SECTION A Genetic Alterations in High-Risk B-Progenitor Acute Lymphoblastic Leukemia Charles G. Mullighan Abstract adulthood, cure rates for ALL fall sharply, and the genetic and biologic determinants of Recent studies profiling genetic alterations in treatment failure remain incompletely B-progenitor acute lymphoblastic leukemia understood (8,9). (B-ALL) at high resolution have identified multiple recurring submicroscopic genetic alterations ALL has long been exceptionally well targeting key cellular pathways in lymphoid cell characterized at the cytogenetic level, and growth, differentiation and tumor suppression. approximately three quarters of childhood ALL A key finding has been that genetic alterations cases harbor recurring gross genetic alterations, disrupting normal lymphoid growth and including chromosomal aneuploidy (high differentiation and are associated with treatment hyperdiploidy and hypodiploidy), and outcome. Notably, genetic alterations targeting rearrangements that dysregulate hematopoietic lymphoid development are present in over two- regulators, transcription factors, and tyrosine thirds of B-ALL cases, including deletions, kinases. These include rearrangements resulting translocations and sequence mutations of the in (e.g. ETV6-RUNX1, TCF3-PBX1, BCR-ABL1, transcriptional regulators PAX5, IKZF1, and EBF1. rearrangement of MLL, and rearrangements of Deletion or mutation of the early lymphoid T cell receptor genes to hematopoietic regulators transcription factor gene IKZF1 is hallmark of and transcription factors in T-lineage ALL) multiple subtypes of ALL with poor prognosis, (10,11). While alterations associated with including BCR-ABL1 positive (Ph+) lymphoid favorable outcome (e.g. high hyperdiploidy and leukemia and a novel subset of BCR-ABL1-like ETV6-RUNX1) are characteristic of childhood ALL cases that have a gene expression profile ALL, and the frequency of BCR-ABL1 similar to that of Ph+ B-ALL, but lack expression (Philadelphia chromosome positive, or Ph+) ALL of BCR-ABL1. In addition to deletion of IKZF1, rises with age (12), the differences in the these BCR-ABL1-like cases commonly harbor frequencies of these recurring gross genetic mutations resulting in aberrant lymphoid chromosomal rearrangements is insufficient to cytokine receptor signaling, including activating fully explain treatment failure in ALL, which mutations of Janus kinases and rearrangement occurs across the spectrum of cytogenetic of CRLF2 (cytokine receptor-like factor 2). These subtypes, including cases that lack cytogenetic findings demonstrate that multiple genetic alterations. Consequently, there has been great alterations disrupting different cellular pathways interest in using genome-wide approaches to are key events in the pathogenesis of high risk identify submicroscopic genetic alterations in ALL, and suggest that novel therapies targeting ALL, and these studies have proven aberrant cytokine receptor signaling may be of exceptionally fruitful in identifying new mutations therapeutic benefit in high risk ALL cases. that target key cellular pathways in B-progenitor and T-lineage ALL. Similar studies have also Acute lymphoblastic leukemia (ALL) is the been informative in T-lineage ALL (13-16), commonest childhood cancer (1,2), and despite however this review focuses primarily on impressive advances in the outcome of therapy B-progenitor ALL and recent studies that have with cure rates now exceeding 80% (3, 4), identified genetic markers of treatment failure in remains a leading cause of cancer-related death this disease. in children and young adults (5-7). Moreover, with increasing age through adolescence and 77 The spectrum of genetic alterations in ALL cases harbor fewer than one copy number insights from genome-wide profiling alteration per case. This suggests that few Multiple studies have used array-based cooperating structural genetic alterations are comparative genomic hybridization and single required to induce leukemia (13, 26). In contrast, nucleotide polymorphism (SNP) microarrays to ETV6-RUNX1 and BCR-ABL1 (Ph+) ALL cases identify submicroscopic genetic alterations in harbor multiple distinct copy number alterations childhood ALL. Fewer studies have examined (13, 27). Deletion ofIKZF1 (IKAROS) is a hallmark young adult and older patients with ALL, and of Ph+ lymphoid leukemia, including both detailed profiling of these cohorts is required. In childhood and adult de novo ALL cases (18, 28) contrast to many solid tumors, which commonly and chronic myeloid leukemia (CML) at harbor multiple gross DNA copy number progression to lymphoid blast crisis (18, 28). alterations (17), ALL his characterized by a Moreover, the presence of IKZF1 alterations is relatively low number of genetic alterations associated with poor outcome in Ph+ ALL (29). approximately six to eight lesions per case (13, IKAROS is a member of a family of zinc-finger 18). However, recurring submicroscopic genetic transcription factors that has multiple, alterarations targeting key cellular pathways and incompletely understood functions in lymphoid genes with key roles in leukemia development development and leukemogenesis, including are a hallmark of ALL (13, 19, 20). These include transcriptional regulation and chromatin mutations targeting transcriptional regulators of remodeling. Normal IKAROS function is required lymphoid development (e.g. PAX5, IKZF1 and for the development of all lymphoid lineages (30- EBF1), cell cycle regulators and tumor 34, 35 ). Expression of aberrant IKAROS isoforms suppressor genes (CDKN2A, CDKN2B, RB1 and in ALL blasts is well recognized, notably that of PTEN), lymphoid signaling genes (CD200, BTLA one isoform, IK6. This isoform lacks the N- and BLNK) and drug response genes (e.g. the terminal zinc fingers of IKAROS and cannot bind glucocorticoid receptor gene NR3C1). Genes DNA, but retains the C-terminal zinc fingers and regulating B lymphoid development are mutated can act in a dominant-negative fashion (36-38). in the majority of B-progenitor ALL cases, most However, SNP array profiling studies of both ALL commonly deletions, sequence mutations or and CML have shown that expression of these translocations of PAX5 (13, 19-21), deletion (and dominant-negative transcripts is determined by less commonly, sequence mutation) of IKZF1 the presence of IKZF1 deletions that involve the (IKAROS) and the IKAROS family members exons corresponding to those deleted in the IKZF2 (HELIOS) and IKZF3 (AIOLOS) and aberrant IKZF1 transcripts and IKZF1 protein deletion of EBF1. These mutations result in loss (18,28). of function in vitro (13), and accelerate the onset Alterations in IKAROS function have previously of ALL in murine models of ALL (22-24). While been reported to have an important role in the the mutations most commonly involve only a pathogenesis of lymphoid tumors. Mice single copy of the affected gene, multiple harboring a dominant-negative mutation in the mutations involving this pathway are common Ikzf1 gene develop aggressive T-lineage in high-risk B-ALL, and a higher number of lymphoproliferative disease (39). Although the lesions in the pathway is associated with poor role of IKAROS in the pathogenesis of Ph+ ALL outcome (25), suggesting the degree of block remains to be fully defined, existing data have in B cell differentiation induced by mutations shown that expression of IK6 impairs B lymphoid in this pathway not only contributes to maturation (40, 41) and pre-B cell receptor leukemogenesis, but also treatment signaling in Ph+ ALL cells (42). Moreover, responsiveness. deletion of Ikzf1 accelerates leukemogenesis in a murine model of Ph+ ALL (24). Genomic profiling of high-risk ALL a central role of IKZF1 A role for IKAROS in the pathogenesis of ALL is also supported by recent data from genome- The frequency and nature of submicroscopic wide association studies in which an inherited genetic alterations in ALL is strongly associated SNP allele at the IKZF1 locus was associated with disease subtype. MLL-rearranged ALL with the risk of childhood ALL, a finding that has 78 SECTION A been identified in multiple studies and patient by BCR-ABL1. A substantial proportion of cohorts (43-45). The mechanistic basis of this BCR-ABL1-like ALL cases have genetic finding remains unclear, although it is notable alterations that result in aberrant cytokine that IKZF1 genotype was associated with the receptor signaling, notably activating Janus level of expressionof IKAROS (43), and the genes kinase (JAK) mutations and rearrangement of at the two other loci found to be associated with CRLF2 (encoding the lymphoid cytokine ALL risk in these studies, ARID5B and CEBPE, receptor gene cytokine receptor like factor 2). also encode transcriptional regulators and genes involved in lymphoid maturation (46,47), Genetic characterization of BCR-ABL1-like, suggesting that germline variation at these loci Ph negative ALL directly influences the risk of ALL. Detailed candidate gene resequencing in high Alterations of IKZF1 are also associated with poor risk ALL, including targets of DNA copy number outcome in Ph- ALL. A Childrens Oncology alteration, dysregulated gene expression, and Group study of over 200 cases of high-risk a subset of receptor and non-receptor tyrosine B-progenitor Ph-negative ALL identified IKZF1 kinases in 187 high-risk B-progenitor ALL cases deletions and sequence mutations in from the Childrens Oncology Group P9906 approximately one third of cases. This study also cohort (25) identified 20 cases with somatic found