Identification of a Novel, Oncogenic and Targetable TPR::ABL2 Fusion Gene in T-Cell Acute Lymphoblastic Leukaemia.

Classifying subtypes of acute lymphoblastic leukemia using silhouette statistics and genetic algorithms

Expression Profile of Selected HOX Genes in Different Childhood ALL Subtypes

The authors conducted a flow cytometry immunophenotyping study in patients with acute lymphoblastic leukemia (ALL) from Natal, Rio Grande do Norte, Brazil. The patients (n = 126) were newly diagnosed using a panel of monoclonal antibodies: CD1a, CD2, CD3, CD4, CD7, CD8, CD10, CD13, CD33, CD14, CD19, CD22, CD79a, CD117, CD34, anti-IgM, anti-TdT, anti-HLA-Dr, and anti-human kappa and lambda light chains. Additional data, such as patients' age and gender, clinical and laboratory findings such as presence of tumor masses, lymphadenopathy, hepatomegaly, splenomegaly, leukemic infiltration in the central nervous system (CNS) were also investigated. Results showed that 56.7% of the cases were B-lineage ALL and 55% were T-cell ALL. Also, we found that males were more affected by the disease, regardless of immunological classification. The correlation between age and immunological subtypes showed that the B-lineage ALL occurred more frequently in patients aged under 15 while the T-cell ALL subtype was more frequent in adults. Immunophenotypic profiles and morphological subtypes showed a direct correlation between L3 subtype and B-lineage ALL, while L1 and L2 subtypes correlated more often with B-cell lineage and T-cell ALL, respectively. Correlation analysis between immunophenotypic and clinical profiles showed that T-cell ALL was more associated with a higher incidence of lymphadenopathy, hepatomegaly, splenomegaly and CNS leukemic infiltration, also showing a greater blast cell count in peripheral blood than the other subgroups. The presented data suggest that immunophenotyping is an important method in the diagnosis, monitoring and prognostic assessment in determining the pathological mechanisms of evolution of ALL.

The application of global gene expression profiling allows to obtain detailed molecular fingerprints of underlying gene expression in any cell of interest. In this work gene expression profiles were generated from a comprehensive cohort of leukemia patients and healthy donors referred to and diagnosed in the Laboratory for Leukemia Diagnostics, Munich, Germany, which is a nation-wide reference center for the diagnosis of hematologic malignancies. Thoroughly characterized clinical samples were analyzed by high-density microarrays interrogating the expression status of more than 33,000 transcripts. In one specific aspect of this work the potential application of gene expression signatures for the prediction and classification of specific leukemia subtypes was assessed. Today the diagnosis and subclassification of leukemias is based on a controlled application of various techniques including cytomorphology, cytogenetics, fluorescence in situ hybridization, multiparameter flow cytometry, and PCR-based methods. The diagnostic procedure is performed according to a specific algorithm, but is time-consuming, cost-intensive, and requires expert knowledge. Based on a very low number of candidate genes it is demonstrated in this work that prognostically relevant acute leukemia subtypes can be classified using microarray technology. Moreover, in an expanded analysis including 937 patient samples representing 12 distinct clinically relevant acute and chronic leukemia subtypes and healthy, non-leukemia bone marrow specimens a diagnostic prediction accuracy of ~95% was achieved. Thus, given these results it can be postulated that the occurring patterns in gene expression would be so robust that they would allow to predict the leukemia subtype using global gene expression profiling technology. This finding is further substantiated through the demonstration that reported differentially expressed genes from the literature, namely pediatric gene expression signatures representing various acute lymphoblastic leukemia (ALL) subtypes, can be used to independently predict the corresponding adult ALL subtypes. Furthermore, it could be demonstrated that microarrays both confirm and reproduce data from standard diagnostic procedures, but also provide very robust results. Parameters such as partial RNA degradation, shipment time of the samples, varying periods of storage of the samples, or target preparations at different time points from either bone marrow or peripheral blood specimens by different operators did not dramatically influence the diagnostic gene expression signatures. In another major aspect of this work gene expression signatures were examined in detail to obtain new insights into the underlying biology of acute promyelocytic leukemia (APL) and t(11q23)/MLL leukemias. In APL, microarrays led to a deeper understanding of morphological and clinical characteristics. Firstly, genes which have a functional relevance in blood coagulation were found to be differentially expressed when APL was compared to other acute myeloid leukemia (AML) subtypes. Secondly, a supervised pairwise comparison between the two different APL phenotypes, M3 and its variant M3v, for the first time revealed differentially expressed genes encoding for biological functions and pathways such as granulation and maturation. With respect to 11q23 leukemias it could be demonstrated that leukemias with rearrangements of the MLL gene are characterized by a common specific gene expression signature. Additionally, in unsupervised and supervised data analysis algorithms ALL and AML cases with t(11q23)/MLL segregated according to the lineage, i.e., myeloid or lymphoid, respectively. This segregation could be explained by a highly differing transcriptional program. Through the use of biological network analyses essential regulators of early B cell development, PAX5 and EBF, were shown to be associated with a clear B-lineage commitment in lymphoblastic t(11q23)/MLL leukemias. Also, the influence of the different MLL translocation partners on the transcriptional program was directly assessed. But interestingly, gene expression profiles did not reveal a clear distinct pattern associated with one of the analyzed partner genes. Taken together, the identified molecular expression pattern of MLL fusion gene samples and biological networks revealed new insights into the aberrant transcriptional program in t(11q23)/MLL leukemias. In addition, a series of analyses was targeted to obtain new insights into the underlying biology in heterogeneous B-lineage leukemias not positive for BCR/ABL or MLL gene rearrangements. It could be demonstrated that the genetically more heterogeneous precursor B-ALL samples intercalate with BCR/ABL-positive cases, but their profiles were clearly distinct from T-ALL and t(11q23)/MLL cases. In conclusion, various unsupervised and supervised data analysis strategies demonstrated that defined leukemia subtypes can be characterized on the basis of distinct gene expression signatures. Specific gene expression patterns reproduced the taxonomy of this hematologic malignancy, provided new insights into different disease subtypes, and identified critical pathway components that might be considered for future therapeutic intervention. Based on these results it is now further possible to develop a one-step diagnostic approach for the diagnosis of leukemias using a customized microarray.

Flow Cytometry Immunophemotyping Evaluation in the Diagnosis of Acute Lymphoblastic Leukemia in Patients from Rio Grande Do Norte State, Brazil

We evaluate clinical significance of recently identified subtypes of acute lymphoblastic leukemia (ALL) in 598 children treated with minimal residual disease (MRD)-directed therapy. Among the 16 B-ALL and 8 T-ALL subtypes identified by next generation sequencing, ETV6-RUNX1, high-hyperdiploid and DUX4-rearranged B-ALL had the best five-year event-free survival rates (95% to 98.4%); TCF3-PBX1, PAX5alt, T-cell, ETP, iAMP21, and hypodiploid ALL intermediate rates (80.0% to 88.2%); and BCR-ABL1, BCR-ABL1-like and ETV6-RUNX1-like and KMT2A-rearranged ALL the worst rates (64.1% to 76.2%). All but three of the 142 patients with day-8 blood MRD <0.01% remained in remission. Among new subtypes, intensified therapy based on day-15 MRD≥1% improved outcome of DUX4-rearranged, BCR-ABL1-like, and ZNF384-rearranged ALL, and achievement of day-42 MRD<0.01% did not preclude relapse of PAX5alt, MEF2D-rearranged and ETV6-RUNX1-like ALL. Thus, new subtypes including DUX4-rearranged, PAX5alt, BCR-ABL1-like, ETV6-RUNX1-like, MEF2D-rearranged and ZNF384-rearranged ALL have important prognostic and therapeutic implications.

Molecular Characterization of a Novel Subtype of Pediatric Acute Lymphoblastic Leukemia.

Role of cranial boost in addition to total body irradiation in the treatment of high risk acute lymphoblastic leukemia

Acute lymphoblastic leukemia (ALL) remains an important cause of morbidity in children and adults. In this article, we highlight advances in the genetics and therapy of three key subtypes of ALL: T-cell ALL, BCR-ABL1 (Philadelphia [Ph] chromosone-positive), and Ph-like ALL. T-ALL is an aggressive disease that accounts for about 15% and 25% of ALL among pediatric and adult cohorts, respectively, and exhibits a multistep nature of cancer initiation and progression. The integration of cytogenetics, molecular biology, and immunophenotype analyses has led to the identification of defined T-ALL subgroups, such as early T-cell precursor ALL and novel lesions with a prognostic role, for which specific inhibitors are being developed. Ph-positive ALL was historically regarded as a subtype of ALL with a poor prognosis, and allogeneic stem cell transplant was recommended for all patients who could undergo this procedure. The deep complete responses seen with combination tyrosine kinase inhibitors (TKIs) and chemotherapy in Ph-positive ALL, and the reports of long-term survival among some patients not undergoing allogeneic stem cell transplant, has raised the question of whether there is a subset of patients who could be cured without this intervention. Ph-like ALL is a subtype of B-progenitor ALL common among older children and adults and associated with a diverse range of genetic alterations that activate kinase signaling. Ph-like ALL is also associated with poor outcome, for which precision medicine trials identifying kinase alterations and testing TKI therapy are being developed.

Genetic alterations that underlie the pathogenesis of individual leukemia subtypes have important prognostic implications in acute lymphoblastic leukemia (ALL). Heretofore, specific genetic abnormalities with prognostic relevance were identified mainly by cytogenetic analysis or reverse-transcriptase polymerase-chain-reaction, and were largely limited to B-lineage ALL cases. Moreover, there is heterogeneity among specific genetic subtypes of ALL and treatment has over-riding impact on the prognostic information conveyed by the genetic abnormalities. In Philadelphia chromosome-positive (Ph+) ALL, an age of 1 to 9 years and a low initial leukocyte count confer a favorable prognosis. In each category of 11q23/MLL abnormality, infants less than 1 year of age have a worse treatment outcome than do older patients. Allogeneic transplantation with HLA-matched related donor improved outcome for patients with Ph+ ALL but not those with the t(4;11) andMLL-AF4 fusion. Using oligonucleotide microarrays, T-cell ALL can now be classified into four major subtypes with prognostic significance:MLL-ENL andHOX11 clusters with favorable prognosis andTAL1 andLYL1 clusters with inferior outcome. In our recent study of 327 ALL cases, distinct expression profiles could not only accurately identify each of the known prognostically important leukemia subtypes, but also individual cases that would eventually relapse or develop therapy-related acute myeloid leukemia. We have also identified a novel ALL subgroup with unique expression profile. Examination of the genes comprising the different expression signatures provided important insights into the biologic differences between the leukemia subgroups. The importance of host pharmacogenetics has also been recognized. Thus, genetic polymorphisms of certain drug-metabolizing enzymes, drug transporters or drug receptors have been linked with treatment efficacy and toxicity, as well as host susceptibility to the development of de novo leukemia or therapy-related second cancers. The best-characterized genes (e.g. thiopurine methyltransferase, cytochrome P450, glutathione transferase, methylenetetrahydrofolate reductase and NAD(P)H: quinone oxidoreductase) will be discussed. Ultimately, patients will be treated with individualized therapy based on leukemic cell gene expression profile and host pharmacogenetics.

A subtype of childhood acute lymphoblastic leukaemia with poor treatment outcome: a genome-wide classification study.

Background: Immunophenotypic analysis of leukemic cells by multicolor flow cytometry using different monoclonal antibodies labeled to various flourochromes, is an important, precise and rapid investigation for diagnosis, classification, prognosis prediction, and minimal residual disease detection in acute lymphoblastic leukemia (ALL). Objectives: The aims of study were to study the expression of CD45, CD34, CD10, human leukocyte antigen-DR (HLA-DR) in B and T-ALL among 114 Iraqi patients and compare findings with other reports worldwide. Patients, Materials and Methods: A retrospective cross-sectional study was conducted on 114 ALL patients of various age groups from different hematology centers in Baghdad who were referred to Flowcytometry Department at the Bone Marrow Transplantation Center/Private Nursing Home Hospital/Medical City for immunophenotypic classification using multi-color flow cytometry from the January 1, 2016, to the August 31, 2016. Results: Out of 114 patients, 71 patients were pediatric patients and 43 were adults. Eighty-three patients were classified as B-cell ALL and 31 patients as T-cell ALL. There was significant association between male gender, high white blood cells count and T-cell ALL subtype. Negative-dim-moderate CD45, positive CD34, and positive HLA-DR expressions were significantly associated with B-cell ALL. Common ALL antigen was seen in 86.7% of the B-cell ALL patients and was accompanied cTdT expression. Aberrant myeloid antigens were observed in 22.9% of B-cell ALL patients and in 35.5% of T-cell ALL. About 4.8% of the patients with B-cell ALL had aberrant T-linage antigens while 6.5% of the patients with T-cell ALL expressed aberrant B-cell lineage antigens. Conclusion: Immunophenotypic expression of ALL cells among Iraqi patients is to somewhat in accordance with various researches worldwide making immunophenotyping flow cytometry a crucial appliance in diagnosis, classification, risk stratification, and minimal residual disease detection in ALL.

Acute lymphoblastic leukemia withTEL-AML1fusion has lower expression of genes involved in purine metabolism and lower de novo purine synthesis

Patterns of Neuropathy in ALL Patients Treated with Vincristine Based Regimens with and without Tyrosine Kinase Inhibitors

Although substantial advances in the identification of cytogenomic subtypes of childhood acute lymphoblastic leukemia (ALL) have been made in recent decades, epidemiologic research characterizing the etiologic heterogeneity of ALL by subtype has not kept pace. The purpose of this review is to summarize the current literature concerning subtype-specific epidemiologic risk factor associations with ALL subtype defined by immunophenotype (e.g., B-cell vs. T-cell) and cytogenomics (including gross chromosomal events characterized by recurring numerical and structural abnormalities, along with cryptic balanced rearrangements, and focal gene deletions). In case-control analyses investigating nongenetic risk factors, home paint exposure is associated with hyperdiploid, MLL-rearranged, and ETV6-RUNX1 subtypes, yet there are few differences in risk factor associations between T- and B-ALL. Although the association between maternal smoking and ALL overall has been null, maternal smoking is associated with an increasing number of gene deletions among cases. GWAS-identified variants in ARID5B have been the most extensively studied and are strongly associated with hyperdiploid B-ALL. GATA3 single nucleotide variant rs3824662 shows a strong association with Ph-like ALL (OR = 3.14). However, there have been relatively few population-based studies of adequate sample size to uncover risk factors that may define etiologic heterogeneity between and within the currently defined cytogenomic ALL subtypes.