Relapse-specific mutations in cytosolic 5’-nucleotidase II in childhood acute lymphoblastic leukemia.

Abstract

9507 Background: Relapsed childhood acute lymphoblastic leukemia (ALL) carries a very poor prognosis despite intensive retreatment that often includes allogeneic stem cell transplantation, due to intrinsic drug resistance. Novel therapeutic approaches are urgently needed and identification of the biological pathways that mediate resistance might provide targets for the effective prevention and treatment of relapsed ALL. However, the spectrum of somatic mutations responsible for ALL relapse is not yet known. Methods: We profiled the transcriptome of matched diagnosis and relapse bone marrow specimens from 10 pediatric B lymphoblastic leukemia patients using massively parallel sequencing technology (RNA-seq) to identify novel mutations specific at disease recurrence. Results: Transcriptome sequencing identified 20 newly acquired novel non-synonymous mutations in 10 relapse specimens not present at initial diagnosis. Two patients harbored relapsed specific mutations in the same gene, NT5C2, which codes for the Cytosolic 5’-nucleotidase II protein at different sites in the protein sequence. Mutations were validated as relapse specific after Sanger resequencing of germline, diagnosis and relapse DNA. RNA-seq coverage of each mutation was >100X, indicating that if a relapsed clone were present at diagnosis it made up < 1% of the bulk leukemia. Full exon sequencing of NT5C2 was completed in 61 additional relapse specimens, identifying 5 additional mutations which were also confirmed as relapse specific. Structural modeling of the relapse-associated mutations in the encoded protein Cytosolic 5’-nucleotidase II suggests alteration of enzyme subunit association/dissociation. Clinically, patients who harbored NT5C2 mutations were more likely to relapse earlier following initial diagnosis than those patients without mutations (p=0.03). Conclusions: Mutations in NT5C2 are associated with the outgrowth of drug resistant cells in childhood ALL.