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Limited duration of complete remission on ruxolitinib in myeloid neoplasms with PCM1-JAK2 and BCR-JAK2 fusion genes.
Ann. Hematol.
PUBLISHED: 09-11-2014
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Rearrangements of chromosome band 9p24 are known to be associated with JAK2 fusion genes, e.g., t(8;9)(p22;p24) with a PCM1-JAK2 and t(9;22)(p24;q11) with a BCR-JAK2 fusion gene, respectively. In association with myeloid neoplasms, the clinical course is aggressive, and in absence of effective conventional treatment options, long-term remission is usually only observed after allogeneic stem cell transplantation (ASCT). With the discovery of inhibitors of the JAK2 tyrosine kinase and based on encouraging in vitro and in vivo data, we treated two male patients with myeloid neoplasms and a PCM1-JAK2 or a BCR-JAK2 fusion gene, respectively, with the JAK1/JAK2 inhibitor ruxolitinib. After 12 months of treatment, both patients achieved a complete clinical, hematologic, and cytogenetic response. Non-hematologic toxicity was only grade 1 while no hematologic toxicity was observed. However, remission in both patients was only short-term, with relapse occurring after 18 and 24 months, respectively, making ASCT indispensable in both cases. This data highlight (1) the ongoing importance of cytogenetic analysis for the diagnostic work-up of myeloid neoplasms as it may guide targeted therapy and (2) remission under ruxolitinib may only be short-termed in JAK2 fusion genes but it may be an important bridging therapy prior to ASCT.
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Identification and functional characterization of imatinib-sensitive DTD1-PDGFRB and CCDC88C-PDGFRB fusion genes in eosinophilia-associated myeloid/lymphoid neoplasms.
Genes Chromosomes Cancer
PUBLISHED: 04-29-2014
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Eosinophilia-associated myeloid neoplasms with rearrangement of chromosome bands 5q31-33 are frequently associated with PDGFRB fusion genes, which are exquisitely sensitive to treatment with imatinib. In search for novel fusion partners of PDGFRB, we analyzed three cases with translocation t(5;20)(q33;p11), t(5;14)(q33;q32), and t(5;17;14)(q33;q11;q32) by 5?-rapid amplification of cDNA ends polymerase chain reaction (5?-RACE-PCR) and DNA-based long-distance inverse PCR (LDI-PCR) with primers derived from PDGFRB. LDI-PCR revealed a fusion between CCDC88C exon 25 and PDGFRB exon 11 in the case with t(5;17;14)(q33;q11;q32) while 5?-RACE-PCR identified fusions between CCDC88C exon 10 and PDGFRB exon 12 and between DTD1 exon 4 and PDGFRB exon 12 in the cases with t(5;14)(q33;q32) and t(5;20)(q33;p11), respectively. The PDGFRB tyrosine-kinase domain is predicted to be retained in all three fusion proteins. The partner proteins contained coiled-coil domains or other domains, which putatively lead to constitutive activation of the PDGFRB fusion protein. In vitro functional analyses confirmed transforming activity and imatinib-sensitivity of the fusion proteins. All three patients achieved rapid and durable complete hematologic remissions on imatinib.
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The KIT D816V expressed allele burden for diagnosis and disease monitoring of systemic mastocytosis.
Ann. Hematol.
PUBLISHED: 10-15-2013
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The activating KIT D816V mutation plays a central role in the pathogenesis, diagnosis, and targeted treatment of systemic mastocytosis (SM). For improved and reliable identification of KIT D816V, we have developed an allele-specific quantitative real-time PCR (RQ-PCR) with an enhanced sensitivity of 0.01-0.1 %, which was superior to denaturing high-performance liquid chromatography (0.5-1 %) or conventional sequencing (10-20 %). Overall, KIT D816 mutations were identified in 146/147 (99 %) of patients (D816V, n?=?142; D816H, n?=?2; D816Y, n?=?2) with SM, including indolent SM (ISM, n?=?63, 43 %), smoldering SM (n?=?8, 5 %), SM with associated hematological non-mast cell lineage disease (SM-AHNMD, n?=?16, 11 %), and aggressive SM/mast cell leukemia?±?AHNMD (ASM/MCL, n?=?60, 41 %). If positive in BM, the KIT D816V mutation was found in PB of all patients with advanced SM (SM-AHNMD, ASM, and MCL) and in 46 % (23/50) of patients with ISM. There was a strong correlation between the KIT D816V expressed allele burden (KIT D816V EAB) with results obtained from DNA by genomic allele-specific PCR and also with disease activity (e.g., serum tryptase level), disease subtype (e.g., indolent vs. advanced SM) and survival. In terms of monitoring of residual disease, qualitative and quantitative assessment of KIT D816V and KIT D816V EAB was successfully used for sequential analysis after chemotherapy or allogeneic stem cell transplantation. We therefore conclude that RQ-PCR assays for KIT D816V are useful complimentary tools for diagnosis, disease monitoring, and evaluation of prognosis in patients with SM.
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Comprehensive mutational profiling in advanced systemic mastocytosis.
Blood
PUBLISHED: 08-19-2013
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To explore mechanisms contributing to the clinical heterogeneity of systemic mastocytosis (SM) and to suboptimal responses to diverse therapies, we analyzed 39 KIT D816V mutated patients with indolent SM (n = 10), smoldering SM (n = 2), SM with associated clonal hematologic nonmast cell lineage disorder (SM-AHNMD, n = 5), and aggressive SM (n = 15) or mast cell leukemia (n = 7) with (n = 18) or without (n = 4) AHNMD for additional molecular aberrations. We applied next-generation sequencing to investigate ASXL1, CBL, IDH1/2, JAK2, KRAS, MLL-PTD, NPM1, NRAS, TP53, SRSF2, SF3B1, SETBP1, U2AF1 at mutational hotspot regions, and analyzed complete coding regions of EZH2, ETV6, RUNX1, and TET2. We identified additional molecular aberrations in 24/27 (89%) patients with advanced SM (SM-AHNMD, 5/5; aggressive SM/mast cell leukemia, 19/22) whereas only 3/12 (25%) indolent SM/smoldering SM patients carried one additional mutation each (U2AF1, SETBP1, CBL) (P < .001). Most frequently affected genes were TET2, SRSF2, ASXL1, CBL, and RUNX1. In advanced SM, 21/27 patients (78%) carried ?3 mutations, and 11/27 patients (41%) exhibited ?5 mutations. Overall survival was significantly shorter in patients with additional aberrations as compared to those with KIT D816V only (P = .019). We conclude that biology and prognosis in SM are related to the pattern of mutated genes that are acquired during disease evolution.
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Impact of additional cytogenetic aberrations at diagnosis on prognosis of CML: long-term observation of 1151 patients from the randomized CML Study IV.
Blood
PUBLISHED: 10-28-2011
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The prognostic relevance of additional cytogenetic findings at diagnosis of chronic myeloid leukemia (CML) is unclear. The impact of additional cytogenetic findings at diagnosis on time to complete cytogenetic (CCR) and major molecular remission (MMR) and progression-free (PFS) and overall survival (OS) was analyzed using data from 1151 Philadelphia chromosome-positive (Ph(+)) CML patients randomized to the German CML Study IV. At diagnosis, 1003 of 1151 patients (87%) had standard t(9;22)(q34;q11) only, 69 patients (6.0%) had variant t(v;22), and 79 (6.9%) additional cytogenetic aberrations (ACAs). Of these, 38 patients (3.3%) lacked the Y chromosome (-Y) and 41 patients (3.6%) had ACAs except -Y; 16 of these (1.4%) were major route (second Philadelphia [Ph] chromosome, trisomy 8, isochromosome 17q, or trisomy 19) and 25 minor route (all other) ACAs. After a median observation time of 5.3 years for patients with t(9;22), t(v;22), -Y, minor- and major-route ACAs, the 5-year PFS was 90%, 81%, 88%, 96%, and 50%, and the 5-year OS was 92%, 87%, 91%, 96%, and 53%, respectively. In patients with major-route ACAs, the times to CCR and MMR were longer and PFS and OS were shorter (P < .001) than in patients with standard t(9;22). We conclude that major-route ACAs at diagnosis are associated with a negative impact on survival and signify progression to the accelerated phase and blast crisis.
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Expression of Transketolase like gene 1 (TKTL1) predicts disease-free survival in patients with locally advanced rectal cancer receiving neoadjuvant chemoradiotherapy.
BMC Cancer
PUBLISHED: 03-22-2011
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For patients with locally advanced rectal cancer (LARC) neoadjuvant chemoradiotherapy is recommended as standard therapy. So far, no predictive or prognostic molecular factors for patients undergoing multimodal treatment are established. Increased angiogenesis and altered tumour metabolism as adaption to hypoxic conditions in cancers play an important role in tumour progression and metastasis. Enhanced expression of Vascular-endothelial-growth-factor-receptor (VEGF-R) and Transketolase-like-1 (TKTL1) are related to hypoxic conditions in tumours. In search for potential prognostic molecular markers we investigated the expression of VEGFR-1, VEGFR-2 and TKTL1 in patients with LARC treated with neoadjuvant chemoradiotherapy and cetuximab.
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Activating CBL mutations are associated with a distinct MDS/MPN phenotype.
Ann. Hematol.
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Activating point mutations in CBL have recently been identified in diverse subtypes of myeloid neoplasms. Because detailed clinical and hematological characteristics of CBL-mutated cases is lacking, we screened 156 BCR-ABL and JAK2 V617F negative patients with myeloproliferative neoplasms (MPN) and overlap syndromes between myelodysplastic syndrome (MDS) and MPN (MPS/MPN) for mutations in exons 8 and 9 of CBL by denaturing high-performance liquid chromatography and direct sequencing. CBL mutations were identified in 16/156 patients (10%), of which five also carried mutations in EZH2 (n?=?3) and TET2 (n?=?2). Comprehensive clinical and hematological characteristics were available from 13/16 patients (81%). In addition to splenomegaly (77%), striking common hematological features were CML-like left-shifted leukocytosis (85%) with monocytosis (85%), anemia (100%), and thrombocytopenia (62%). Thrombocytosis was not observed in any patient. Relevant bone marrow features (n?=?12) included hypercellularity (92%) with marked granulopoiesis (92%), nonclustered microlobulated megakaryocytes (83%), and marrow fibrosis (83%). Nine deaths (progression to secondary acute myeloid leukemia/blast phase, n?=?7; cytopenia complications, n?=?2) were recorded. Three-year survival rate was 27%, possibly indicating poor prognosis of CBL mutated MDS/MPN patients.
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Ruxolitinib as potential targeted therapy for patients with JAK2 rearrangements.
Haematologica
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JAK2 fusion genes are rare but recurrent abnormalities associated with diverse, clinically heterogeneous hematologic malignancies. Here we assess the JAK1/2 inhibitor ruxolitinib as therapy for patients with JAK2-rearrangement associated myeloproliferative neoplasms (MPN). Ruxolitinib-treated Ba/F3 cells transformed to IL3 independence by ETV6-JAK2 showed reduced proliferation and survival (IC(50) = 370 nM) compared with KG1A or Ba/F3 cells transformed by BCR-ABL1, SPBN1-FLT3 and ZMYM2-FGFR1 (IC(50) > 10 ?M for all). Inhibition was associated with reduced phosphorylation of ETV6-JAK2, ERK, STAT5 and AKT. Primary cell growth from 2 patients with JAK2 rearrangement and one patient with JAK2 amplification was assessed in methylcellulose assays. Reduced colony growth was seen for all patients in ruxolitinib-treated cultures compared with healthy controls (n=7). Fluorescence in situ hybridization showed reduced growth of JAK2-rearrangement positive colonies compared to JAK2-rearrangement negative colonies. Our data, therefore, provide evidence that ruxolitinib is a promising therapy for treatment of patients with JAK2 fusion genes.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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