BACKGROUND Philadelphia chromosome–like acute lymphoblastic leukemia (Ph-like ALL) is characterized by a gene-expression profile similar to that of BCR–ABL1–positive ALL, alterations of lymphoid transcription factor genes, and a poor outcome. The frequency and spectrum of genetic alterations in Ph-like ALL and its responsiveness to tyrosine kinase inhibition are undefined, especially in adolescents and adults. METHODS We performed genomic profiling of 1725 patients with precursor B-cell ALL and detailed genomic analysis of 154 patients with Ph-like ALL. We examined the functional effects of fusion proteins and the efficacy of tyrosine kinase inhibitors in mouse pre-B cells and xenografts of human Ph-like ALL. RESULTS Ph-like ALL increased in frequency from 10% among children with standard-risk ALL to 27% among young adults with ALL and was associated with a poor outcome. Kinase-activating alterations were identified in 91% of patients with Ph-like ALL; rearrangements involving ABL1, ABL2, CRLF2, CSF1R, EPOR, JAK2, NTRK3, PDGFRB, PTK2B, TSLP, or TYK2 and sequence mutations involving FLT3, IL7R, or SH2B3 were most common. Expression of ABL1, ABL2, CSF1R, JAK2, and PDGFRB fusions resulted in cytokine-independent proliferation and activation of phosphorylated STAT5. Cell lines and human leukemic cells expressing ABL1, ABL2, CSF1R, and PDGFRB fusions were sensitive in vitro to dasatinib, EPOR and JAK2 rearrangements were sensitive to ruxolitinib, and the ETV6–NTRK3 fusion was sensitive to crizotinib. CONCLUSIONS Ph-like ALL was found to be characterized by a range of genomic alterations that activate a limited number of signaling pathways, all of which may be amenable to inhibition with approved tyrosine kinase inhibitors. Trials identifying Ph-like ALL are needed to assess whether adding tyrosine kinase inhibitors to current therapy will improve the survival of patients with this type of leukemia. (Funded by the American Lebanese Syrian Associated Charities and others.)

Genetic alterations activating NOTCH1 signaling and T cell transcription factors, coupled with inactivation of the INK4/ARF tumor suppressors are hallmarks of T-ALL, but detailed genome-wide sequencing of large T-ALL cohorts has not been performed. Using integrated genomic analysis of 264 T-ALL cases, we identify 106 putative driver genes, half of which were not previously described in childhood T-ALL (e.g. CCND3, CTCF, MYB, SMARCA4, ZFP36L2 and MYCN). We described new mechanisms of coding and non-coding alteration, and identify 10 recurrently altered pathways, with associations between mutated genes and pathways, and stage or subtype of T-ALL. For example, NRAS/FLT3 mutations were associated with immature T-ALL, JAK3/STAT5B mutations in HOX1 deregulated ALL, PTPN2 mutations in TLX1 T-ALL, and PIK3R1/PTEN mutations in TAL1 ALL, suggesting that different signaling pathways have distinct roles according to maturational stage. This genomic landscape provides a logical framework for the development of faithful genetic models and new therapeutic approaches.

Recent genomic studies have identified chromosomal rearrangements defining new subtypes of B-progenitor acute lymphoblastic leukemia (B-ALL), however many cases lack a known initiating genetic alteration. Using integrated genomic analysis of 1,988 childhood and adult cases, we describe a revised taxonomy of B-ALL, incorporating 23 subtypes defined by chromosomal rearrangements, sequence mutations, or heterogeneous genomic alterations, many of which show marked variation in prevalence according to age. Two subtypes have frequent alterations of the B lymphoid transcription factor gene PAX5. One, PAX5alt (7.4%), has diverse PAX5 alterations (rearrangements, intragenic amplifications or mutations), and a second subtype is defined by PAX5 p.Pro80Arg and biallelic PAX5 alterations. We show that p.Pro80Arg impairs B lymphoid development and promotes the development of B-ALL with biallelic Pax5 alteration in vivo. These results demonstrate the utility of transcriptome sequencing to classify B-ALL and reinforce the central role of PAX5 as a checkpoint in B lymphoid maturation and leukemogenesis.

Despite great progress in curing childhood acute lymphoblastic leukemia (ALL), survival after relapse remains poor. We analyzed survival after relapse among 9585 pediatric patients enrolled on Children's Oncology Group clinical trials between 1988 and 2002. A total of 1961 patients (20.5%) experienced relapse at any site. The primary end point was survival. Patients were subcategorized by the site of relapse and timing of relapse from initial diagnosis. Time to relapse remains the strongest predictor of survival. Patients experiencing early relapse less than 18 months from initial diagnosis had a particularly poor outcome with a 5-year survival estimate of 21.0±1.8%. Standard risk patients who relapsed had improved survival compared with their higher risk counterparts; differences in survival for the two risk groups was most pronounced for patients relapsing after 18 months. Adjusting for both time and relapse site, multivariate analysis showed that age (10 þ years) and the presence of central nervous system disease at diagnosis, male gender, and T-cell disease were significant predictors of inferior post-relapse survival. It can be noted that there was no difference in survival rates for relapsed patients in earlier vs later era trials. New therapeutic strategies are urgently needed for children with relapsed ALL and efforts should focus on discovering the biological pathways that mediate drug resistance.

Philadelphia chromosome-like (Ph-like) acute lymphoblastic leukemia (ALL) is a high-risk subtype characterized by genomic alterations that activate cytokine receptor and kinase signaling. We examined the frequency and spectrum of targetable genetic lesions in a retrospective cohort of 1389 consecutively diagnosed patients with childhood B-lineage ALL with high-risk clinical features and/or elevated minimal residual disease at the end of remission induction therapy. The Ph-like gene expression profile was identified in 341 of 1389 patients, 57 of whom were excluded from additional analyses because of the presence of - (n = 46) or - (n = 11). Among the remaining 284 patients (20.4%), overexpression and rearrangement of (- or -) were identified in 124 (43.7%), with concomitant genomic alterations activating the JAK-STAT pathway (, ,) identified in 63 patients (50.8% of those with rearrangement). Among the remaining patients, using reverse transcriptase polymerase chain reaction or transcriptome sequencing, we identified targetable ABL-class fusions (, ,, and ) in 14.1%, rearrangements or fusions in 8.8%, alterations activating other JAK-STAT signaling genes (, ,) in 6.3% or other kinases (, ,) in 4.6%, and mutations involving the Ras pathway (, ,, ) in 6% of those with Ph-like ALL. We identified 8 new rearrangement partners for 4 kinase genes previously reported to be rearranged in Ph-like ALL. The current findings provide support for the precision-medicine testing and treatment approach for Ph-like ALL implemented in Children's Oncology Group ALL trials.

T-cell acute lymphoblastic leukaemia (T-ALL) is induced by the transformation of T-cell progenitors and mainly occurs in children and adolescents. Although treatment outcome in patients with T-ALL has improved in recent years, patients with relapsed disease continue to have a poor prognosis. It is therefore important to understand the molecular pathways that control both the induction of transformation and the treatment of relapsed disease. In this Review, we focus on the molecular mechanisms responsible for disease induction and maintenance. We also compare the physiological progression of T-cell differentiation with T-cell transformation, highlighting the close relationship between these two processes. Finally, we discuss potential new therapies that target oncogenic pathways in T-ALL.

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Relapsed childhood acute lymphoblastic leukemia (ALL) carries a poor prognosis despite intensive retreatment, due to intrinsic drug resistance1-2. The biological pathways that mediate resistance are unknown. Here we report the transcriptome profiles of matched diagnosis and relapse bone marrow specimens from ten pediatric B lymphoblastic leukemia patients using RNA-sequencing. Transcriptome sequencing identified 20 newly acquired novel non-synonymous mutations not present at initial diagnosis, of which two patients harbored relapse specific mutations in the same gene, NT5C2, a 5′-nucleotidase. Full exon sequencing of NT5C2 was completed in 61 additional relapse specimens, identifying five additional cases. Enzymatic analysis of mutant proteins revealed that base substitutions conferred increased enzymatic activity and resistance to treatment with nucleoside analogue therapies. Clinically, all patients who harbored NT5C2 mutations relapsed early, or within 36 months of initial diagnosis (p=0.03). These results suggest that mutations in NT5C2 are associated with the outgrowth of drug resistant clones in ALL.