Mutations in the RAS Signaling, B-Cell Development, TP53/RB1, and JAK Signaling Pathways Are Common in High Risk B-Precursor Childhood Acute Lymphoblastic Leukemia (ALL): A Report From the Children's Oncology Group (COG) High-Risk (HR) ALL TARGET Project.

Abstract 85

Despite the overall favorable treatment outcome for childhood ALL, about 20% of patients still experience relapse. Moreover, many ALL cases lack sentinel chromosomal alterations, and the genetic factors contributing to leukemogenesis in these cases are poorly understood. Genome-wide profiling of DNA copy number alterations (CNA) coupled with focused candidate gene resequencing has identified novel genetic alterations that contribute to leukemogenesis and are associated with treatment outcome in ALL. However, large-scale analysis of somatic sequence mutations in ALL has not yet been performed. As part of the COG HR-ALL TARGET (Therapeutically Applicable Research to Generate Effective Targets) Project, 125 genes (selected based on recurrent CNAs, gene expression profiles and known cancer genes) were sequenced in 187 HR B-precursor ALL patients enrolled in COG P9906. The entire coding region and UTRs of each gene were sequenced and more than 98% of the targeted nucleotides have high quality sequence coverage. We found that somatic mutations are frequent in genes that encode for proteins involved in signal transduction, B-cell development and p53/RB signaling. A notable finding was the presence of somatic mutations resulting in constitutive activation of RAS signaling in at least 39% of the cohort. Seventy-three cases have at least one mutation in NRAS (30), KRAS (28), PTPN11 (9), FLT3 (7) and NF1 (6), including 7 patients with multiple mutations (KRAS and NRAS (3), FLT3 and NF1 (1), PTPN11 and FLT3 (1), PTPN11 and FLT3 (1), PTPN11 and NRAS (1), PTPN11 and KRAS (1)). There was no association between RAS pathway mutations and event-free survival or cumulative incidence of relapse in this HR patient cohort. Notably, RAS pathway mutations were uncommon in ALL cases with TCF3-PBX1 (1/22 cases) or MLL translocations (2/18 cases), but occurred frequently in cases lacking known sentinel cytogenetic lesions (68/145 cases, 47%, p<0.0001). Sequence mutations that are known or predicted to impair normal B-cell development were observed in at least 14% of the cohort (PAX5 (21), IKZF1 (7)), with confirmatory germline sequencing underway to clarify whether lymphoblast sequence alterations present in other B-cell development genes including BLK, ETV6, IKZF3, TCF3, RAG1, and BCL11A are somatic or germline. Sequence mutations disrupting TP53/RB1 signaling ((TP53 (10), RB1 (4), CDKN2A (4)) occurred in 10% of cases. Activating sequence mutations in JAK family members (JAK2 (16) and JAK1 (3)) were present in 10% of the cases. Of the 103 cases (55%) with at least one sequence mutation in these four (RAS signaling, B-cell development, p53/RB, and JAK) pathways, 30 have somatic mutations in multiple pathways. When both CNAs and sequence mutations are considered, 94% of the cases have lesions in at least one of the pathways and 31% of the cases have somatic alterations in all three of the B-cell development, RAS, and p53/RB1 signaling pathways, suggesting that aberrations in multiple pathways may be central to development of HR-ALL. Our results contrast sharply with known genetic alterations in T-cell ALL, where RAS pathway alterations are relatively uncommon (<10% of cases). PTEN somatic mutations and deletions occur in approximately 25% of cases of T-cell ALL, but we found no PTEN alterations in this cohort, suggesting differential activation of RAS and PI-3K signaling pathways in T-cell and HR B-precursor ALL. Validation of putative deleterious mutations in other sequenced genes (e.g., TBL1XR1, BLK, and CREBBP) is ongoing. These results from the TARGET COG HR-ALL Pilot Project confirm and extend prior knowledge of genetics of this subtype of ALL and point the way to new potential therapeutic strategies for this patient population.