Additional Mutations in SRSF2, ASXL1 and/or RUNX1 Identify a High Risk Group of Patients with KIT D816V+ Advanced Systemic Mastocytosis
Authors: Jawhar Mohamad , Schwaab Juliana , Schnittger Susanne , Meggendorfer Manja , Sotlar Karl , Horny Hans-Peter , Metzgeroth Georgia , Kluger Sebastian , Naumann Nicole , Haferlach Claudia , Haferlach Torsten , Valent Peter , Hofmann Wolf-Karsten , Fabarius Alice Charlotte , Cross Nicholas C.P. , Reiter Andreas 
Published: October 2019

Abstract
The presence of multiple additional mutations was recently reported in >90% of patients with KIT D816V+ advanced systemic mastocytosis (SM) (Schwaab et al., Blood 2013). In the vast majority of cases, additional mutations occur prior to KIT D816V indicating a multi-mutated stem cell disease with phenotype modification through KIT D816V (Jawhar et al., Leukemia 2015). We now sought to clarify the prognostic impact of individual mutations and the number of mutated genes in 70 multi-mutated patients with ISM (n=26), aggressive SM (ASM, n=34) and mast cell leukemia (MCL, n=10). All patients had an associated clonal hematologic non-mast-cell disease (AHNMD), such as a myelodysplastic/myeloproliferative syndrome unclassified (MDS/MPNu, n=28), chronic myelomonocytic leukemia (CMML, n=21), chronic eosinophilia leukemia (CEL, n=9), acute myeloid leukemia (AML, n=8) or primary mylofibrosis (PMF, n=4). Mutational analysis was performed by next-generation sequencing in the following genes: ASXL1, CBL, ETV6, EZH2, IDH2, JAK2, KRAS, NRAS, RUNX1, SF3B1, SRSF2, TET2 and U2AF1. The most frequently identified mutated genes were TET2 (n=33, 47% of cases), SRSF2 (n=30, 43%), ASXL1 (n=20, 29%), RUNX1 (n=16, 23%) and JAK2 (n=11, 16%). The molecular data were correlated with overall survival (OS), clinical characteristics (e.g. B- and C-findings), bone marrow (BM) findings, serum tryptase levels and KIT D816V allele burden in peripheral blood. With a median observation time of 3.7 years, 25 of 70 patients (36%) have died. Three-year OS probability was 71%. In univariate analysis, OS was adversely influenced by mutations in SRSF2 ([hazard ratio (HR) 5.9, 95% confidence interval (CI)] [2.2-15.4], p<0.0001), ASXL1 (HR 3.4 [1.5-7.7], p=0.002) and RUNX1 (HR 2.4 [1.1-5.4], p=0.03) but was not influenced by mutations in TET2 (p=0.3) or JAK2 (p=0.1). In multivariate analysis of the most frequently mutated genes, SRSF2 remained as the best independent poor risk marker for OS. Furthermore, inferior OS was significantly associated with the number of mutated genes in SRSF2/ASXL1/RUNX1 (S/A/R), with median OS not reached for 0 mutation (n=25), 3.9 years for 1 mutation (n=27) and 2.7 years for >=2 mutations (n=18). The 3-year OS was 90% in patients with 0 mutation, 73% in patients with 1 mutation and 42% in patients with >=2 mutations in the S/A/R gene panel. Pairwise significantly different OS probabilities were observed for the comparisons 0 vs. 1 mutation (HR 9.2 [2.0-42.3], p=0.001), 0 vs. >=2 mutations (HR 4.1 [1.9-8.8], p<0.0001) and 1 and >=2 mutations in the S/A/R gene panel (HR 2.5 [1.1-6.0], p=0.04) (Figure 1). Regarding clinical characteristics, the S/A/R mutation profile is significantly related to C-findings including hepatosplenomegaly, albumin <35g/L, alkaline phosphatase >150 IU/L, ascites and weight loss, and other important disease-related parameters including bone marrow mast cell infiltration, serum tryptase levels and KIT D816V+ allele burden. We therefore conclude that a) presence and number of mutations in the S/A/R gene panel are adversely associated with advanced disease and poor survival in KIT D816V+ SM patients and b) the inclusion of molecular markers should be considered in upcoming prognostic scoring systems for SM patients.
Figure 1. Overallsurvival (OS) of 70 multi-mutated KIT D816V+ advanced systemic mastocytosis patients depending on number of mutated genes in SRSF2/ASXL1/RUNX1 (S/A/R) gene panel. HR [hazard ratio 95% confidence interval (CI)]. Figure 1. Overallsurvival (OS) of 70 multi-mutated KIT D816V+ advanced systemic mastocytosis patients depending on number of mutated genes in SRSF2/ASXL1/RUNX1 (S/A/R) gene panel. HR [hazard ratio 95% confidence interval (CI)].

Disclosures
Schnittger: MLL: Equity Ownership. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Sotlar:Ludwig-Maximilians-University: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Valent:Bristol-Myers Squibb: Honoraria; Pfizer: Honoraria; Celgene: Honoraria; Ariad: Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Cross:Qiagen: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding.