Leukemia- and lymphoma-associated flow cytometric, cytogenetic, and molecular genetic aberrations in healthy individuals

Most leukemia and lymphoma cases are characterized by specific flow cytometric, cytogenetic and molecular genetic aberrations, which can also be detected in healthy individuals in some cases. The authors review the literature concerning monoclonal B-cell lymphocytosis, and the occurrence of chromosomal translocations t(14;18) and t(11;14), NPM-ALK fusion gene, JAK2 V617F mutation, BCR-ABL1 fusion gene, ETV6-RUNX1(TEL-AML1), MLL-AF4 and PML-RARA fusion gene in healthy individuals. At present, we do not know the importance of these aberrations. From the authors review it is evident that this phenomenon has both theoretical and practical (diagnostic, prognostic, and therapeutic) significance. Orv. Hetil., 2012, 153, 531–540.

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The Decrease of JAK2 V617F Allele Burden in Leukemia Transformation of An Elderly Patient with Myelofibrosis.

Blood ◽

10.1182/blood.v114.22.4990.4990 ◽

2009 ◽

Vol 114 (22) ◽

pp. 4990-4990

Author(s):

Su-Jiang Zhang ◽

Jianyong Li ◽

Wei Xu

Keyword(s):

Conflicts Of Interest ◽

Molecular Genetic ◽

Myeloproliferative Neoplasm ◽

Jak2 V617f ◽

Jak2 V617f Mutation ◽

Leukemia Cells ◽

Elderly Aml ◽

Genetic Abnormalities ◽

V617f Mutation

Abstract Abstract 4990 Recently there were two different model about clone genesis of acute myeloid leukemia (AML) transformed from pre-existing JAK2 V617F positive myeloproliferative neoplasm (MPN). One model showed the leukemia cells were come from JAK2 V617F negative clone, however, the other indicated that the leukemia cells were still developed from JAK2 V617F positive clone. Here, we report a elderly AML patient who was developed from pre-existing myelofibrosis (MF) with homozygous JAK2 V617F mutation. In leuekmic transformation phase, heterozygous JAK2 V617F mutation was identified, supported the idea that the leukemia cells may be come from JAK2 V617F negative clone. Moreover, no other cytogenetic and molecular genetic abnormalities were further found. After one course of CAG regimen, complete remission was achieved. Further follow-up is still in progress. Disclosures No relevant conflicts of interest to declare.

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High Frequency of Concurrent JAK2 V617F Mutation and BCR/ABL Fussion Gene in a Cohort (18/142) of Mexican Patients with MPD

Blood ◽

10.1182/blood.v120.21.1766.1766 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1766-1766 ◽

Cited By ~ 2

Author(s):

Rosa Maria Arana Trejo ◽

Veronica A Gonzalez ◽

Israel Saldivar ◽

Beatriz SÃ°nchez ◽

Yolanda Lugo ◽

...

Keyword(s):

Bone Marrow ◽

Tyrosine Kinase ◽

Tyrosine Kinase Inhibitor ◽

High Frequency ◽

Kinase Inhibitor ◽

Fusion Gene ◽

Jak2 V617f ◽

Jak2 V617f Mutation ◽

Number Of Patients ◽

V617f Mutation

Abstract Abstract 1766 The MPD are characterized by clonal neoplastic proliferation of one or more of the myeloid lineage in the bone marrow. The BCR/ABL fusion gene is a key genetic marker for CML an the mutation JAK2 V617F is present by over 95% of patients with PV and more than 50% of patients with ET and IMF. Both JAK2 V617F and BCR/ABL result in perturbation of tyrosine kinases and their downstream signaling pathway. The coexistence of BCR/ABL fusion gene and JAK2 V617F mutation have been described in some cases. In the differential diagnosis of MPD vs CML we performed both determinations BCR/ABL fusion gene and JAK2 V617F mutation and observed high frequency of the coexistence. Material and Methods: We performed 350 JAK2V617F and 1500 BCR-ABL PCR assays in LAOH, between January 2011 and July 2012, for the patients with suspected MPD vs CML that were sent from diferents hospitals. This cases without definitive diagnosis were maked both test and the cohort was of 142 patients. Peripheral blood or bone marrow samples were included of 56 cases with CML, 48 cases of isolated thrombocytosis, 34 cases in which total blood count disclosed elevation of ≥2 myeloid cell types and 4 cases with PV diagnosis. RNA and DNA were extracted using Trizol® reagent and Quiagen® commercial kit, the RT-PCR from b3a2/b2a2 BCR-ABL transcript was performed according to protocols standarized. For JAK2 V617F we using the allele-specific PCR method and the restiction enzyme method with BsaXI enzyme wich restriction site is present in exon 14 of JAK2. To validate the specificity of our result we sequenced the PCR products in 10 cases and two normal controls. Results. Of the 142 patients evaluated for both JAK2 V617F and BCR-ABL fusion gene, 18 patients were positive for both (12.7%). Ten with CML de novo, seven cases with ET, and one case in follow-up with imatinib. The average age of these JAK2 V617F mutation and BCR-ABL positive patients (n=18) was 63.5 (ranging from 45–82) and in patients JAK2 V617F mutation and BCR-ABL negative (n=124) were 58 years old (rank 18–85). The patient with CML in therapy with imatinib by one year, was present increase WBC; and his molecular analysis for JAK2 V617F was positive and also BCR/ABL+. The patient′s treatment regimen was modified, with initiation of hydroxyurea and imatinib; the patient achieved complete hematological response. Discussion. The others cases reports in the literature are most often a JAK2 V617F+ MPN developed in the setting of a previously diagnosed CML undergoing treatment with a tyrosine kinase inhibitor, or alternately, CML developed months to years after patients had been diagnosed with a JAK2 V617F+ MPN. In this report the most of our cases the BCR-ABL and JAK2 V617F positive cells were detected before the treatment. This studies have suggested two possibilities to account for expression of BCR-ABL and JAK2 V617F anomalies concomitant in a single patient. The first is that there are two clones each having BCR-ABL and JAK2 V617F mutation and another possibility is a single clone concurrently possesses both BCR-ABL and JAK2 V617F. It remains to be seen whether or not this patients would benefit from combined treatment with tyrosine kinase inhibitor and interferon or hydroxyurea. The clinical following up of this patients help us to response this questions. We have the highest frequency of concurrent JAK2 V617F mutation and BCR-ABL fusion gene reported, but is neccessary determined the true incidence in a greater number of patients with CML and MPD. Disclosures: No relevant conflicts of interest to declare.

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Analysis of genetic aberrations in pediatric high-grade gliomas

Advances in molecular oncology ◽

10.17650/2313-805x-2020-7-3-37-47 ◽

2020 ◽

Vol 7 (3) ◽

pp. 37-47

Author(s):

M. A. Zaytseva ◽

A. P. Shekhtman ◽

L. I. Papusha ◽

E. F. Valiakhmetova ◽

L. A. Yasko ◽

...

Keyword(s):

Fusion Gene ◽

Clinical Care ◽

Molecular Genetic ◽

Driver Mutations ◽

High Grade ◽

Missense Mutations ◽

Genetic Aberrations ◽

Genomic Landscape ◽

Wide Range ◽

High Grade Gliomas

Background. High-grade gliomas are characterized by a wide range of genetic abnormalities. The heterogeneous genomic landscape of pediatric high-grade gliomas allows identifying distinct subgroups of the disease in children and young adults. Most importantly, these subgroups differ by clinical course and prognosis, as well as treatment response to standard therapy.Objective: to assess the profile of molecular genetic markers of high-grade gliomas in children.Materials and methods. In the current study, we examine the frequency of H3F3A, Hist1H3B, BRAF, IDH1 / 2 mutations, the copy number alterations of CDKN2A / 2B genes and the expression of ETV6‑NTRK3 fusion gene in a cohort of 53 pediatric high-grade gliomas.Results. Driver mutations and CDKN2A / 2B deletions were observed in 24 (45 %) and 15 (28 %) of 53 tumors, respectively. Overall, the studied high-grade gliomas harbored 41 genetic aberrations including 24 (58.5 %) somatic missense mutations, 1 (2.4 %) genetic variant of unknown clinical significance, 1 (2.4 %) oncogenic fusion gene and 15 (36.6 %) deletions of the tumor suppressor genes.Conclusion. These findings point to the importance of molecular profiling of tumors for the optimal clinical care and development of new approaches to treatment aimed at molecular targets for personalized anticancer therapies.

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A NUP98-HOXD13 Fusion Gene Impairs B Lymphocyte Development and Function In a Mouse Model for Myelodysplastic Syndrome

Blood ◽

10.1182/blood.v116.21.279.279 ◽

2010 ◽

Vol 116 (21) ◽

pp. 279-279

Author(s):

Abdul Gafoor A. Puthiyaveetil ◽

Bettina Heid ◽

David L. Caudell

Keyword(s):

B Cells ◽

Transgenic Mice ◽

B Cell ◽

B Lymphocytes ◽

Marginal Zone ◽

Fusion Gene ◽

Flow Cytometric Analysis ◽

Flow Cytometric ◽

Cell Fractions ◽

And Function

Abstract Abstract 279 Transgenic mice which express the fusion gene NUP98-HOXD13 (NHD13) have been shown to develop characteristic features of Myelodysplastic syndrome (MDS) including impaired hematopoietic differentiation and peripheral blood cytopenias in the presence of normocellular or hypercellular bone marrow (BM). It is evident that B-cells play a role in the progression of MDS by immune modulation or as direct targets of mutations resulting in ALL, or as cells that influence the BM microenvironment in which a neoplastic myeloid clone evolves. Choi and colleagues suggested a block in differentiation during the early development of B lymphocytes in the BM of NHD13 mice leading to lymphopenia consistent with the observation in some MDS patients. In this study, we sought to further delineate the role of NHD13 on B lymphocytes which escaped the initial differentiation block in the BM. We hypothesized that NHD13 impairs maturation and function of IgM+ B lymphocytes contributing to immunodeficiency. To study this, we performed blood smear examination, Complete Blood Counts (CBC), quantitative ELISA for antibody concentrations, and flow cytometric analysis of B cell fractions from the BM and spleen in 8–12 week-old transgenic and wild type (WT) mice. CBCs revealed significant lymphopenia and ELISA showed higher IgM concentrations (n=10, p<0.001), reduced levels of IgG1 (n=10, p<0.05) and IgE (n=10, p<0.01). The IgG2a, IgG2b, and IgG3 antibody levels were comparable to WT counterparts. Flow cytometric analysis of BM and splenic B cell fractions revealed reduced numbers of B cells in Hardy fractions D and F (n=10, p<0.01) indicative of impaired differentiation prior to these stages; splenic fractions in NHD13 mice were comparable to WT controls. Next, to assess the peripheral maturation and functional efficiency of B lymphocytes in the context of a comprehensive immune stimulation, a cohort of five WT and five preclinical transgenic mice were injected with 100 μ g dinitrophenol (DNP) followed by a booster dose on day 21. Mice were euthanized on day 28 and whole blood, spleen, lymphnodes and BM were harvested. CBC evaluation revealed significant lymphopenia in NHD13 mice (n=5, p<0.001). Quantitative ELISA for DNP specific antibodies showed comparable levels of serum IgM and significantly reduced levels of serum IgG1 (n=5, p<0.001), IgG2a (n=5, p<0.001), IgG2b (n=5, p<0.01), IgG3 (n=5, p<0.001) and IgE (n=5, p<0.01). Flow cytometric analysis of peripheral blood showed reduced numbers of B220+ IgM+ B cells (n=5, p<0.01), but comparable percentages of CD4+ and CD8+ T-cells. Detailed flow cytometric analysis of B-cell fractions in the BM and spleen of DNP-stimulated mice revealed a reduction in subpopulations of B lymphocytes. The earliest B cell lineage population, Pre-Pro B, was comparable to the WT controls. Hardy Pro B fraction B (n=5, p<0.001) and Pre B fractions E (n=5, p<0.01) and F (n=5, p<0.01) from BM of stimulated mice were significantly reduced in contrast to fractions C and C', which were higher (n=5, p<0.05 and p<0.001 respectively), indicative of cell growth arrest at these stages. Flow cytometry of splenic B-cell fractions from the DNP-stimulated mice showed significantly lower Transitional 1 (n=5, p<0.01), Follicular (n=5, p<0.05) and Marginal Zone (n=5, p<0.001) populations upon antigenic stimulation suggestive of defective clonal expansion of IgM+ cells even after escaping the block in the BM. Histopathology of the spleen revealed smaller lymphoid follicles with poorly developed mantle and marginal zone regions in the transgenic mice when compared to WT controls, consistent with the flow cytometric data. This study indicates that when NHD13 mice are immunologically challenged, B lymphocytes undergo impaired differentiation in the BM and maturation in the spleen, as well as reduced antibody class switching and subsequently lower antibody production. Analysis of B cell subsets during development and specific IgG/IgE antibody production, suggest that the NHD13 transgene might impair VDJ gene recombination and class switch recombination that are critical during these phases of B cell development. Disclosures: No relevant conflicts of interest to declare.

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Myeloproliferative Neoplasm with BCR-JAK2 Fusion Gene As the Result of t(9;22)(p24,11.2) in a Brazilian Patient

Blood ◽

10.1182/blood.v120.21.4808.4808 ◽

2012 ◽

Vol 120 (21) ◽

pp. 4808-4808

Author(s):

Daniela Ferreira Dias ◽

Marcelo Bellesso ◽

Rodrigo Santucci ◽

Renata Campos Elias ◽

Veronica Ramos Oliveira ◽

...

Keyword(s):

Tyrosine Kinase ◽

Tyrosine Kinase Inhibitors ◽

Kinase Inhibitors ◽

Fusion Gene ◽

Jak2 V617f ◽

Jak2 V617f Mutation ◽

Rt Pcr ◽

Hematologic Response ◽

V617f Mutation

Abstract Abstract 4808 Introduction Chronic Myeloid Leukemia (CML) is the most well described disease result of t(9;22)(q34,q11.2). This chromosomal rearrangement leads to well-know BCR-ABL fusion that promotes tyrosine kinase activity. There are others oncogenic BCR fusion found such as PDGFRA (4q12), FGFR1(8p12) that causes myeloproliferative disorders (MD). JAK2 gene is one of the 4 genes members of JAK family. The JAK2 V617F mutation which results from a G –>T transversion at nucleotide 1849 in exon 14 of the JAK2 gene, the consequence of which is substitution of valine by phenylalanine at codon 617 is associated with MD and it is a major diagnosis criterion for Primary Myeloficrosis, Polycythaemia vera and Essential thrombocytemia. There are described a lot chromosomal translocations involving the JAK2 locus. We report an extremely rare case with BCR-JAK2 fusion gene as the result of t(9;22)(p24,q11.2) for the first time in Brazilian people, and it is the 6thcase all of the world. Case Report In April 2010, a 54 years old male patient presented fatigue, abdominal pain and splenomegaly. A blood count revealed leukocytosis 93.380/mm3 with a predominance of neutrophils and left shift. Conventional cytogenetic analysis was performed and it was evidenced 46,XY, t(9;22)(p24;q11.2) in 90% metaphases examined, due to expected association it was promoted BCR-ABL1 fusion gene and it was not detected by using RT-PCR. He was treated with imatinib 400mg/day because the involvement of BCR gene. After three months he presented weight loss, progressive splenomegaly without hematologic response and it was modified to Dasatinib 150mg/day plus hydroxyureia 3g/day. In August 2011, due to not hematologic response, it was stopped Dasatinib treatment and nowadays patient has been treating with hydroxyureia 1.5g/day. His last follow up in May 2012, blood count was abnormal Hb 16.8g/dl leukocytes 7730/mm3 and low platelets count 32.000/mm3. The differential count showed 65.3% segmented granulocytes, 13.6% eosinophlis, 1.6% basophil, 2.6% monocytes, 16.9% lymphocytes. It was repeated conventional Karyotyping and it was evidenced 46,XY, t(9;22)(p24;q11.2) in all of metaphases examined. The presence of BCR-ABL rearrangement was excluded by using the fluorescence in situ hybridization (FISH) using a BCR-ABL probe. In addition, it was not evidenced FIP1L1-PDGFRa fusion gene and JAK2 V617F mutation by using RT-PCR. Discussion We have described a male patient with MD with t(9;22)(p24;q11.2) wich leads to the BCR-JAK2 fusion and it was not evidenced BCR-ABL1, FIP1L1-PDGFRa fusion genes and JAK2 V617F mutation by using RT-PCR. Moreover, patient has not been achieved hematologic response with tyrosine kinase inhibitors: imatinib and dasatinib. In the five cases reported three presented MD, one Acute Myeloid Leukemia and one Acute Lymphoblastic Leukemia. Only in one case report it was prescribed imatinib and the patient lost the follow up (Table1). The BCR-JAK2 fusion protein contain the coiled-coil dimerization domain of BCR and the protein tyrosine Kinase domain (JH1) of JAK2. It was not possible to define what would be the best therapy, because tyrosine kinase inhibitors may not be effective to the BCR-JAK2 fusion. Maybe in MD presentation, we could return to pre- tyrosine kinase inhibitors era based on treatments with hydroxyureia, subcutaneous cytarabine and interferon for patients that were not potential candidates for allogeneic transplant. Disclosures: No relevant conflicts of interest to declare.

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