Genomic Characterization and Experimental Modeling Of BCR-ABL1-Like Acute Lymphoblastic Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 232-232 ◽  
Author(s):  
Kathryn G. Roberts ◽  
Yongjin Li ◽  
Debbie Payne-Turner ◽  
Richard C. Harvey ◽  
Jinjun Cheng ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Young Adult ◽  
High Risk ◽  
Tyrosine Kinase ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Hematopoietic Progenitor Cell ◽  
Oncology Group ◽  
Genomic Landscape ◽  
Primary Mouse

Abstract BCR-ABL1-like B-progenitor acute lymphoblastic leukemia (B-ALL) accounts for 10-15% of childhood B-ALL and is characterized by alteration of IKZFI, a gene expression profile similar to BCR-ABL1 ALL and poor outcome. Using next-generation sequencing, we have shown that BCR-ABL1-like ALL patients harbor genetic alterations activating kinase pathways that are sensitive to tyrosine kinase inhibitors (TKIs), and have shown that refractory BCR-ABL1-like ALL is responsive to TKIs in vivo (Weston et al., J. Clin. Oncol 2013). Furthermore, the outcome of ALL in adolescent and young adult (AYA) patients is inferior to children, yet the genetic basis underlying treatment failure is poorly understood. To define the frequency and genomic landscape of BCR-ABL1-like ALL in children, adolescents, and young adults we have extended our studies to include 665 high-risk childhood (<16 years, 14% BCR-ABL1-like), 370 adolescent (16-21 years, 21% BCR-ABL1-like) and 161 young adult (21-39 years; 26% BCR-ABL1-like) B-ALL cases from the Children's Oncology Group, St Jude Children's Research Hospital, Eastern Cooperative Oncology Group, MD Anderson Cancer Center and the Alliance - CALGB trials. Event-free survival (EFS) for BCR-ABL1-like cases was inferior to non BCR-ABL1-like cases with 5-year EFS rates of 40.0±7.1 vs 85.0±3.3 (p<0.0001) for adolescent cases and 16.1±8.5 vs 57.9±8.0 (p=0.006) for young adult cases. In each age group, 50-60% of BCR-ABL1-like cases harbored rearrangements of CRLF2 (IGH@-CRLF2 or P2RY8-CRLF2) (Fig. 1). To characterize the full spectrum of kinase lesions in the remaining BCR-ABL1-like ALL cases we performed mRNA-seq on pediatric (n=39), adolescent (n=21) and young adult (n=22) cases, and whole genome (WGS; n=18) or exome sequencing (n=10) on cases with matched tumor and normal material. Fusion transcripts were identified using deFuse and CICERO, a novel assembly-based structural variation detection method specifically designed for mRNA-seq analysis. We identified 23 different kinase rearrangements involving 7 tyrosine kinase or cytokine receptor genes. These consist of 5 ABL1, 2 PDGFRB, 8 JAK2 fusions and 2 EPOR translocations to IGH@ and IGK@ loci, along with new fusions involving the tyrosine kinases ABL2 (n=3), CSF1R (n=1), AKT2 (n=1) and STAT5B (n=1). We performed frequency testing for 15 of these fusions on 555 cases from the COG AALL0232 trial of high-risk B-ALL. Several alterations were recurrent in BCR-ABL1-like ALL, including NUP214-ABL1, RCSD1-ABL2, SSBP2-CSF1R, PAX5-JAK2 and EPOR translocations. Notably, we did not identify any of these fusions in non BCR-ABL1-like cases. The frequency of ABL1/ABL2 and EPOR translocations was consistent across all age groups (∼16% and 7% of BCR-ABL1-like cases, respectively), while JAK2 rearrangements were more common in young adult than in pediatric and adolescent ALL (12%). Importantly, ∼10% of BCR-ABL1-like ALL cases lacked a kinase-activating alteration on analysis of mRNA-seq data. Notably, we identified two additional cases with IL7R or SH2B3 sequence mutations, indicating the requirement for complementary approaches such as WGS to fully define the genomic landscape of BCR-ABL1-like ALL. Current functional studies include the development of experimental models using the Ba/F3 hematopoietic progenitor cell line, primary mouse pre-B cultures and the generation of xenografts to determine the role of these alterations in leukemogenesis, and to enable testing of targeted therapies. For example, we show that RCSD1-ABL1 and SSBP2-CSF1R confer factor-independent growth and constitutive activation of JAK/STAT pathways in Ba/F3 cells. Furthermore, RCSD1-ABL1 and SSBP2-CSF1R are both sensitive to the TKIs, imatinib (IC50 378nM and 327nM, respectively) and dasatinib (IC50 2.1nM and 2.5nM, respectively). Together, these complementary approaches will further define the genetic landscape of both pediatric and AYA ALL, and facilitate the development of diagnostic and therapeutic strategies to improve the treatment outcome for high-risk BCR-ABL1-like ALL patients. Disclosures: Hunger: Bristol Myers Squibb: Consultancy.

scholarly journals Genetic Alterations Activating Kinase and Cytokine Receptor Signaling in High-Risk Acute Lymphoblastic Leukemia

Cancer Cell ◽  
2012 ◽  
Vol 22 (2) ◽  
pp. 153-166 ◽  
Author(s):  
Kathryn G. Roberts ◽  
Ryan D. Morin ◽  
Jinghui Zhang ◽  
Martin Hirst ◽  
Yongjun Zhao ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Cytokine Receptor ◽  
Receptor Signaling

scholarly journals BCR/ABL1-Like Acute Lymphoblastic Leukemia: From Diagnostic Approaches to Molecularly Targeted Therapy

2021 ◽  
pp. 1-9
Author(s):  
Anna Płotka ◽  
Krzysztof Lewandowski
Keyword(s):  
Flow Cytometry ◽  
Targeted Therapy ◽  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Diagnostic Analysis ◽  
Cell Precursor ◽  
Molecularly Targeted Therapy ◽  
Diagnostic Approaches

<b><i>Background:</i></b> <i>BCR/ABL1</i>-like acute lymphoblastic leukemia is a newly recognized high-risk subtype of ALL, characterized by the presence of genetic alterations activating kinase and cytokine receptor signaling. This subtype is associated with inferior outcomes, compared to other B-cell precursor ALL. <b><i>Summary:</i></b> The recognition of <i>BCR/ABL1</i>-like ALL is challenging due to the complexity of underlying genetic alterations. Rearrangements of <i>CRLF2</i> are the most frequent alteration in <i>BCR/ABL1</i>-like ALL and can be identified by flow cytometry. The identification of <i>BCR/ABL1</i>-like ALL can be achieved with stepwise algorithms or broad-based testing. The main goal of the diagnostic analysis is to detect the underlying genetic alterations, which are critical for the diagnosis and targeted therapy. <b><i>Key Messages:</i></b> The aim of the manuscript is to review the available data on <i>BCR/ABL1</i>-like ALL characteristics, diagnostic algorithms, and novel, molecularly targeted therapeutic options.

scholarly journals Targetable kinase gene fusions in high-risk B-ALL: a study from the Children’s Oncology Group

Blood ◽  
2017 ◽  
Vol 129 (25) ◽  
pp. 3352-3361 ◽  
Author(s):  
Shalini C. Reshmi ◽  
Richard C. Harvey ◽  
Kathryn G. Roberts ◽  
Eileen Stonerock ◽  
Amy Smith ◽  
...  
Keyword(s):  
High Risk ◽  
Tyrosine Kinase ◽  
Effective Treatment ◽  
Pediatric Patients ◽  
Genetic Alterations ◽  
Gene Fusions ◽  
Oncology Group ◽  
Kinase Gene ◽  
Tyrosine Kinase Signaling ◽  
Key Points

Key Points Ph-like ALL is characterized by a diverse array of genetic alterations activating cytokine receptor and tyrosine kinase signaling. Pediatric patients with Ph-like ALL can be identified in real time for effective treatment stratification.

Thymic Stromal Lymphopoietin Stimulation of Pediatric Acute Lymphoblastic Leukemias with CRLF2 Alterations Induces JAK/STAT and PI3K Phosphosignaling

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 410-410 ◽  
Author(s):  
Sarah K Tasian ◽  
Michelle Y Doral ◽  
Brent L Wood ◽  
Michael J Borowitz ◽  
J. Racquel Collins-Underwood ◽  
...  
Keyword(s):  
High Risk ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Thymic Stromal Lymphopoietin ◽  
Jak Inhibitor ◽  
Oncology Group ◽  
Term Survival ◽  
Phosphoflow Cytometry ◽  
Stimulation Of

Abstract Abstract 410 Collaborative genomic profiling efforts though the National Cancer Institute's TARGET Initiative and the Children's Oncology Group have identified CRLF2 and JAK mutations in a subset of children with high-risk acute lymphoblastic leukemia (ALL), but few biochemical studies have assessed the functional sequelae of these genetic alterations. CRLF2 encodes the thymic stromal lymphopoietin (TSLP) receptor chain, which heterodimerizes with the IL-7 receptor alpha chain (IL-7Rα). Children with high CRLF2-expressing ALL detected by gene expression profiling have high rates of minimal residual disease at end-induction (Day 29), and approximately 70% of these patients ultimately relapse (Harvey et al., Blood 2010). We hypothesize that characterization of aberrant signaling networks in these leukemias will facilitate identification of potential targets for small molecule inhibitor therapies. Using phosphoflow cytometry, we analyzed the phosphorylation status of key signaling molecules after stimulation with TSLP, IL-7, or pervanadate (an irreversible proximal membrane phosphatase inhibitor used as a positive control) in 2 human ALL cell lines with CRLF2 and JAK2 mutations and in 43 fresh or cryopreserved diagnostic primary patient samples, 27 of which overexpressed CRLF2 through P2RY8-CRLF2 fusion or CRLF2-IgH translocation and 16 of which did not have CRLF2 or JAK mutations (controls). Cells were rested in serum-free media for 60 minutes at 37°C, then stimulated with TSLP, IL-7, or pervanadate for 30 minutes to induce signaling. Cells were also exposed to the JAK inhibitor XL019 (Exelixis) for 60 minutes and/or subsequently stimulated with the aforementioned cytokines or pervanadate to determine the effects of JAK inhibition on signaling. Cells were then processed for phosphoflow cytometry according to our previously published methodologies (Kotecha et al., Cancer Cell 2008). High CRLF2-expressing leukemias (n = 27) with or without concomitant JAK mutations demonstrated strong surface staining of the TSLP receptor, as well as CD10, CD19, and CD127 (IL-7Rα). In vitro stimulation of leukemic blasts with TSLP elicited phosphorylation of STAT5 and S6, but not ERK 1/2, in leukemias with JAK and/or CRLF2 alterations. Control leukemias without CRLF2 and JAK mutations (n=16) did not stain for the TSLP receptor, and TSLP stimulation did not elicit phosphosignaling through the JAK/STAT, PI3K, or MAPK pathways. STAT5 and S6 phosphorylation in the high CRLF2-expressing leukemias was further abrogated by in vitro JAK inhibition with XL019. Surprisingly, despite flow cytometric staining for CD127, stimulation with IL-7 did not elicit phosphosignaling through these epitopes in high CRLF2-expressing or control leukemic blasts, although it did predictably phosphorylate STAT5 in control T and non-blast B cells contained within the primary patient leukemia samples. These results suggest that the JAK/STAT and PI3K pathways, but not the MAPK pathway, are involved in TSLP receptor signaling in high CRLF2-expressing ALL +/− JAK mutations and may represent druggable targets. Phosphoflow cytometry is an efficient method of interrogating intracellular signaling at a single-cell level in primary human samples and, furthermore, can be used for rapid identification of patients at time of leukemia diagnosis with high CRLF2-expressing ALL who exhibit the TSLP phosphosignature. Therapy for this subset of high-risk patients might be modified to include a targeted therapeutic (such as a JAK inhibitor) to improve initial treatment responses and, ultimately, to enhance long-term survival. To this end, we have developed a Children's Oncology Group Phase I clinical trial of JAK inhibition for patients with relapsed or refractory leukemias (including those with CRLF2 and JAK mutations) and will validate the use of phosphoflow cytometry and other biologic assays to assess in vivo target inhibition during therapy. We ultimately envision incorporation of JAK inhibitor therapy into a systemic chemotherapy backbone for patients with high CRLF2-expressing ALL. Disclosures: No relevant conflicts of interest to declare.

Novel Chromosomal Rearrangements and Sequence Mutations in High-Risk Ph-Like Acute Lymphoblastic Leukemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 67-67
Author(s):  
Kathryn G. Roberts ◽  
Ryan D Morin ◽  
Jinghui Zhang ◽  
Martin Hirst ◽  
Richard C. Harvey ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Cytokine Receptor ◽  
Negative Regulator ◽  
Rna Seq ◽  
Kinase Signaling ◽  
Rt Pcr ◽  
B Lineage

Abstract Abstract 67 Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy, and relapsed B-lineage ALL remains a leading cause of cancer death in young people. Recent genomic analyses by our group and others identified a unique subtype of BCR-ABL-negative, high-risk B-ALL, with deletion or mutation of IKZF1 and a gene expression profile similar to BCR-ABL1-positive ALL (Ph-like ALL). Up to 50% of Ph-like patients harbor rearrangements of the cytokine receptor gene, CRLF2, with concomitant JAK mutations detected in ∼30%. However, the nature of genetic alterations activating kinase signaling in the remaining cases is unknown. To identify novel genetic alterations in Ph-like ALL, we performed transcriptome sequencing (RNA-seq) on 11 cases of Ph-like B-ALL (10 from the P9906 Children's Oncology Group trial and 1 from the St Jude Total XV study), and whole genome sequencing (WGS) on two of these. Using multiple complementary analysis pipelines including deFuse, Mosaik, CREST and CONSERTING, we identified novel rearrangements, structural variations and sequence mutations dysregulating cytokine receptor and kinase signaling in 10 cases. Putative rearrangements and sequence mutations were validated using RT-PCR, genomic PCR and Sanger sequencing. The spectrum of alterations included 3 cases with known IGH@CRLF2 rearrangement, 2 cases with the NUP214-ABL1 rearrangement, 1 case each with the in-frame fusions EBF1-PDGFRB, BCR-JAK2 or STRN3-JAK2, and 1 case with a cryptic IGH@-EPOR rearrangement. Detailed analysis of RNA-seq data revealed a 7.5 kb insertion of EPOR downstream of the enhancer domain in the IGH@ locus, which was not detected by fluorescence in situ hybridization. WGS identified an in-frame activating insertion in the transmembrane domain of IL7R (L242>FPGVC) in 1 index case, and recurrence screening identified similar IL7R sequence mutations in 8 cases from the P9906 cohort (N=188). This patient also harbored a focal homozygous deletion removing the first two exons of SH2B3 that was not evident by SNP array analysis. SH2B3 encodes LNK, a negative regulator of JAK2 signaling. Notably, all patients harbor genetic lesions affecting B-lymphoid development (e.g IKZF1), suggesting these events cooperate to drive B-lineage ALL. To determine the frequency of each fusion, candidate RT-PCR was performed on 231 cases from the COG AALL0232 trial of high-risk B-ALL, 40 (17%) of which were identified as Ph-like using Predictor Analysis of Microarrays (PAM). The EBF1-PDGFRB fusion was detected in 3 additional patients, each containing an intact PDGFRB kinase domain. No additional cases of NUP214-ABL1, BCR-JAK2, or STRN3-JAK2 were identified. Phosphoflow analysis on 3 primary ALL samples demonstrated increased CKRL phosphorylation in the NUP214-ABL1 case and tyrosine phosphorylation in the cases with BCR-JAK2 and STRN3-JAK2 fusions. Importantly, this activation was reduced with the tyrosine kinase inhibitors (TKI) imatinib, dasatinib and the T315I inhibitor XL228 in cells harboring the ABL1 fusion, and the JAK2 inhibitor, XL019, in the JAK2-rearranged samples. Furthermore, the novel EBF1-PDGFRB fusion transformed Ba/F3 cells to growth factor independence, induced constitutive activation of pSTAT5, pAkt, pERK1/2, and responded with low IC50 values to imatinib, dasatinib and the specific PDGFRB/FGFR inhibitor, dovitinib. Using complementary genomic approaches we show that rearrangements, sequence mutations and DNA copy number alterations dysregulating cytokine receptor and kinase signaling are a hallmark of Ph-like ALL. These data support the screening of patients at diagnosis to identify those with Ph-like ALL, characterize the genomic lesions driving this phenotype, and to determine those that may benefit from TKI treatment. Disclosures: Hunger: Bristol-Myers Squibb: Author's children own stock in BMS, Membership on an entity's Board of Directors or advisory committees.

scholarly journals Improved Survival for Children and Adolescents With Acute Lymphoblastic Leukemia Between 1990 and 2005: A Report From the Children's Oncology Group

2012 ◽  
Vol 30 (14) ◽  
pp. 1663-1669 ◽  
Author(s):  
Stephen P. Hunger ◽  
Xiaomin Lu ◽  
Meenakshi Devidas ◽  
Bruce M. Camitta ◽  
Paul S. Gaynon ◽  
...  
Keyword(s):  
Clinical Trials ◽  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Children And Adolescents ◽  
Lymphoblastic Leukemia ◽  
Survival Rates ◽  
Population Based ◽  
Oncology Group ◽  
Clinical Covariates ◽  
The Impact

Purpose To examine population-based improvements in survival and the impact of clinical covariates on outcome among children and adolescents with acute lymphoblastic leukemia (ALL) enrolled onto Children's Oncology Group (COG) clinical trials between 1990 and 2005. Patients and Methods In total, 21,626 persons age 0 to 22 years were enrolled onto COG ALL clinical trials from 1990 to 2005, representing 55.8% of ALL cases estimated to occur among US persons younger than age 20 years during this period. This period was divided into three eras (1990-1994, 1995-1999, and 2000-2005) that included similar patient numbers to examine changes in 5- and 10-year survival over time and the relationship of those changes in survival to clinical covariates, with additional analyses of cause of death. Results Five-year survival rates increased from 83.7% in 1990-1994 to 90.4% in 2000-2005 (P < .001). Survival improved significantly in all subgroups (except for infants age ≤ 1 year), including males and females; those age 1 to 9 years, 10+ years, or 15+ years; in whites, blacks, and other races; in Hispanics, non-Hispanics, and patients of unknown ethnicity; in those with B-cell or T-cell immunophenotype; and in those with National Cancer Institute (NCI) standard- or high-risk clinical features. Survival rates for infants changed little, but death following relapse/disease progression decreased and death related to toxicity increased. Conclusion This study documents ongoing survival improvements for children and adolescents with ALL. Thirty-six percent of deaths occurred among children with NCI standard-risk features emphasizing that efforts to further improve survival must be directed at both high-risk subsets and at those children predicted to have an excellent chance for cure.

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