Low level CpG island promoter methylation predicts a poor outcome in adult T-cell acute lymphoblastic leukemia

Abstract

Cancer cells undergo massive alterations in their DNA methylation patterns which result in aberrant gene expression and malignant phenotypes. Abnormal DNA methylation is a prognostic marker in several malignancies, but its potential prognostic significance in adult T-cell acute lymphoblastic leukemia (T-ALL) is poorly defined. Here, we performed methylated DNA immunoprecipitation to obtain a comprehensive genome-wide analysis of promoter methylation in adult T-ALL (n=24) compared to normal thymi (n=3). We identified a CpG hypermethylator phenotype that distinguishes two T-ALL subgroups and further validated it in an independent series of 17 T-lymphoblastic lymphoma. Next, we identified a methylation classifier based on nine promoters which accurately predict the methylation phenotype. This classifier was applied to an independent series of 168 primary adult T-ALL treated accordingly to the GRAALL03/05 trial using methylation-specific multiplex ligation-dependent probe amplification. Importantly hypomethylation correlated with specific oncogenic subtypes of T-ALL and identified patients associated with a poor clinical outcome. This methylation-specific multiplex ligation-dependent probe amplification based methylation profiling could be useful for therapeutic stratification of adult T-ALL in routine practice. The GRAALL-2003 and -2005 studies were registered at http://www.clinicaltrials.gov as #NCT00222027 and #NCT00327678, respectively.

Copyright© 2020 Ferrata Storti Foundation.

Figures

Figure 1.

Genome-wide promoter methylation-array hierarchical clustering in T-cell acute lymphoblastic leukemias. (A) Unsupervised hierarchical clustering of 24 adult T-cell acute lymphoblastic leukemias (T-ALL) based on the genome-wide promoter methylation (MeDIP-array). The hypermethylated (hyperM; group 1) and intermediate methylated (interM; group 2) clusters are indicated. (B) Supervised clustering of T-ALL samples along with three human thymi using the differentially methylated signature obtained between groups 1 and 2 (panel A).

Figure 2.

Genome-wide promoter methylation-array hierarchical clustering in T-lymphoblastic lymphomas. (A) Unsupervised hierarchical clustering of 17 T-lymphoblastic lymphomas (T-LBL) based on genome-wide promoter methylation (MeDIP-array). The hypermethylated (hyperM; group 1) and intermediate methylated (interM; group 2) methylated clusters are indicated. (B) Supervised clustering of T-LBL samples, one thymoma and three thymi, using the differentially methylated signature obtained between groups 1 and 2 (panel A). (C) Venn diagram representing the overlap between the differentially methylated promoters between hyperM and interM subgroups found in T-ALL and T-LBL samples. Statistical significance was assessed by a Hypergeometric test.

Figure 3.

Targeted promoter methylation analysis in GRAALL 03/05 T-cell acute lymphoblastic leukemias series. (A) List of the nine gene promoters classifier allowing methylation status prediction. (B) Representative ratio charts of methylation specific-multiplex ligation-dependent probe amplification (MS-MLPA) analysis for one normal thymus and two T-cell acute lymphoblastic leukemias (T-ALL) from the training series belonging to the intermediate methylated (interM) subgroup and the hypermethylated (hyperM) subgroup respectively. Top panels refer to the MLPA (undigested) reference panel and the bottom panel the MS-MLPA (digested with HhAI restriction enzyme) panel. (C) Methylation ratio was assessed by MS-MLPA for T-ALL from the training series and according to their methylation subgroup and for three normal thymi. (D) Methylation ratio assessed by MS-MLPA for 168 adult T-ALL included in GRAALL03/05 trial and according to the driver oncogene involved (TLX1, TLX3, HOXA, SIL-TAL1). (E) Methylation ratio according to the early thymic precursor (ETP) phenotype.

Figure 4.

Outcome of patients according to the methylation ratio. (A) and (B) Kaplan-Meyer graphs according to the methylation status, hypomethylated (hypoM) cases (Q1) versus the others (Q2-Q4) for cumulative incidence of relapse (CIR) and overall survival (OS), respectively, for patients included in the GRAALL03-05 trial.