Curr Hematol Malig Rep (2012) 7:133–143 the pseudokinase domain of JAK2 [40–42], and are associated with very poor outcome [46, 47]. Therapeutically important, JAK2 mutant-CRLF2–mediated and CRLF2 mutant–mediated transformation are sensitive to JAK inhibition in vitro, suggesting that ALL patients with CRLF2 overexpression may benefit from future kinase inhibitor approaches [41]. Intrachromosomal Amplification of Chromosome 21 Intra- chromosomal amplification of chromosome 21 (iAMP21) occurs at an incidence up to 2 % in older children with B- cell precursor ALL. iAMP21 was originally identified as a consequence of routine screening for the presence of the ETV6-RUNX1 fusion by FISH [48] and is defined by at least three copies of the RUNX1 gene, commonly concomitant with deletion of the subtelomeric regions of chromosome 21. Recently, the common region of amplification on chro- mosome 21 was better refined to a 5.1-Mb region that included RUNX1, miR-802, and genes mapping to the DS critical region. Recurrent abnormalities affecting IKZF1 (22 %), CDKN2A/B (17 %), PAX5 (8 %), ETV6 (19 %), and RB1 (37 %) and secondary to chro- mosome 21 rearrangements were identified [49]. iAMP21 has been shown to be linked to a dismal outcome when patients are treated with standard therapy, because it is associated with an increased risk of both early and late relapses [50, 51]. Prognostic Factors Based on Conventional and Next-Generation Sequencing Technologies Whole genome/exome-sequencing approaches have been demonstrated to provide a comprehensive view of the landscape of genetic alterations in leukemia identifying crucial markers for diagnosis, risk-stratification, and po- tentially novel target therapies. Screening for BCR-ABL1 mutations in Philadelphia-positive ALL allows to iden- tify patients who may benefit from second-generation TKIs or from novel compounds targeting the T315I mutation [52]. An extensive Sanger resequencing of 120 candidate cancer genes in diagnostic leukemia sam- ples from 187 children and adolescents with high-risk B-ALL treated with augmented post-induction chemo- therapy on the Children’s Oncology Group P9906 pro- tocol and integrated analysis with genome-wide CNAs and gene expression profiles revealed recurrent somatic alterations in key signaling pathways, including B-cell development/differentiation (68 %), TP53/RB tumor sup- pressor pathway (54 %), Ras signaling (50 %), and Janus kinases (11 %). The frequency of mutations with- in the four major pathways varied markedly across genetic subtypes with a striking association of B-cell development/differentiation and JAK2 mutations with leukemias expressing a BCR-ABL1-like gene expression 137 profile and of Ras signaling pathway mutations with cases defined by a distinct gene expression profile cou- pled with focal ERG deletion [53]. For most of these genes the prognostic impact remains to be determined, but they highlighted important new therapeutic targets for selected patient subsets. The landscape of genetic alterations in B-ALL was extended by the identification of loss-of-function mutations (either deletions or se- quence mutations) of CREBBP, encoding the transcrip- tional coactivator and histone acetyltransferase CREB- binding protein or CBP, in 19 % of relapsed samples from children with B-cell progenitor ALL [54 ]. Func- tionally, the mutations impair histone acetylation and transcriptional regulation of CREBBP targets, including glucocorticoid responsive genes, likely influencing treat- ment responsiveness in ALL. CREBBP mutations are preserved at relapse when present at diagnosis, acquired at relapse but occurring in minor subclones at diagnosis, or reduplicated to homozygosity at relapse when heterozygous at diagnosis [54 ]. Although the prognostic impact remains to be determined, the observation that the CREBBP mutations impair regulation of glucocorticoid-responsive genes, and that the mutations are selected for at relapse, suggests that these alterations may influence the likelihood of relapse. Therefore, novel therapeutic approaches, such as histone deacetylase inhibitors (HDAC) [55], directed at modulating protein acet- ylation may be useful in high-risk ALL. Recently, new mo- lecular prognostic markers have been identified. The analysis of 265 first-relapse patients enrolled in the German Acute Lymphoblastic Leukemia Relapse Berlin-Frankfurt-Münster 2002 (ALL-REZ BFM 2002) trial for sequence and copy number alterations of the TP53 gene, by using direct sequenc- ing and multiplex ligation-dependent probe amplification, identified copy number and sequence alterations of TP53 in 12.4 % of B-cell precursor ALL samples [56]. In more than half of cases (54 %) TP53 alterations were gained at relapse. Mutations were strikingly associated with non- response to chemotherapy and poor event-free survival and overall survival rates, allowing the identification of patients at high risk of treatment failure. To overcome the limits of conventional Sanger sequencing whole exome and transcriptome NGS approaches have been subsequently pursued in two pediatric ETV6-RUNX1 (TEL-AML1) positive ALL [57] and in an adult patient with BCR-ABL1 positive ALL [58], respectively. Both studies identified novel somatic mutations, some of them occurred in genes previously described to be mu- tated in other types of cancer, but none was found to be recurrent in an extended series of ALL samples. Although the true pathogenetic significance of the mutations must await future functional evaluations, these studies provided a first estimate of the mutational burden of ETV6-RUNX1 and BCR-ABL1-positive ALLs.