Abstract PL02-02: The molecular pathology of acute leukemia
Authors: James R. Downing
Journal: Cancer Research (2011)

Abstract A goal of modern cancer research is to define the underlying genetic lesions that are responsible for driving the formation, maintenance, and metastatic potential of cancer. The hope is that the information gained from these genomic studies will result in the identification of new diagnostic and prognostic markers, as well as rational molecular targets against which novel therapeutic agents can be developed. Although these hopes are being realized across the spectrum of human cancers, their direct influence on clinical care is probably most evident in the treatment of hematopoietic malignancies. Specifically, genomic data have significantly advanced our understanding of the pathogenesis of both acute lymphoblastic leukemia (ALL) and acute myeloblastic leukemia (AML), and have resulted in the identification of molecular targets against which new therapies have been developed. Despite the progress made, however, we still lack a full understanding of the genetic lesions that underlie the process of leukemogenesis. To approach this problem, my laboratory has applied genomic methodologies including single nucleotide polymorphism (SNP) arrays, candidate gene resequencing, and whole genome and transcriptome sequencing to pediatric acute leukemia. To date, we have analyzed over 800 leukemias for copy number alterations (CNAs), along with a smaller subset of cases for DNA-sequence mutations. Our analyses have revealed marked heterogeneity in the number and type of lesions across the different subtypes of acute leukemia. For B progenitor ALL the most remarkable finding was that over 60% of cases had deletion, amplification, point mutation or structural rearrangement in genes that encode key regulators of normal B cell development and differentiation. The PAX5 gene was the most frequent target of somatic mutation, being altered in 30% of cases. Moreover, the frequency of mutations in specific B cell regulatory genes varied markedly across the different subtypes of ALL, with a near obligate deletion of IKZF1 in BCR-ABL1 ALL, a clinically aggressive ALL subtype. To explore the clinical significance of the identified CNAs, we extended our studies to a cohort of 221 high-risk ALL patients treated through the Children's Oncology Group. This analysis revealed mutations of IKZF1 as an independent predictor of a high risk of relapse. Importantly, the mutations of IKZF1 were associated with an expression signature similar to BCR-ABL1 containing cases, raising the possibility that within this subgroup of ALLs there might exist activating mutations in kinase signaling pathways. To explore this possibility, we sequenced the tyrosine kinome within a subset of these cases. Our analysis revealed heterozygous somatic mutations in JAK1 or JAK2 in a significant percentage of the IKZF1 deleted cases, with the mutations occurring in highly conserved residues in the kinase and pseudokinase domains. These mutations were shown to lead to constitutive JAK-STAT activation and growth factor independence. Surprisingly, a subset of the JAK mutant cases had marked overexpression of the cytokine receptor-like factor 2 (CRLF2). This overexpression was shown to be the result of an interstitial deletion of the pseudoautosomal region 1 of chromosomes X and Y that juxtaposed the first, noncoding exon of the purinergic receptor P2RY8 with the coding region of CRLF2, leading to overexpression of the wild-type CRLF2 protein. This rearrangement was identified in 7% of B-progenitor ALLs and 53% of ALLs associated with Down syndrome. Taken together, these data suggested that an activated JAK kinase and overexpression of CRLF2 directly cooperate in leukemogenesis. These data also suggest that inhibitors of JAK kinases may be rational therapeutic agents in these high-risk patients. Clinical protocols to test this hypothesis are now in development. Despite the impressive cure rates that have been achieved for pediatric ALL, treatment failures remain a formidable problem with relapsed ALL ranking as the fourth most common childhood malignancy. Moreover, cure rates for relapsed ALL remain dismal with overall survival rate of only 30%. To explore the genetic basis of relapse and drug resistance, we performed CNAs on matched diagnosis and relapse samples from 61 patients with ALL. In the majority of cases, the diagnosis and relapse samples showed different patterns of CNAs, with the abnormalities acquired at relapse preferentially affecting genes involved in cell cycle regulation and B cell development. Although the diagnosis and relapse samples were genetically related, most relapse samples lacked some of the CNAs present at diagnosis, suggesting that the cells responsible for relapse are ancestral to the primary leukemia cells. Backtracking studies demonstrated that cells corresponding to the relapse clone were often present as minor sub-populations at diagnosis. These data suggest that genomic abnormalities contributing to relapse are selected for during treatment and that the signaling pathways affected by these acquired alterations may be rational targets for therapeutic intervention. Using the approaches outlined above, we have also analyzed pediatric AML. Our results demonstrated that, in contrast to pediatric ALL, de novo AML was characterized by a very low burden of genomic alterations, with a mean of less than 2 somatic CNAs per leukemia. Even more surprising was the observation that 34% of the leukemias lacked any identifiable copy number alterations. The only exception to this was acute megakaryocytic leukemias, which was characterized by a high number of CNAs. These initial studies have provided important insights into the underlying molecular pathology of pediatric acute leukemias. However, these studies have been limited by a rather low level of resolution. To extend these studies to a higher level of resolution, we have recently initiated a collaborative effort called the St. Jude Children's Research Hospital, Washington University Pediatric Cancer Genome Project. This project has as its explicit goal to sequence the whole genome of matched tumor and normal samples from 600 pediatric cancer patients. For the leukemias, we have focused on the high-risk subtypes of infant ALL, early T-progenitor ALL, and acute megakaryocytic leukemia. The early data generated through this project illustrates the remarkable power of these approaches to define the underlying genetic lesions in pediatric cancer. The generated data should serve as an important resource for the community of cancer researchers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr PL02-02. doi:10.1158/1538-7445.AM2011-PL02-02