Introduction: Ph+ ALL comprises ~5% of childhood and adolescent ALL. Prior to development of tyrosine kinase inhibitor (TKI) therapy, survival rates were poor. Less than half of patients (pts) survived despite treatment with intensive chemotherapy and frequent use of allogeneic hematopoietic stem cell transplant (HSCT) in first remission (CR1). The Children's Oncology Group (COG) AALL0031 trial (Schultz, JCO 2009) and the EsPhALL trial (Biondi, Lancet Oncology 2012) showed adding imatinib to different intensive chemotherapy backbones improved event-free (EFS) and overall survival (OS) in pediatric Ph+ ALL. Dasatinib is attractive to study in Ph+ ALL because it is a dual ABL/SRC TKI that is 300 times more potent than imatinib in vitro, is active against most ABL1 TKI domain mutations that cause imatinib resistance, and accumulates in the central nervous system (CNS), a sanctuary site for ALL where imatinib penetration is poor. Methods: We conducted a phase 2 trial of dasatinib added to the EsPhALL chemotherapy backbone in pediatric (>1-17.99 years (yrs) of age) Ph+ ALL pts at COG sites in North America and Australia and EsPhALL sites in Italy and the United Kingdom. Protocol therapy added continuous daily dasatinib (60 mg/m2) at day 15 of induction chemotherapy. The study measured minimal residual disease (MRD) by Ig/TCR PCR, flow cytometry, and BCR - ABL1 RT-PCR, with clinical actions based upon a single method, in this hierarchical order. Pts with MRD ≥ 0.05% at the end of block Ib (day 78) and those with MRD 0.005-0.05% at end of Ib who remained MRD positive at any detectable level after three additional high-risk (HR) chemotherapy blocks underwent HSCT in CR1. Dasatinib treatment post HSCT was optional. The remaining pts received chemotherapy plus daily dasatinib for 2 yrs, with cranial irradiation limited to CNS3 pts. The primary study endpoint was 3-year EFS assessed when all patients completed 3 years of follow-up. Results: From April 2012 to May 2014, 109 pts enrolled; 3 did not meet inclusion criteria and received no trial therapy. The median age was 9.0 yrs (range 1-17), 54% were males, and 80% were Caucasian. 71% had CNS1 status at baseline, 24% CNS2, and 5% CNS3. Safety analysis included all treated pts (N=106) and efficacy analysis included all treated Ph+ ALL pts (N=104; 2 pts were retrospectively diagnosed with blast crisis CML). The database lock date was 8/17/16; at this time all pts had completed therapy and 75% had ≥3 yrs of follow-up. Two pts discontinued dasatinib for toxicity (1 allergy and 1 prolonged myelosuppression post HSCT). Nineteen pts met study criteria for HSCT, and 15 received HSCT in CR1 (14.2% of pts). The remaining 91 pts (85.8%) received EsPhALL chemotherapy plus dasatinib without HSCT. Patients tolerated dasatinib combined with chemotherapy well. The primary toxicity was febrile neutropenia and infection: Grade 3+ febrile neutropenia occurred in 75.5% of pts, Grade 3+ sepsis in 18.9%; and Grade 3+ bacteremia in 13.2%. Elevated ALT (21.7%) and AST (10.4%) were the only non-hematologic, non-infectious Grade 3+ adverse events attributed to dasatinib reported in >10% of pts. Relevant Grade 3+ non-hematologic, non-infectious toxicities attributed to dasatinib included pleural effusion (3.8%), edema (3.8%), hemorrhage (2.8%), and cardiac failure (0.8%). No cases of pulmonary hypertension or pulmonary arterial hypertension were reported. All 104 treated Ph+ ALL pts achieved CR. As of the database lock date, 33 events had occurred including 5 deaths (3 in HR3 and 2 in reinduction) due to proven or suspected infection in the 91 patients receiving chemotherapy plus dasatinib, 2 deaths from infection post-HSCT in the 15 HSCT pts, and 26 relapses (chemotherapy 22/86; HSCT 4/12). Sites of relapse included isolated bone marrow (BM; 14), CNS (4), BM+CNS (3), BM+other (2), and other (3). At the time of the interim analysis the 3-yr EFS is 66.0% (95% CI, 54.8-75.0) and the 3-yr OS is 92.3% (95% CI, 85.2-96.1); updated results with all patients having at least 3 years of follow-up will be presented. Conclusions: Addition of dasatinib to the EsPhALL chemotherapy regimen is safe and effective in pediatric Ph+ ALL pts. With only 14% of pts undergoing SCT in CR1, as compared to 80% in the EsPhALL imatinib trial, this trial demonstrates similar outcomes with 3-yr EFS/OS 66.0%/92.3% in this trial vs. published 4-yr EFS/OS 61.9%/72.1% in the EsPhALL imatinib trial. Disclosures Hunger: Novartis: Consultancy; Jazz Pharmaceuticals: Honoraria; Erytech Pharmaceuticals: Consultancy; Amgen: Consultancy, Equity Ownership. Saha: Shire: Research Funding. Gastier Foster: Bristol-Myers Squibb: Research Funding. Cazzaniga: Italian Association for Cancer Research: Research Funding; Fondazione Tettamanti onlus: Employment. Borowitz: Beckman Coulter: Honoraria; Becton-Dickinson Biosciences: Research Funding; HTG Molecular: Honoraria. Gramatges: Bristol Meyer Squibb: Research Funding. Sun: Baxalta: Consultancy. Swanink: Bristol-Myers Squibb: Employment. Schrappe: Baxalta: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; JAZZ Pharma: Consultancy, Research Funding; SigmaTau: Consultancy, Research Funding; Medac: Consultancy, Research Funding. Healey: Bristol-Myers Squibb: Employment, Equity Ownership; Pfizer: Equity Ownership.

No abstract

Introduction B progenitor acute lymphoblastic leukemia (B-ALL) is a leading cause of childhood cancer death. Many chimeric genes have been identified and led to a refined classification of B-ALL and tailored therapies. Still, up to 30% of B-ALL cannot be classified into established subtypes, and the outcome for many is poor. Methods To identify novel subtypes of B-ALL, we performed integrative genomic analysis including transcriptome sequencing (RNA-seq) of 1,988 cases from St. Jude, Children's Oncology Group and adult cooperative group studies and analyzed chromosomal rearrangements, gene-expression profiles (GEP), somatic mutations and chromosome-level copy-number alterations. Cases lacking known or putative subtype-defining alterations underwent whole genome sequencing. Effects on proliferation and transformation of novel lesions were assessed by retroviral expression in cell lines and point-mutation knock-in mice using CRISPR/Cas9 genome editing. Results Using integrated genetic alterations and gene expression profiling, we classified 23 B-ALL subtypes (Table and Figure). Three groups included cases with similar GEP as canonical subtypes (ETV6-RUNX1, KMT2A-rearranged, and ZNF384-rearranged), but lacking the expected drivers (e.g., ETV6-RUNX1-like, n=42). Eighteen cases (0.9%) had rearrangements of BCL2, MYC and/or BCL6 showing a distinct GEP; they were mostly adults (n=16) with very poor outcome. These alterations, rarely seen in ALL, are identical to those observed in "double/triple hit" lymphoma, and are of pre-B immunophenotype. Eight cases with tightly clustered GEP comprised a novel subtype defined by IKZF1 N159Y missense mutation. N159Y is in the DNA-binding domain of IKZF1, and is known to perturb IKZF1 function, with distinct nuclear mislocalization and induction of aberrant intercellular adhesion. We identified two subtypes with distinct GEP characterized by PAX5 alterations. One, herein termed PAX5 altered (PAX5alt), comprised 148 (7.4%) cases, was characterized by diverse PAX5 alterations including rearrangements (n=57), sequence mutations (n=46) and/or focal intragenic amplifications (n=8). These PAX5 alterations were found in 73.6% of PAX5alt cases and different alteration types were mutually exclusive. Other PAX5 alterations, including deletions and large-scale amplifications were also assessed using SNP array, but were not enriched in the PAX5alt group. Clinically, PAX5alt pediatric and adult patients had favorable (96.8±3.2%) and intermediate (42.1±10.2%) 5-year overall survival (OS), respectively. The other GEP distinct subtype comprised 44 cases, all with PAX5 P80R missense mutations. In 30 of these cases, PAX5 P80R was homozygous due to deletion of the wild-type (WT) PAX5 allele or copy-neutral loss of heterozygosity. Of the other 14 cases with heterozygous PAX5 P80R mutations, 13 had a second frameshift (n=7), nonsense (n=2) or deleterious missense (n=4) PAX5 mutation. Four of the remaining 1,944 cases also had the PAX5 P80R mutation, but all were heterozygous with preservation of a WT PAX5 allele, consistent with the notion that homozygous or compound heterozygous PAX5 P80R mutation is the hallmark of this subtype. Adult PAX5 P80R cases (n=14) showed better 5-year OS (61.9±13.4%) than those in PAX5alt subtype (42.1±10.2%). To examine the effects of PAX5 P80R on B-cell maturation, WT PAX5, PAX5 P80R, V26G and P34Q were expressed in Pax5-/- lineage-depleted bone marrow cells. Expression of WT PAX5, PAX5 V26G and P34Q resulted in near complete rescue of B cell differentiation; however, expression of PAX5 P80R blocked the differentiation at the pre-pro-B stage of B-cell maturation. Further, Pax5 P80R heterozygous or homozygous mice developed pre-B-ALL with a median latency of 166 and 87 days, respectively, with heterozygous mice acquiring alterations on the second allele. In contrast, Pax5+/- mice, and those harboring G183S mutation observed in familial leukemia, do not spontaneously develop B-ALL. Conclusions These results show the utility of transcriptome sequencing in defining subtypes and founding genetic alterations in B-ALL, provide a revised taxonomy of the disease across the age spectrum, and reinforce the central role of PAX5 as a checkpoint in B lymphoid maturation and leukemogenesis. Disclosures McKay: ImmunoGen Inc.: Employment. Tallman:Orsenix: Other: Advisory board; AROG: Research Funding; BioSight: Other: Advisory board; Cellerant: Research Funding; AbbVie: Research Funding; Daiichi-Sankyo: Other: Advisory board; ADC Therapeutics: Research Funding. Stock:Jazz Pharmaceuticals: Consultancy. Konopleva:Stemline Therapeutics: Research Funding. Relling:Shire Pharmaceuticals: Research Funding. Mullighan:Cancer Prevention and Research Institute of Texas: Consultancy; Amgen: Honoraria, Speakers Bureau; Abbvie: Research Funding; Loxo Oncology: Research Funding; Pfizer: Honoraria, Research Funding, Speakers Bureau.