ASXL1 Mutations Predict for Resistance to Chemotherapy and Inferior Outcome in Younger Adult Patients with Acute Myeloid Leukemia (AML): A Study of the German-Austrian AMLSG
Authors: Paschka Peter , Schlenk Richard F. , Herzig Julia , Aulitzky Teresa , Gaidzik Verena I. , Bullinger Lars , Habdank Marianne , Corbacioglu Andrea , Späth Daniela , Köhne Claus-Henning , Kündgen Andrea , von Lilienfeld-Toal Marie , Held Gerhard , Horst Heinz A. , Götze Katharina , Bentz Martin , Krauter Jürgen , Ganser Arnold , Döhner Hartmut , Döhner Konstanze 
Published: October 2019

Abstract
Abstract 412

Background:
Somatic mutations of the ASXL1 (Additional Sex Comb-Like 1) gene on chromosome 20q11.1 were identified in various myeloid malignancies with the highest incidence reported in CMML (∼40%) and lower frequencies in MDS, AML, CML, and myeloproliferative neoplasia. The ASXL1 protein has been suggested to act as chromatin modifier and the highly conserved C-terminal plant homeo-domain (PHD) finger is presumably critical for its function. ASXL1 mutations cluster in exon 12 and are mainly frameshift mutations predicted to remove the PHD domain. We and others (Paschka et al., Haematologica 2011;96(s2);425; Chou et al., Blood 2010;116:4086–94) have recently reported first results on the unfavorable prognostic impact of ASXL1 mutations in AML. However, the clinical relevance of these mutations still needs to be elucidated in larger AML cohorts.

Methods:
Mutational analyses of ASXL1 were performed on diagnostic samples from 1429 patients with AML aged 18 to 61 years. All patients were intensively treated on one of two AMLSG trials [AML HD98A (n=745), Schlenk et al., J Clin Oncol. 2010;28:4642–8; AMLSG 07–04(n=684), NCT00151242]. GeneScan-based fragment analysis of several amplicons spanning ASXL1 exon 12 was used to screen for ASXL1 mutations. Samples showing altered GeneScan profiles were amplified in a second PCR reaction, and the amplicons were sequenced to confirm the mutation and to determine the mutation type. Patients were also assessed for the presence of NPM1, FLT3 (ITD and TKD), CEBPA, IDH1/2, RUNX1, and DNMT3A mutations by standard PCR-based methods.

Results:
ASXL1 mutations were detected in 90 (5.9%) of 1429 patients. All mutations were heterozygous frameshifts predicted to cause loss of the PHD finger. The most common mutation was a duplication of guanine at position 1934 (c.1934dup) identified in 59% (53/90) of the mutated cases. Three other ASXL1 mutations were detected in more than one patient: c.1900_1922del (n=16), c.1934del (n=5), c.1960dup (n=3). The majority of mutations (91%) clustered within or around a glycine-rich protein domain spanning amino acids 642–685. Patients with ASXL1 mutations were older (P<.0001), more frequently males (P=.03), and they more frequently had secondary AML (P=.02). They also showed lower values for WBC (P=.01), bone marrow (P=.0002) and circulating (P=.001) blasts, and LDH serum levels (P=.007). ASXL1 mutations were more frequent in patients exhibiting intermediate-risk cytogenetics (P=.06). Among the ASXL1 mutated cases, 42% were cytogenetically normal (CN), 23% had other intermediate-risk cytogenetics, 26% had high-risk, and 9% low-risk [t(8;21) n=7, t(15;17) n=1, no inv(16)/t(16;16)] cytogenetics. ASXL1 mutations were associated with RUNX1 (P<.001) and IDH2R140 mutations (P=.006). In contrast, ASXL1 mutations were less frequent in patients with NPM1 (P<.001), FLT3-ITD (P<.001), and DNMT3A (P=.02) mutations. The median follow-up for survival was 4.8 years. Patients with an ASXL1 mutation had an inferior complete remission (CR) rate compared with ASXL1 wildtype patients (57% vs 74%; P<.001); the same was true for the subset of CN-AML (58% vs 77%; P=.04). In both, the entire cohort (P=.04) and in CN-AML (P=.07) ASXL1 mutations were associated with a worse event-free-survival, whereas no impact of these mutations was observed on relapse-free-survival. Patients with ASXL1 mutation had a shorter overall survival (OS) compared to those with ASXL1 wildtype (P=.002; 4-year OS rates, 32% vs 47%). The adverse impact of ASXL1 mutations on OS was also present in the subgroup of CN-AML (P=.009; 4-year OS rates, 33% vs 48%). In multivariable analysis, ASXL1 mutation was in trend an unfavorable factor for OS in CN-AML (hazard ratio: 1.46; P=.087).

Conclusions:
ASXL1 mutations were identified in ∼6% of younger AML patients and were associated with intermediate-risk cytogenetics. ASXL1 mutations frequently occurred with RUNX1 and IDH2R140 mutations, and were less frequent in NPM1 and FLT3-ITD mutated AML. ASXL1 mutations were associated with lower CR rate and inferior OS. The exact role of ASXL1 in normal hematopoiesis still needs to be defined in functional studies as well as the potential role of mutated ASXL1 protein in mediating resistance to chemotherapy.

Disclosures:
Kündgen: Celgene: Honoraria; Novartis: Honoraria.