Comprehensive Molecular Profiling of T-Cell Acute Lymphoblastic Leukemia Identifies Mutations in 76 (97.4%) of 78 Patients

Abstract

Abstract 2548 Introduction:At present, the diagnosis of T-ALL is based on immunophenotyping and specific chromosomal rearrangements. However, the knowledge about recurrent somatic mutations is limited. Patients and Methods: We studied a cohort of 78 adult T-ALL cases (n=33 early, n=33 cortical, n=2 mature T-ALL, n=10 subtype not available), including 57 males and 21 females. Age ranged from 18.8–87.7 yrs (median: 42 yrs). A deep-sequencing assay was used to investigate for specific molecular alterations in genes involved in transcriptional regulation: NOTCH1, FBXW7, CDKN2A, DNMT3A, FLT3-ITD, FLT3-TKD, NPM1, PTEN, and RUNX1. Further, chromosome banding analysis and FISH with probes for DNMT3A (2p23), SEC63 (6q21), MYB (6q23), CDKN2A (9p21), PTEN (10q23), CDKN1B (12p13) and TP53 (17p13), as well as CDKN2B promoter methylation analyses were performed. Results: Cytogenetic data was available in 68 patients: normal karyotype: n=22 (2 of these harbored a PICALM-MLLT10-rearrangement), SIL-TAL1-rearrangement: n=3, t(5;14)(q35;q32): n=2, t(10;14)(q24;q11)/t(7;10)(q34;q24): n=9, t(10;11)(p13;q21): n=3, other abnormalities n=29. Importantly, molecular mutations were detected in 67/78 patients (85.6%). In detail, NOTCH1 was the most frequently mutated gene (55/77 cases, 71.4%). Other alterations were detected in DNMT3A (16/78; 20.5%); RUNX1 (13/78; 16.6%); FBXW7 (11/75; 14.6%); PTEN (7/78; 10.0%); CDKN2A (3/58; 5.2%); FLT3-ITD (2/78; 2.5%); and FLT3-TKD (1/70; 1.4%). By FISH analyses, heterozygous deletions of the following loci were observed: DNMT3A (1/43; 2.3%), SEC63 (7/43; 16.3%), PTEN (1/32, 3.1%), CDKN1B (8/43; 18.6%) and TP53 (3/43; 7.0%). CDKN2A deletions were detected in 30/72 (41.6%) cases: n=14 heterozygous, n=15 homozygous, n=1 showed a clone with a heterozygous and a subclone with a homozygous deletion. Further, the CDKN2B promoter methylation status was analyzed. Here, 36/74 (48.6%) cases demonstrated hypermethylation. As such, when combining molecular mutations, CDKN2A deletions, and CDKN2B hypermethylation, in median 2 alterations per case were observed (range 1–5). Moreover, almost every patient (76/78) harbored at least one aberration resulting in a mutation rate of 97.4%. Interestingly, considering alterations in the group of cyclin-dependent kinase inhibitors (CDKN2A/1B deletions, CDKN2A mutations, and CDKN2B hypermethylation), 61/78 (78.2%) cases carried at least one such alteration. With respect to associations amongst molecular mutations, no specific pattern was observed except for a strong correlation between RUNX1 and DNMT3A mutations, i.e. 6/13 RUNX1 mutated cases concomitantly harbored DNMT3A mutations (p=0.021). Furthermore, we observed that DNMT3A and RUNX1 alterations were associated with higher age (DNMT3A: mean±SD 60.9±16 vs. 39.6±16 years; p<0.001; RUNX1: mean±SD 55.4±18 vs. 41.7±18 yrs; p=0.013) whereas PTENmut were associated with younger age (mean±SD 32.9±10 vs. 45.0±19 yrs; p=0.019). With regard to cytogenetics, DNMT3A was significantly correlated with normal karyotype (9/23, 39.1% vs. 6/45, 13.3%; p=0.028). Moreover, RUNX1mut were associated with lower WBC count (mean±SD 26.4±41 vs. 63.4±90 cell count G/L; p=0.025). With respect to immunophenotypes, cases with RUNX1mut showed a trend to be associated with early T-ALLs (9/23, 39.1% vs. 6/45, 13.3%; p=0.082). CDKN2B hypermethylation was significantly correlated with early T-ALLs (21/32, 65.6% vs. 10/31, 32.2%; p=0.012). In contrast, FBXW7mut were associated with the cortical subgroup (1/32, 3.1% vs. 9/32, 28.1%; p=0.013). With regard to clinical outcome, patients with RUNX1mut had a shorter overall survival (OS) compared to RUNX1wt patients (alive at 2 yrs: 44.4% mutated vs. 64.0% wild-type, p=0.011). Further, for NOTCH1mut cases (alive at 2 yrs: 67.4% mutated vs. 33.6% wild-type, p=0.060) a trend towards a better OS was detectable. Conclusions: 1. T-ALL is characterized by a high number of genetic alterations since 46/68 (67.6%) showed cytogenetic aberrations. In addition, at least one molecular alteration was observed in 76/78 (97.4%) patients. 2. The most frequent alterations observed were mutations in NOTCH1, DNMT3A, RUNX1 and FBXW7. 3. The cyclin-dependent kinase inhibitors were altered by deletion, mutation or hypermethylation in 78.2% of cases. 4. RUNX1 mutations are associated with shorter and NOTCH1 mutations with longer OS. Disclosures:Grossmann:MLL Munich Leukemia Laboratory: Employment. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Weissmann:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Artusi:MLL Munich Leukemia Laboratory: Employment. Schindela:MLL Munich Leukemia Laboratory: Employment. Stadler:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.