia at diagnosis and had follow up data available (n = 23). a Comparison of the 3 molecular groups shows that cases in group 3 (those with additional mutated genes that do not include SF3B1) have an inferior OS compared to all other cases (median 14 months vs. 65 months; P < 0.0004). b OS for cases with mutations in 2 or more additional genes (excluding DNMT3A and TET2) was significantly worse than that for cases with 0 or 1 additional mutations (median 18 months vs. 50 months; P = 0.001). Of the 9 cases with mutations in ≥2 additional genes, 8 were in group 3

Fig. 4

a Clonal hierarchy for 4 patients. The numbers indicate the number of colonies with that genotype, e.g. for case E11825, 5 colonies were mutant for TET2 and SF3B1 but not STAT5B and 9 colonies were mutant for all 3 genes. Light grey indicates an inferred ancestral clone. b Sequential analysis of two cases with increasing eosinophil counts (values indicated are ×10⁹/L). Case E11825 had an elevated eosinophil count at diagnosis, with eosinophilia increasing over time co-incident with an increase in STAT5B N642H vaf. Case E13661 was diagnosed initially with PV in 1988 and STAT5B N642H was not detected in a sample taken 6 years later. Slightly elevated eosinophil counts were first noted in 2014, and a sample from this time was positive for STAT5B N642H. The mutant vaf and eosinophil counts increased over the next 4 years