EZH2 molecular alterations drive micromegakaryocyte expansion in MDS defining a specific high-risk group

Abstract BackgroundMyelodysplastic neoplasm/syndrome (MDS) subtypes are categorized by genetic aberrations leading to specific morphological dysplastic features. Micromegakaryocytes (microMK) are a defining morphological alteration of myeloid dysplasia in adults, although unrelated to any specific genetic alteration. We previously demonstrated a poor outcome of MDS patients with microMK and, more recently, we reported that most of them presented concurrent EZH2 and RUNX1 mutations (RUNX1mut+EZH2mut) [Blood (2023) 142 (Supplement 1): 4116]. RUNX1 mutations (RUNX1mut) genotype is also described in often co-occurence with monosomy 7 or deletion of the long arm of chromosome 7 (del(7q)/-7), i.e. the locus harboring EZH2, suggesting a potential cooperative role. The present study aimed to examine the correlation between RUNX1mut and EZH2mut or del(7q)/-7 with the presence of microMK, and to analyse the prognostic impact of these co-occurrences in MDS. MethodsOur cohort included MDS patients with RUNX1mut, EZH2mut and/or have del(7q)/-7 alterations from 9 international institutions. As a control group, we use MDS without these genetic alterations. MDS was diagnosed per the 5th WHO criteria. Genetic profiling included G-banding cytogenetics and NGS. Cases were categorized by the presence (≥3% of the megakaryocytes) or absence and the percentage of microMK. The Molecular International Prognostic Scoring System (IPSS-M) was employed for patients' risk stratification. Overall survival (OS) was correlated using Cox regression. The statistical analysis was performed using R version 3.5.1. ResultsClinical and molecular data from 259 MDS patients were analysed: 125 MDS with mutations of RUNX1, EZH2 or del(7q)/-7 (19 EZH2mut, 35 RUNX1mut, 38 del(7q)/-7, 21 RUNX1mut+EZH2mut and 8 RUNX1mut+del(7q)/-7) and 134 MDS controls (MDScontrol). The median follow-up was 27.8 (95%CI, 22.5 – 44.7) months. The median age was 71 years (IQR, 64-80) and 65% (n=170) of the patients were male, with not significant differences among groups. The proportion of patients with microMK was significantly lower in the MDScontrol compared to all other groups (7.8% vs 78.9% EZH2mut, 41.2% RUNX1mut, 50% del(7q)/-7, 85.7% RUNX1mut+EZH2mut, and 54.5% RUNX1mut+del(7q)/-7; p<0.0001). Those groups with EZH2 mutated gene (EZH2mut and RUNX1mut+EZH2mut) harboured similarly high frequencies of patients with microMK (p=0.86); both with significantly more microMK cases than RUNX1mut group (p=0.014 and p=0,028, respectively). The median percentage of microMK in EZH2mut alone or in combination with RUNX1 was also higher than in the RUNX1mutgroup (3%, 33% vs 0%; p=0.02 and p<0.0001, respectively). However, a significantly higher percentage of microMK was detected in RUNX1mut+EZH2mut than in EZH2mut group (p=0.037). In our cohort, TP53 mutations and del(5q) were absent in EZH2-mutated groups. ASXL1 mutations were significantly enriched in RUNX1mut+EZH2mut (80%) and EZH2mut (63.1%) cases, compared to other groups (19.5%; p < 0.001). Patients with RUNX1mut+EZH2mut, RUNX1mut+del(7q)/-7, and del(7q)/-7 had a worse OS than controls (median OS: 23.1, 13.5, 14.3 vs. 82.2 months, respectively; p<0.001). These groups remain prognostic despite including age and hematopoietic stem cell transplantation (HSCT) in the multivariate analysis (p<0.001). MDS with RUNX1mut+EZH2mut and RUNX1mut+del(7q)/-7 patients were all categorized into the moderate-high and high IPSS-M risk groups, compared to 18.6% cases in MDScontrolgroup, 44.4% in EZH2mut group and 59% in RUNX1mut group (p<0.01). Stratifying the overall cohort by IPSS-M categories, patients in the moderate-high IPSS-M group with doble alteration, RUNX1mut with EZH2mut or del(7q)/-7, showed significantly worse survival than control cohort (15.9 vs 22.4 months; p=0.039). ConclusionsOur study demonstrates the association of EZH2 gene mutation with the presence of microMK in MDS patients, and suggests that, the co-occurrence with RUNX1 mutations further increases the bone marrow microMK levels and may contribute to a poorer prognosis. Patients with combined RUNX1 and EZH2 mutations or del(7q)/-7 alterations share similarly unfavourable outcomes.