Novel DNA Methylation Markers for MRD Identified from ALL-Discordant Twins

Background: Minimal residual disease (MRD) refers to the remaining leukemic clones in post-therapeutic patients, which may lead to relapse of acute lymphoblastic leukemia (ALL). ALL-discordant co-twin design provides an efficient way to develop more clinically applicable molecular markers for MRD monitoring of ALL. Methods: Whole-genome and whole-methylome sequencing were performed on peripheral blood samples. DNA methylation gene panel was analyzed by an amplicon sequencing strategy. XGBoost and random forest model was applied to train and evaluate the candidate gene panel in 133 ALL-discordant sibling pairs and 1670 extra public methylation datasets. Findings: No evidence showed that any leukemia-associated genomic mutations were detected in the blood of B-ALL-discordant twins at complete remission (CR). 26 recurrent differential methylated regions (DMRs) were commonly identified. A 10-gene diagnosis panel were identified from 133 ALL-discordant siblings, which showed 80·8% specificity and 86·1% sensitivity. RT-PCR revealed that aberrant DNA methylation in introns of PRDM16 and LCP2 correlates with their differential transcript usage, implying their potential roles in ALL pathogenesis. Finally, we obtained a clinically applicable 4-probe panel that displayed highly promising predictive value both in blood and bone marrow samples: 98·89% sensitivity and 94·12% specificity to diagnose ALL and 100% sensitivity and 100% specificity to differentiate relapse from remission patients. Interpretation: We proved DNA methylation aberrations exist in blood samples of B-ALL patients at CR and established a 4-probe DNA methylation marker that can be potentially used in clinics for more sensitive, fast and standardized MRD monitoring in ALL. Funding Statement: Scientific Research Innovation Key Program (No. 2012213-5), Research-based Disciplines Program (No.201314-3), Research-based Talents Program (No.201314-6) of Beijing 307 Hospital Affiliated, the State Key Development Program for Basic Research of China (No. 2003CB515509 and 2009CB522401), the National High Technology Research and Developmental Program of China (2012AA02A201), and the innovative R&D team program of Guangdong Province (2009010016). Declaration of Interests: None to declare Ethics Approval Statement: Ethics permission approved by the human ethics committee of Beijing 307 Hospital.