Molecular Analysis of Myelodysplastic Syndrome with Isolated Del(5q) Reveals a Broad Spectrum of Clinically Relevant Mutations: A Study on 119 Patients and 26 Genes
Authors: Meggendorfer Manja , Alpermann Tamara , Haferlach Claudia , Schnittger Susanne , Kern Wolfgang , Haferlach Torsten 
Published: October 2019

Abstract
Introduction: The WHO category of “Myelodysplastic syndrome with isolated del(5q)” shows a good prognosis. It also demonstrates sensitivity for specific treatment such as lenalidomide. However, in some patients it evolves to secondary AML. Underlying pathobiological mechanisms are still under debate.
Aim: To determine the frequency of mutations in a 26 gene panel and to investigate a mutation pattern combined with clinical data and prognostic information.
Patients and Methods: We investigated 119 patients (85 female, 34 male) having MDS with isolated del(5q), strictly classified according to WHO classification 2008 with respect to cytomorphology and cytogenetics (blasts below 5% in the bone marrow and 5q deletion sole). All patients underwent molecular analyses by a myeloid gene panel containing ASXL1, BCOR, BRAF, CBL, DNMT3A, ETV6, EZH2, FLT3-TKD, GATA1, GATA2, IDH1, IDH2, JAK2, KIT, NRAS, KRAS, MPL, NPM1, PHF6, RUNX1, SF3B1, SRSF2, TET2, TP53, U2AF1, and WT1. The library was generated with the ThunderStorm (RainDance Technologies, Billerica, MA) and sequenced on a MiSeq instrument (Illumina, San Diego, CA).
Results: Most patients harbored 1 mutation (60/119, 50%), while 2, 3, and 4 mutations per patient occurred less frequently (18/119, 15%; 2/119, 2%; and 1/119, 1%, respectively). In 38/119 patients (32%) no gene mutation was identified in addition to the del(5q). In the total cohort the most frequently mutated genes were DNMT3A and TP53 (21/119, 18% each), followed by SF3B1 (20/119, 17%), TET2 (14/119, 12%), ASXL1 (9/119, 8%), and JAK2 (7/119, 6%). Although these 6 gene mutations overlapped rarely and occurred frequently as sole mutations, they were not completely mutually exclusive. The mutation frequencies of all other analyzed genes were below 5%. Dividing the patients in groups defined by a bone marrow blast count of <2% and 2-5%, as described in IPSS-R, we could not detect any correlation to the mutation number per patient. However, patients that had no mutation were younger compared to patients with at least 1 mutation (70 vs. 76 years, p=0.009). But there was no difference between these 2 patient groups in white blood cell count, hemoglobin level, or platelet count. Taking single genes into account revealed that TP53 and SF3B1 mutations (mut) correlated with higher age (78 vs. 73 years, p=0.047; 78 vs. 73 years, p=0.050, respectively). Addressing the correlations of ring sideroblasts (RS) >15% and SF3B1mut showed that also in MDS with isolated del(5q) these two parameters significantly correlate with a mean of 19% RS (range: 0-80%) in SF3B1mut and only 1% RS in SF3B1 wildtype patients (wt; range: 0-12%, p<0.001). Looking at prognostic relevance of gene mutations surprisingly showed that SF3B1mut patients had a significantly worse outcome than SF3B1wt patients (median overall survival (OS) 31 vs. 91 months, p=0.008). Comparing the mutation frequency of TP53 in MDS with isolated del(5q) with all other MDS (Haferlach et al, Leukemia 2014) resulted in a significant higher mutation rate in MDS with isolated del(5q) (21/119 (18%) vs. 49/781 (6%), p<0.001). However, we did not find a prognostic impact of TP53mut in our cohort. Our patients were unselected and median OS was 91 months. Thus, our cohort may include a larger proportion of patients earlier in their clinical course compared to cohorts enrolled in treatment studies. Therefore the negative impact of TP53mut may become obvious later or even not before treatment needs to be started. Of note, 50% (60/119) of our patients were only under observation or received red blood cells or erythropoietin only. In contrast, an increasing number of gene mutations per patient showed a very strong trend towards a worse outcome with a median OS of 90 months in patients with no or 1 additional mutation in comparison to patients with more than 1 mutation (median OS: 36 months, p=0.061).
Conclusion: 1) In myelodysplastic syndrome with isolated del(5q) the 5 most frequently mutated genes are comparable to all other MDS (Haferlach et al, Leukemia 2014). 2) In contrast, TP53 is more frequently mutated in MDS with isolated del(5q). 3) Ring sideroblasts >15% correlate with SF3B1mut. 4) SF3B1mut lead to significantly worse OS. 5) Increasing numbers of mutations show negative prognostic impact.

Disclosures
Meggendorfer: MLL Munich Leukemia Laboratory: Employment. Alpermann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.