Characterization of 14 NUP214-ABL1 Fusions in T-Cell Acute Lymphoblastic Leukaemia (T-ALL) Exhibits a Genomic Heterogeneity.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2075-2075
Author(s):  
Carlos Graux ◽  
Marina Lafage ◽  
Nicole Dastugue ◽  
Francine Mugneret ◽  
Roland Berger ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Kinase ◽  
Cultured Cells ◽  
Outcome Data ◽  
Tyrosine Kinase Domain ◽  
Fish Analysis ◽  
Rt Pcr ◽  
Genomic Amplification ◽  
Chromosomal Alterations ◽  
Male Predominance

Abstract The recently described episomal amplification of the NUP214-ABL1 fusion in 6% of T-ALL leads to the expression of a constitutively activated chimeric tyrosine kinase sensitive to imatinib. We collected additional cases in order to better characterize this new entity with respect to genetic presentations and clinical course. We collected 14 new NUP214-ABL1 positive cases by FISH (LSI-BCR/ABL1 ES, Vysis and ABL1 break-apart home-made probes) or RT-PCR (fusion transcript) screening. FISH analysis detected episomal amplification of NUP214-ABL1 in 11 patients with a highly variable number of nuclei with amplification (<1% to 90%). Interestingly, one case showed a higher percentage of nuclei with amplification when using frozen non cultured cells rather than cultured fixed cells (10% vs 1%) suggesting loss of episomes during culture. FISH showed also intrachromosomal amplification of NUP214-ABL1 in two cases: one presented a HSR at the original 9q34 site without detectable episomes, the other associated HSR (probably on a chromosome 10), episomes (<1% of nuclei) and 9q34 chromosomal insertions including NUP214 and ABL3′ that encode the tyrosine kinase domain but not ABL5′, on variable chromosomes including 14p (33%). One NUP214-ABL1 RT-PCR positive case did not show any FISH aberration. Median age: 16 y (3–45) with a male predominance (10:4). There were no T-cell lymphoblastic lymphoma. Immunophenotype (EGIL): mature (n=2), cortical (n=6) or pre-T (n=4). Karyotype: structural chromosomal alterations in 8 patients (including 4 with 10q24/HOX11 rearrangements), only numerical chromosomal alterations in 4 (including 2 with + 8), normal in 1, failed in 1. All samples with available information (10/14) showed a HOX11 or HOX11L2 abnormality. Among the 13 cases with available outcome data, we observed 5 early relapses, including both patients with NUP214-ABL1 HSR, and 1 refractory ALL. These observations emphasize the interest of combining both (quantitative) RT-PCR and FISH for the screening and characterisation of NUP214-ABL1 fusion and amplification, demonstrate the coexistence of different NUP214-ABL1 genomic presentations in one patient (episomal amplification, 9q34 insertions, HSR) compatible with the model in which genomic amplification start with episome formation in order to create the NUP214-ABL1 fusion followed by their amplification and optional secondary reintegration, confirm occurrence of NUP214-ABL1 in T-ALL with HOX11 and HOX11L2 involvement, raise the question of the rather worse prognosis for cases with intrachromosomal amplification as previously suggested, raise the signification of minor NUP214-ABL1 clones and variable genomic presentations in the leukemogenesis of this subgroup of T-ALL that could potentially benefit from imatimib. ° on behalf of the GFCH (Groupe Francophone de Cytogénétique Hématologique) and the BCGHO (Belgian Cytogenetic Group for Hematology and Oncology).

Fusion of SFQP to ABL in a Patient with a T(1;9)(p34;q34) and Acute Lymphocytic Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4444-4444
Author(s):  
Nicholas C.P. Cross ◽  
Andrew J. Chase ◽  
Milton Drachenberg ◽  
W. Mark Roberts ◽  
Jerry Z. Finklestein ◽  
...  
Keyword(s):  
Complete Remission ◽  
Fusion Gene ◽  
Chimeric Protein ◽  
Sh2 Domain ◽  
Lymphocytic Leukemia ◽  
Tyrosine Kinase Domain ◽  
Fish Analysis ◽  
Fusion Partner ◽  
Rt Pcr ◽  
Polypyrimidine Tract Binding Protein

Abstract We have investigated a child who presented with pre-B ALL and an acquired t(1;9)(p34;q34). BCR-ABL was not detected by RT-PCR or FISH analysis, however FISH did indicate that the ABL gene at 9q34 was disrupted. To identify the putative partner locus in this case, a modified 5′RACE strategy was employed that selected against normal ABL transcripts. Several clones were recovered in which ABL was fused to SFPQ (also known as PSF), a gene mapping to 1p34 that encodes a polypyrimidine tract-binding protein-associated splicing previously identified as a fusion partner of the helix-loop-helix transcription factor TFE3 in papillary renal cell carcinomas. Both SFPQ-ABL and reciprocal ABL-SFPQ transcripts were detectable by RT-PCR, and disruption of these two genes was further confirmed by amplification and sequencing of the forward genomic breakpoint. SFPQ-ABL, the likely oncogenic product, is predicted to encode a protein that retains the coiled coil domain of SFPQ and the entire tyrosine kinase domain and C-terminal sequences of ABL. The breakpoint in ABL was downstream of that seen for other ABL fusion genes and the chimeric protein is predicted to lack the ABL-encoded SH3 domain and part of the SH2 domain. The patient was treated according to the Children’s Cancer Group Protocol 1961 and subsequently received augmented BFM therapy with doxorubicin and double delayed intensification. He achieved complete remission but suffered extramedullary testicular relapse at 4.5 years. Following orchiectomy and intensive chemotherapy he remains in complete remission more than 6 years after diagnosis. We conclude that SFPQ-ABL is a novel fusion gene associated with ALL. Although the patient here responded to conventional chemotherapy, SFPQ-ABL is likely to be sensitive to imatinib and use of this agent might be considered in further cases.

Activating FLT3 Mutations in CD4+/CD8− Pediatric T-Cell Acute Lymphoblastic Leukemias.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1465-1465
Author(s):  
Pieter Van Vlierberghe ◽  
Jules P.P. Meijerink ◽  
Ronald W. Stam ◽  
Wendy van der Smissen ◽  
Elisabeth R. van Wering ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Kinase ◽  
Point Mutations ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Activating Mutations ◽  
Activated State ◽  
Flt3 Mutations ◽  
Genetic Aberration ◽  
High Level

Abstract Activating mutations in the FMS-like tyrosine kinase 3 gene (FLT3) are the most common genetic aberration in acute myeloid leukemia (AML). Internal tandem duplications (ITD) in the juxtamembrane (JM) domain, or point mutations (PM) in the activation loop of the tyrosine kinase domain lead to a constitutive activated state of the FLT3 tyrosine kinase. Recently, FLT3 mutations were identified in a cohort of 69 adult T-ALL patients, showing that this genetic abnormality is not only restricted to myeloid leukemias. To validate the incidence of FLT3 mutations in pediatric T-ALL and investigate its relation to outcome and other clinical and immunophenotypical parameters, we screened 72 diagnostic pediatric T-ALL samples. FLT3/ITD mutations were identified in 2/72 pediatric T-ALLs (2.7%), whereas 0/72 showed point mutations in the kinase domain. Immunophenotypic analysis revealed a similar profile for both FLT3 mutated patient samples, i.e. TdT+, CD2+, CD5+, CD7+, CD4+/CD8−, cytoplasmic CD3+, surface CD3− and CD10−. Although representing early T-cell differentiation stages for both patient samples, these cases seem to have a more advanced immunophenotype compared to the FLT3 mutated adult T-ALL cases, previously described (CD34+, CD4−/CD8−). Both FLT3 mutated patients showed high level LYL1 and LMO2 expression. In addition, both pediatric samples contained a HOX11L2 translocation, which was not present in the FLT3 mutated adult T-ALL cases. The first FLT3 mutated patient suffered a relapse 13 months after initial diagnosis, whereas the other is still in continued complete remission for 61+ months. Interestingly, the relapse material showed no FLT3/ITD mutation, indicating that the FLT3 mutated T-ALL subclone seems to be effectively eradicated by current chemotherapy. These data suggest that the application of FLT3 inhibitors for FLT3-mutated T-ALLs, as recently suggested in literature, may not further improve treatment outcome in pediatric T-ALL.

scholarly journals Efficient CD28 signalling leads to increases in the kinase activities of the TEC family tyrosine kinase EMT/ITK/TSK and the SRC family tyrosine kinase LCK

1998 ◽  
Vol 330 (3) ◽  
pp. 1123-1128 ◽  
Author(s):  
Spencer GIBSON ◽  
Ken TRUITT ◽  
Yiling LU ◽  
Ruth LAPUSHIN ◽  
Humera KHAN ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
T Cell Activation ◽  
Tyrosine Kinases ◽  
Cell Activation ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain

Optimal T cell activation requires crosslinking of the T cell receptor (TCR) concurrently with an accessory receptor, most efficiently CD28. Crosslinking of CD28 leads to increased interleukin 2 (IL2) production, inhibition of anergy and prevention of programmed cell death. Crosslinking of CD28 leads to rapid increases in tyrosine phosphorylation of specific intracellular substrates including CD28 itself. Since CD28 does not encode an intrinsic tyrosine kinase domain, CD28 must activate an intracellular tyrosine kinase(s). Indeed, crosslinking of CD28 increases the activity of the intracellular tyrosine kinases EMT/ITK and LCK. The phosphatidylinositol 3-kinase (PI3K) and GRB2 binding site in CD28 is dispensable for optimal IL2 production in Jurkat T cells. We demonstrate herein that murine Y170 (equivalent to human Y173) in CD28 is also dispensable for activation of the SRC family tyrosine kinase LCK and the TEC family tyrosine kinase EMT/ITK. In contrast, the distal three tyrosines in CD28 are required for optimal IL2 production as well as for optimal activation of the LCK and EMT/ITK tyrosine kinases. The distal three tyrosines of CD28, however, are not required for recruitment of PI3K to CD28. Furthermore, PI3K is recruited to CD28 in JCaM1 cells which lack LCK and in which EMT/ITK is not activated by ligation of CD28. Thus optimal activation of LCK or EMT/ITK is not obligatory for recruitment of PI3K to CD28 and thus is also not required for tyrosine phosphorylation of the YMNM motif in CD28. Taken together the data indicate that the distal three tyrosines in CD28 are integral to the activation of LCK and EMT/ITK and for subsequent IL2 production.

Ph’-Positive (Ph’+) Chronic Myeloid Leukemia (CML) Presenting as Ph’+ T-Lymphoblastic Lymphoma (LL) Resistant to High-Dose Chemotherapy and Gleevec.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4869-4869
Author(s):  
Lorenzo Falchi ◽  
Giorgia Desantis ◽  
Fabrizio Liberati ◽  
Roberta Pace ◽  
Maria G. Morandi ◽  
...  
Keyword(s):  
T Cell ◽  
Soft Tissues ◽  
Blast Crisis ◽  
Chronic Phase ◽  
Malignant Cell ◽  
Whole Body ◽  
High Dose ◽  
Fish Analysis ◽  
Rt Pcr ◽  
Entire Treatment Period

Abstract BACKGROUND Extramedullary blast crisis (BC) of Ph’+ CML is infrequent and commonly affects bone, lymphoid tissue, skin and soft tissues and central or peripheral nervous system. Most of nodal lymphoid tumors occuring in the setting of CML derive from T-cell precursors and represent the evolution of CML to a lymphoid nodal BC. CASE HISTORY In February 2004, a 52 year-old patient underwent diagnostic wide biopsy of a nasopharingeal mass that caused severe acute respiratory symptoms. At the histological examination, the nasopharyngeal mucosa exhibited a diffuse pattern of infiltration by neoplastic cells with a characteristic single-file arrangement. The cells showed typical convoluted nuclei with one or two nucleoli and abundant cytoplasm (L2, lymphoblasts). The malignant cell population expressed a preT-cell immunophenotype: cytoplasmic CD3(+), CD43 (+), TdT (+/−), CD34(+), CD4(−), CD8(−). Thus, the diagnosis of T-cell LL was formulated. Whole-body CT scan revealed nasopharingeal mass, retropharyngeal, laterocervical, axillary, inguinal enlarged nodes and splenomegaly. Laboratory tests indicated leukocytosis (58,000/μl) with a differential WBC count typical of CML in chronic phase (myeloblasts &lt;1%). This diagnosis was confirmed by a low alkaline phosphatase value (score: 1), histological features of bone marrow (BM) biopsy, classical cytogenetics (presence of the Ph’ in 100% of 25 metaphases analysed and absence of other cytogenetic abnormalities) and FISH evaluation (D-FISH bcr-abl in interphase and metaphase, Oncor probe). The nested RT-PCR (JQ Guo et al., Leukemia;2002,15:2447) disclosed the presence of the hybrid protein p210 (b2,a2), but not that of p190. A laterocervical node was excised to perform cytogenetic and molecular analyses in order to determine whether the T-cell LL was an unrelated disease or the expression of an extramedullary BC. The histology confirmed the presence of a uniform population of T-lymphoblasts in which classical cytogenetic analysis disclosed the following kariotype: 49,XY,t(9;22)(q34;q11),+9,+19,+der(22)t(9;22)(q34;q11) and 50–52,XY t(9;22)(q34;q11),+6+9,+9q+,+19,+20,+22q−. The FISH analysis showed the presence of bcr-abl gene in all cells analyzed and the presence of multiple copies of this gene as well as of double Ph’ chromosome. Nested RT-PCR showed the presence of both p210 and p190 transcripts. These findings indicated that the T-cell LL was an extramedullary BC of a CML simultaneously diagnosed in chronic phase in BM and peripheral blood. The patient was initially treated with vincristine, daunomicine, asparaginase and prednisone combined with Gleevec (800mg/die). Consolidation therapy, consisting of high-dose Ara-C (4g/m2 for 4 days) and mithoxantrone (10mg/m2 for 2 days), followed by a mieloablative course with mithoxantrone (60 mg/m2) and melphalan (180 mg/m2) and autologous stem cell support (ASCT) was administered after an initial clinical response. Gleevec was given during the entire treatment period. A fugacious complete clinical-hematological and a partial cytogenetic (FISH: 12% of cells bcr-abl+) and molecular (number of bcr-abl/104 ABL copies= 184, real-time quantitative RT-PCR: J Gabert et al., Leukemia;2003:1) remission was documented after ASCT. The patient refused further consolidation treatment with Gleevec and died, due to progressive disease, in December 2004.

Efficacy of Imatinib-Based Therapy in a Patient with Resistant NUP214-ABL1 T-Cell Acute Lymphoblastic Leukemia.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4329-4329
Author(s):  
Jacques Delaunay ◽  
Raouf Ben Abdelali ◽  
Cyrille Touzeau ◽  
Patrice Chevallier ◽  
Guillaume Thierry ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Lymphoblastic Leukemia ◽  
Pcr Analysis ◽  
Fish Analysis ◽  
Molecular Response ◽  
Rt Pcr ◽  
First Case ◽  
Cell Acute Lymphoblastic Leukemia

Abstract T-cell acute lymphoblastic leukemia (T-ALL) is a rare disease which represents 25% of adult ALL. In T-ALL, many cryptic abnormalities can only be detected by FISH or RQ-PCR. Herein, we describe the case of a 25 year-old man presenting with a NUP214-ABL1 T-ALL. This rearrangement is observed in about 5% of adult T-ALL, results in ABL1 amplification and has been associated with sensitivity to imatinib in vitro At diagnosis, the patient presented with multiple lymphadenopathies and a high WBC (120 G/L). Blast cells expressed T (CD3, CD5, CD7, and CD8) and myeloid (CD13 and CD33) markers. Karyotype was normal, while FISH analysis revealed HOX11L2 rearrangement and extrachromosomal ABL amplification. Presence of NUP214 exon32-ABL1 exon2 transcript and overexpression of HOX11L2 were both confirmed by RT-PCR. Treatment induction according to the GRAALL-2003 protocol, consisted of a corticosteroïd prephase (d1–7) followed by 5-drug induction (doxorubicin, cyclophosphamide, vincristine, corticosteroid and asparaginase) (d8–21). Because of ALL resistance and presence of severe infection, treatment was stopped at d21 and salvage therapy associating imatinib (400 mg, bid), vincristine (2 mg d1,d8,d15,d21) and dexamethasone (30 mg/m2 d1–2,d8–9,d15–16,d21–22) was initiated at d25. At d50, the patient had reached complete hematological response. He then received monthly consolidation courses combining imatinib (400 mg bid d1–28), vincristine (2 mg d1) and prednisolone (40mg/m2 d1–7). After the second course, he reached major molecular response at d120 (NUP214/ABL1 = 1x10-3 by RQ-PCR) but relapsed at d150. Karyotype was still normal and FISH analysis showed persistance of HOX11L2 rearrangement but loss of extrachromosomal ABL amplification and presence of a third ABL signal (cryptic rearrangement) in the majority of metaphases. Quantitative RT-PCR analysis detected a low level of NUP214/ABL1. We initiated a second salvage combining dasatinib (70 mg bid) and Hyper-CVAD, with achievement of a second hematological CR. After two consolidation courses, the patient is still in hematological CR (8 months) and complete molecular response. He will receive a double cord blood transplant In conclusion, we report here the first case of transitory imatinib efficacy in a patient with NUP214/ABL1 T-ALL, followed by achievement of a secondary remission with dasatinib, despite the partial loss of NUP214-ABL1, in favor of the use of ABL inhibitors in this specific T-ALL subset. We also raise the hypothesis that NUP214/ABL1 may be a secondary oncogenic event.

Multilineage dysplasia has no impact on biologic, clinicopathologic, and prognostic features of AML with mutated nucleophosmin (NPM1)

Blood ◽  
2010 ◽  
Vol 115 (18) ◽  
pp. 3776-3786 ◽  
Author(s):  
Brunangelo Falini ◽  
Katja Macijewski ◽  
Tamara Weiss ◽  
Ulrike Bacher ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Who Classification ◽  
Hox Genes ◽  
Kinase Domain ◽  
World Health ◽  
Tyrosine Kinase Domain ◽  
Male Predominance ◽  
Prognostic Features ◽  
Multilineage Dysplasia ◽  
Health Organization

Abstract NPM1-mutated acute myeloid leukemia (AML) is a provisional entity in the 2008 World Health Organization (WHO) classification of myeloid neoplasms. The significance of multilineage dysplasia (MLD) in NPM1-mutated AML is unclear. Thus, in the 2008 WHO classification, NPM1-mutated AML with MLD is classified as AML with myelodysplasia (MD)–related changes (MRCs). We evaluated morphologically 318 NPM1-mutated AML patients and found MLD in 23.3%. Except for a male predominance and a lower fms-related tyrosine kinase 3–internal tandem duplication (FLT3-ITD) incidence in the MLD+ group, no differences were observed in age, sex, cytogenetics, and FLT3-–tyrosine kinase domain between NPM1-mutated AML with and without MLD. NPM1-mutated AML with and without MLD showed overlapping immunophenotype (CD34 negativity) and gene expression profile (CD34 down-regulation, HOX genes up-regulation). Moreover, overall and event-free survival did not differ among NPM1-mutated AML patients independently of whether they were MLD+ or MLD−, the NPM1-mutated/FLT3-ITD negative genotype showing the better prognosis. Lack of MLD impact on survival was confirmed by multivariate analysis that highlighted FLT3-ITD as the only significant prognostic parameter in NPM1-mutated AML. Our findings indicate that NPM1 mutations rather than MLD dictate the distinctive features of NPM1-mutated AML. Thus, irrespective of MLD, NPM1-mutated AML represents one disease entity clearly distinct from AML with MRCs.

FLT3 Is Fused to ETV6 in a Myeloproliferative Disorder with a t(12;13)(p13;q12) Translocation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2901-2901
Author(s):  
Hoang Anh Vu ◽  
Phan Thi Xinh ◽  
Seiko Shimizu ◽  
Michihiko Masuda ◽  
Toshiko Motoji ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Blot Analysis ◽  
Fusion Proteins ◽  
Fusion Gene ◽  
Hematologic Malignancies ◽  
Myeloproliferative Disorder ◽  
Tyrosine Kinase Domain ◽  
Common Mutation ◽  
Fish Analysis ◽  
Fusion Partner

Abstract The FLT3, located at band 13q12 and encoding a receptor tyrosine kinase (RTK), is one of the most frequently mutated genes in hematologic malignancies including ALL, MDS, and AML. The most common mutation of the FLT3 is an internal tandem duplication in exons 14 and 15, whereas other mutations have also been found at and around codon 835 of exon 20. These activating mutations promote constitutive RTK activity in the absence of ligand, proposing FLT3 as an attractive therapeutic target for directed inhibition. However, many questions with regards to the biology of FLT3 and its role in leukemogenesis remain to be clarified. Despite its highly frequent mutations, FLT3 has never been reported to fuse to any other genes, a phenomenon usually observed in other RTKs. Here, we report a case of a novel fusion gene between FLT3 and ETV6 at 12p13, a well-known target for a number of translocations. The patient, a 68-year-old female, was diagnosed as myeloproliferative disorder with hypereosinophilia in May 2002. Peripheral blood showed WBC 33.6x106/L (3% myelocytes, 33.5% neutrophils, 54% eosinophils, 1.5% basophils, 1.0% monocytes and 7% lymphocytes), Hb 119g/L and platelet counts 5,450x106/L. The bone marrow (BM) was marked hypercellular with 0.9% blasts, 6.0% promyelocytes, 15.6% myelocytes, 8.1% immature eosinophils and 19.2% mature eosinophils. Karyotype of BM cells was 46, XX, t(12;13)(p13.1;q12.3–13)[28]/46, XX[2]. Under the suspicion of Ph-negative CML, she was treated with IFNα with no response. Then, HU was started and her WBC decreased to 30x106/L. FISH analysis showed that the breakpoint at 12p13 occurred within ETV6, while the breakpoints at 13q12 occurred at two locations, within FLT3 or CDX2. To identify the fusion partner of ETV6, 3′-RACE PCR was performed. Sequence analysis of PCR-products revealed 4 types of ETV6/FLT3 transcripts. These fusion transcripts were confirmed by Northern blot analysis. Each ETV6/FLT3 transcript contained the entire helix-loop-helix domain of ETV6 (exons 1 to 4 or 5) and almost all of the functional domains of FLT3 including the tyrosine kinase domain (from exons 14, 16 or 17), suggesting that the resultant chimeric protein would be constitutively activated FLT3 kinase. Of them, three are in-frame fusion, presumably encoding for the approximately 58, 62, and 83 kD fusion proteins. However, Western blot analysis showed only expression of the 58 and 83 kD proteins. RT-PCR detected the reciprocal FLT3/ETV6 transcript, comprising the FLT3 exons 1 to 13 frameshifly fused to the ETV6 exons 6 to 8, within which a stop codon appeared at codon 33- downstream from the fusion point. Functional studies to assess the oncogenic properties of these fusion proteins are now in progress. Our findings provide an evidence that FLT3 is also involved in hematologic malignancies as a fusion gene.

Recurrent Fusion of PCM1 to JAK2 in Atypical Chronic Myeloid Leukemia and Acute Leukemia Associated with the t(8;9)(p21-22;p23-24).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2963-2963
Author(s):  
Andreas Reiter ◽  
Christoph Walz ◽  
Ann Watmore ◽  
Claudia Schoch ◽  
Ilona Blau ◽  
...  
Keyword(s):  
Acute Leukemia ◽  
Hematological Malignancies ◽  
Coiled Coil ◽  
Janus Kinase ◽  
Tyrosine Kinase Domain ◽  
Fish Analysis ◽  
Rt Pcr ◽  
Idiopathic Myelofibrosis ◽  
Secondary Aml ◽  
Atypical Cml

Abstract We have identified a novel, recurrent t(8;9)(p21–22;p23–24) in six patients with diverse hematological malignancies: atypical CML (n=4), secondary AML following idiopathic myelofibrosis (n=1) and pre B-ALL (n=1). Because of the involvement of several different tyrosine kinases in atypical CML, we focused our analysis on this class of gene. Initial FISH studies of one patient indicated that the janus kinase 2 gene (JAK2), located at 9p24, was disrupted. RACE-PCR was then used to identify the 8p21 partner gene as PCM1, a large centrosomal protein that contains multiple coiled-coil domains. RT-PCR and FISH analysis confirmed the fusion in this case, and also identified PCM1-JAK2 in the five other t(8;9) patients (RT-PCR and FISH, n=4; RT-PCR only, n=1; FISH only, n=1). Four different types of in-frame mRNA junction were identified, but in all cases the chimeric mRNA is predicted to encode a protein that retains several of the predicted coiled-coil domains from PCM1 and the entire tyrosine kinase domain of JAK2. Reciprocal JAK2-PCM1 mRNA could not be amplified in any patient. Clinically, 4 patients displayed CML-like hyperplasia with variable degrees of myelofibrosis and eosinophilia. Similar to typical CML, the clinical course of these patients was variable: one is alive 11 months after allogeneic stem cell transplantation, one transformed to acute leukemia 5 years after diagnosis, one died 4 days after presentation and one achieved a major cytogenetic response with interferon but died due to neurodegenerative disease and pneumonia at 7.5 years. Of the two remaining patients, one presented with secondary AML following idiopathic myelofibrosis and remains in remission 15 years after diagnosis following intensive chemotherapy and maintenance with interferon. The final patient died shortly after induction therapy for pre B-ALL. In conclusion, PCM1-JAK2 is a novel recurrent fusion gene in hematological malignancies that is likely to deregulate hemopoiesis in a manner similar to BCR-ABL. Patients with PCM1-JAK2 disease are attractive targets for signal transduction therapy.

Amplification at 7q22 Targets CDK6 in T-Cell Lymphoma.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3563-3563
Author(s):  
Stefan Nagel ◽  
Ellen Leich ◽  
Hilmar Quentmeier ◽  
Corinna Meyer ◽  
Andreas Zettl ◽  
...  
Keyword(s):  
Cell Cycle ◽  
T Cell ◽  
Cell Line ◽  
Copy Number ◽  
Cell Lymphoma ◽  
T Cell Lymphoma ◽  
Rt Pcr ◽  
Genomic Amplification ◽  
Genetic Event ◽  
T Cell Lymphomas

Abstract Genomic amplification is microscopically visible extrachromosomally as double-minute chromosomes (DMIN) or intrachromosomally as homogeneously stained regions (HSR). Both DMIN and HSR comprise iterated copies of discrete genomic regions (amplicons) effecting copy number increases associated with upregulation of target genes. In cancer cells, these usually comprise oncogenes such as MYC. Cytogenetic analysis of the anaplastic large cell lymphoma (ALCL) cell line SU-DHL-1 revealed an HSR on the long arm region of chromosome 7 (7q). Detailled analysis of this region by fluorescence in situ hybridization (FISH) using tilepath BAC clones identified an amplicon corresponding to 86–95 Mb (NCBI Build 36.1) at 7q22. Expression analysis using reverse transcription (RT)-PCR of candidate oncogenes mapped within the amplicon, comprising DBF4, SRI, AKAP9, GATAD1, CDK6 and PPP1R9A, highlighted cyclin-dependent kinase 6 (CDK6) as a plausible amplification target. With reference to other ALCL cell lines, including SR-786, lacking this amplicon, SU-DHL-1 displays upregulation of CDK6 at both the RNA and protein levels as indicated by semiquantitative RT-PCR, real-time quantitative (RQ)-PCR and immuno-cytochemistry. Another ALCL cell line, L-82, containing a larger amplicon (7q21-31), also showed enhanced CDK6 expression as analyzed by RQ-PCR. For a functional test both SU-DHL-1 and SR-786 cells were treated with rapamycin, an inhibitor of the AKT-pathway which is involved in degradation of CDK-inhibitor p27. In contrast to SR-786, SU-DHL-1 cells are resistant to the effects of rapamycin on both proliferation and G1 cell cycle arrest, implying that CDK6 overexpression may confer proliferative advantage. Copy number analysis of several tumor-relevant genes using a commercial PCR-based approach (MLPA) determined a nearly 5-fold amplification of the CDK6 gene in SU-DHL-1. Interphase FISH confirmed copy number increase in SU-DHL-1 and identified CDK6 amplification in 1/77 ALCL and 4/38 peripheral T-cell lymphoma patient samples, indicating that this genetic event is relatively rare among T-cell lymphomas. Taken together, we have identified an amplicon at 7q22 in T-cell lymphoma cells targeting CDK6 which is an important cell cycle regulator and probably connected to the increased proliferative capacity of these malignant cells. Although CDK6 amplification seems to be infrequent overall, it strengthens the importance of this gene in regulating proliferation among T-cell lymphomas and may, therefore, represent a potential molecular target for therapeutic intervention in these disease entities.

scholarly journals Fine-tuning of substrate preferences of the Src-family kinase Lck revealed through a high-throughput specificity screen

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Neel H Shah ◽  
Mark Löbel ◽  
Arthur Weiss ◽  
John Kuriyan
Keyword(s):  
T Cell ◽  
Tyrosine Kinase ◽  
Cell Signaling ◽  
High Throughput ◽  
Tyrosine Kinases ◽  
Catalytic Domain ◽  
Surface Display ◽  
Fine Tuning ◽  
Tyrosine Kinase Domain ◽  
T Cell Signaling

The specificity of tyrosine kinases is attributed predominantly to localization effects dictated by non-catalytic domains. We developed a method to profile the specificities of tyrosine kinases by combining bacterial surface-display of peptide libraries with next-generation sequencing. Using this, we showed that the tyrosine kinase ZAP-70, which is critical for T cell signaling, discriminates substrates through an electrostatic selection mechanism encoded within its catalytic domain (Shah et al., 2016). Here, we expand this high-throughput platform to analyze the intrinsic specificity of any tyrosine kinase domain against thousands of peptides derived from human tyrosine phosphorylation sites. Using this approach, we find a difference in the electrostatic recognition of substrates between the closely related Src-family kinases Lck and c-Src. This divergence likely reflects the specialization of Lck to act in concert with ZAP-70 in T cell signaling. These results point to the importance of direct recognition at the kinase active site in fine-tuning specificity.

Sign in / Sign up

Export Citation Format

Share Document