KIF5B-RET: Discovery of a novel fusion oncogene in lung adenocarcinomas by a systematic screen for tyrosine kinase fusions and identification of patients for a RET targeted therapy trial.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. 7578-7578
Author(s):  
Yoshiyuki Suehara ◽  
Maria E. Arcila ◽  
Alexander Edward Dela Cruz Drilon ◽  
Tatsuo Ito ◽  
Lu Wang ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Kinase Domain ◽  
Positive Sample ◽  
The Novel ◽  
Rt Pcr ◽  
Dimerization Domain ◽  
Phase 2 Trial ◽  
Related Variant ◽  
Rapid Amplification ◽  
Lung Adenocarcinomas

7578 Background: The mutually exclusive pattern of major targetable driver oncogenes in lung adenocarcinomas (ADC) suggests that other similar driver oncogenes may exist. We therefore performed a systematic screen for tyrosine kinase (TK) fusions in cases without known driver oncogenes by measuring aberrantly high RNA expression of kinase domain (KD) exons relative to more 5’ exons. Methods: We studied 74 patients whose lung ADC lacked mutations in KRAS, EGFR, BRAF, HER2, and ALK fusions. A NanoString-based assay was designed to query the transcripts of 90 TKs at two points: 5’ to the KD and within or 3’ to the KD. Tumor RNAs were hybridized to the NanoString probes and analyzed for outlier 3’ to 5’ expression ratios. The assay was validated on samples with known ALK and ROS fusions. Presumed novel fusion events were followed up by rapid amplification of cDNA ends (RACE) and confirmatory RT-PCR. Results: The NanoString assay identified aberrant 5’ to 3’ ratios in ROS and RET in 2 cases, respectively, out of 74. RACE analysis isolated a novel GOPC-ROS fusion in the former and a novel KIF5B-RET fusion in the latter, both confirmed by RT-PCR. Further screening by RT-PCR for KIF5B-RET identified one more positive sample in the study set that had not been detected by NanoString. At the RNA level, both fusions joined exon 15 of KIF5B to exon 12 of RET, thus retaining a portion of the dimerization domain of KIF5B and the entire KD of RET, analogous to RET fusions in papillary thyroid carcinoma (TC). One KIF5B-RET patient was a 60 y.o. female never smoker, the other, a 73 y.o. male former smoker. Conclusions: The novel KIF5B-RET fusion described here and also recently reported by Ju YS et al. (Genome Res, Dec 22, 2011) defines a new subset of lung ADC with a potentially targetable driver oncogene. Based on these genetic data and the preclinical activity of the RET inhibitor XL184 (Exelixis) in papillary TC and its known activity in medullary TC with RET mutations, we have initiated prospective testing for KIF5B-RET as part of our lung ADC screening panel in anticipation of a planned phase 2 trial with XL184 in patients with KIF5B-RET or related variant RET fusions.

Clinical Course and Significance of the Novel FLT3-Y842C Mutation in a Patient with AML Treated with PKC412 Monotherapy.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2537-2537
Author(s):  
T. Kindler ◽  
F. Breitenbuecher ◽  
S. Kasper ◽  
E. Estey ◽  
F. Giles ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Clinical Course ◽  
Novel Mutation ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Activation Loop ◽  
The Novel ◽  
Constitutive Activation

Abstract We recently identified a novel mutation (Y842C) within the tyrosine kinase domain of FLT3 in a patient treated with PKC410 monotherapy (ASH 2003, # 4681). Here, we present follow up studies including the clinical course of the patient and frequency analysis in 110 patients with AML. In addition, we characterized the novel mutation using overexpression of FLT3-Y842C in 32D cells. AML M2 was diagnosed in a 63 year old, male patient in 1993. After having experienced his second relapse upon standard therapy the patient was refractory to alemtuzumab treatment. Due to reduced performance status the patient was not eligible to standard chemotherapy and was enrolled into a phase II trial investigating PKC412. On conventional FLT3 mutation analysis the patient was considered to be FLT3 wild-type. Upon 8 and 29 days of treatment complete clearance of PB blast counts and BM blast infiltration was observed, respectively. Daily substitution of G-CSF resulted in transient recovery or the patients ANC′s. Since the patient showed an excellent clinical responsiveness, we reasoned whether the patient may have a yet unidentified FLT3 mutation. Sequence analysis revealed a novel point mutation in exon 21 of FLT3 (Y842C). Protein analysis of primary AML blasts showed constitutive FLT3 tyrosine-phosphorylation, ex vivo treatment with PKC412 caused significant inhibition of FLT3 and STAT5 activation. Further, in vivo analysis of FLT3 tyrosine-phosphorylation during the course of PKC412 treatment showed complete suppression of FLT3 activation within 8 days. Overexpression of FLT3-Y842C in 32D cells resulted in constitutive activation of FLT3 and STAT5 as well as in factor independent proliferation. Treatment with PKC412 caused inhibition of FLT3 tyrosine-phosphorylation, factor independent growth and apoptotic cell death. To further investigate the clinical significance of the novel Y842C mutation, the tyrosine kinase domain of FLT3 was investigated in 110 patients with AML using sequence analysis. Altogether, the novel mutation Y842C was identified in 2 patients, FLT-ITD in 22 patients and D835 in 7 patients, respectively. It is interesting to note that the recently described crystal structure of FLT3 reveals a critical role for Y842 in regulating the switch from the closed to the open (=active) conformation of the FLT3 activation loop. Since our data is consistent with the concept that the Y842C mutation results in constitutive activation of FLT3, it is tempting to speculate that the exchange of tyrosine for cysteine at position 842 disrupts the autoinhibited state of the FLT3 activation loop. Given that the novel mutation described here could only be identified by direct sequencing, it is likely that the number of mutations in this region of FLT3 is currently underestimated. Thus, extended sequence analysis of this mutational hotspot may be helpful in further defining the spectrum of TKI-sensitive FLT3 mutations in AML.

scholarly journals Shc and Enigma Are Both Required for Mitogenic Signaling by Ret/ptc2

1998 ◽  
Vol 18 (4) ◽  
pp. 2298-2308 ◽  
Author(s):  
Kyle Durick ◽  
Gordon N. Gill ◽  
Susan S. Taylor
Keyword(s):  
Tyrosine Kinase ◽  
Molecular Complexes ◽  
Kinase Domain ◽  
Potential Interaction ◽  
Dominant Negative ◽  
Tyrosine Kinase Domain ◽  
Chimeric Proteins ◽  
Cell Periphery ◽  
Mitogenic Signaling ◽  
Dimerization Domain

ABSTRACT Ret/ptc2 is a constitutively active, oncogenic form of the c-Ret receptor tyrosine kinase. Like the other papillary thyroid carcinoma forms of Ret, Ret/ptc2 is activated through fusion of the Ret tyrosine kinase domain to the dimerization domain of another protein. Investigation of requirements for Ret/ptc2 mitogenic activity, using coexpression with dominant negative forms of Ras and Raf, indicated that these proteins are required for mitogenic signaling by Ret/ptc2. Because activation of Ras requires recruitment of Grb2 and SOS to the plasma membrane, the subcellular distribution of Ret/ptc2 was investigated, and it was found to localize to the cell periphery. This localization was mediated by association with Enigma via the Ret/ptc2 sequence containing tyrosine 586. Because Shc interacts with MEN2 forms of Ret, and because phosphorylation of Shc results in Grb2 recruitment and subsequent signaling through Ras and Raf, the potential interaction between Ret/ptc2 and Shc was investigated. The PTB domain of Shc also interacted with Ret/ptc2 at tyrosine 586, and this association resulted in tyrosine phosphorylation of Shc. Coexpression of chimeric proteins demonstrated that mitogenic signaling from Ret/ptc2 required both recruitment of Shc and subcellular localization by Enigma. Because Shc and Enigma interact with the same site on a Ret/ptc2 monomer, dimerization of Ret/ptc2 allows assembly of molecular complexes that are properly localized via Enigma and transmit mitogenic signals via Shc.

EGFR Mutations in Non–Small-Cell Lung Cancer: Analysis of a Large Series of Cases and Development of a Rapid and Sensitive Method for Diagnostic Screening With Potential Implications on Pharmacologic Treatment

2005 ◽  
Vol 23 (4) ◽  
pp. 857-865 ◽  
Author(s):  
Antonio Marchetti ◽  
Carla Martella ◽  
Lara Felicioni ◽  
Fabio Barassi ◽  
Simona Salvatore ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Tyrosine Kinase ◽  
Direct Sequencing ◽  
Kinase Domain ◽  
Small Cell ◽  
Sscp Analysis ◽  
Egfr Mutations ◽  
Small Cell Lung ◽  
Lung Carcinomas ◽  
Lung Adenocarcinomas

Purpose It has been reported that EGFR mutations in lung carcinomas make the disease more responsive to treatment with tyrosine kinase inhibitors. We decided to evaluate the prevalence of EGFR mutations in a large series of non–small-cell lung carcinomas (NSCLCs) and to develop a rapid and sensitive screening method. Patients and Methods We examined 860 consecutive NSCLC patients for EGFR mutations in exons 18, 19, and 21 using a dual technical approach—direct sequencing of polymerase chain reaction (PCR) products and PCR single-strand conformation polymorphism (SSCP) analysis. Moreover, all lung adenocarcinomas were analyzed for K-ras mutations at codon 12 by allele-specific oligoprobe hybriditations. Results There were no EGFR mutations in 454 squamous carcinomas and 31 large cell carcinomas investigated. Thirty-nine mutations were found in the series of 375 adenocarcinomas (10%). Mutations were present in 26% of 86 bronchioloalveolar carcinomas (BACs) and in 6% of 289 conventional lung adenocarcinomas; P = .000002. EGFR mutations and K-ras mutations were mutually exclusive. A multivariable analysis revealed that BAC histotype, being a never smoker, and female sex were independently associated with EGFR mutations (odds ratios: 4.542, 3.632, and 2.895, respectively). The SSCP analysis was accurate and sensitive, allowing identification of mutations that were undetectable (21% of cases) by direct sequencing. Conclusion Mutations in the EGFR tyrosine kinase domain define a new molecular type of lung carcinoma, more frequent in particular subsets of patients. The SSCP assay is a rapid and reliable method for the detection of EGFR kinase domain mutations in lung cancer.

SPTBN1-FLT3 in Atypical Chronic Myeloid Leukemia Transforms Ba/F3 Cells to IL-3 Independence and Is Sensitive to Both Tyrosine Kinase Inhibitors and Immunotherapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2009-2009
Author(s):  
Francis H. Grand ◽  
Sameena Iqbal ◽  
Lingyan Zhang ◽  
Nigel H. Russell ◽  
Andrew Chase ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chronic Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Fusion Gene ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Unrelated Donor ◽  
Rt Pcr

Abstract We have identified a patient who presented with BCR-ABL negative chronic myeloid leukemia (CML) and an acquired 46XX, t(2;13;2;21) (p13;q12;q33;q11.2) in all bone marrow metaphases examined. Fluorescence in situ hybridization (FISH) using probes flanking the FLT3 gene at 13q12 suggested that this gene was disrupted. 5′-RACE PCR using primers to the region of FLT3 encoding the tyrosine kinase domain identified a novel in-frame mRNA fusion between exon 3 of SPTBN1 (spectrin, beta, non-erythrocytic 1 isoform 2, NM 178313) on chromosome 2p16 and exon 13 of FLT3 (NM 004119). Juxtaposition of SPTBN1 and FLT3 was confirmed by two color FISH and amplification of the genomic DNA breakpoint confirmed a fusion between intron 3 of SPTBN1 and intron 12 of FLT3. The SPTBN1-FLT3 fusion gene is predicted to be translated into a 570 amino acid chimeric protein that retains two coiled-coil domains from SPTBN1 and 424 amino acids from FLT3, including the entire tyrosine kinase domain. Since the t(2;13) is readily visible by cytogenetic analysis but has not been reported previously it seems likely that SPTBN1-FLT3 is uncommon. However to test if FLT3 might be involved more widely in BCR-ABL negative CML we analysed 40 cases by RT-PCR. Two cases were positive for the FLT3 internal tandem duplication (ITD) but mutation of residue D835 was not observed. Expression of the SPTBN1-FLT3 fusion transformed the interleukin 3 (IL-3)-dependent cell line Ba/F3 to growth factor independence and was accompanied by constitutive phosphorylation of the fusion protein and the downstream substrate ERK1/2. The growth of transformed cells was inhibited in a dose-dependent fashion by SU11567 and PKC142, but not by imatinib mesylate. The patient was initially treated with hydroxyurea and subsequently underwent an unrelated donor bone marrow transplant. She relapsed cytogenetically at 4 years but responded to donor lymphocyte infusion (DLI), achieving sustained cytogenetic and molecular (nested RT-PCR) remission. We conclude that SPTBN1-FLT3 is a rare abnormality in BCR-ABL negative CML that is responsive to both targeted signal transduction therapy and immunotherapy by DLI.

NIDDM associated with mutation in tyrosine kinase domain of insulin receptor gene

Diabetes ◽  
1992 ◽  
Vol 41 (4) ◽  
pp. 521-526 ◽  
Author(s):  
S. Cocozza ◽  
A. Porcellini ◽  
G. Riccardi ◽  
A. Monticelli ◽  
G. Condorelli ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Insulin Receptor ◽  
Receptor Gene ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Insulin Receptor Gene

The tyrosine kinase TEK is differentially expressed in non-small cell lung adenocarcinomas.

2020 ◽  
Author(s):  
Shahan Mamoor
Keyword(s):  
United States ◽  
Tyrosine Kinase ◽  
Lung Adenocarcinoma ◽  
Microarray Data ◽  
The United States ◽  
Small Cell ◽  
Small Cell Lung ◽  
Significant Differential Expression ◽  
Lung Cancers ◽  
Lung Adenocarcinomas

Non-small cell lung adenocarcinoma (NSCLC) is a leading cause of death in the United States and worldwide (1, 2). We mined published microarray data (3, 4, 5) to discover genes associated with NSCLC. We identified significant differential expression of the tyrosine kinase TEK in tumors from patients with NSCLC. TEK may be of relevance to the initiation, progression or maintenance of non-small cell lung cancers.

Characteristics of BCR/ABL1 kinase domain mutations in the patients with chronic myeloid leukemia who are primary resistant to the imatinib therapy

2019 ◽  
Vol 11 (1) ◽  
pp. 27-33
Author(s):  
I Dmytrenko ◽  
J Minchenko ◽  
I Dyagil
Keyword(s):  
Overall Survival ◽  
Chronic Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Myeloid Leukemia ◽  
High Efficiency ◽  
Kinase Domain ◽  
Imatinib Therapy ◽  
Missense Mutations ◽  
Primary Resistance ◽  
Kinase Domain Mutations

The chronic myeloid leukemia (CML) development is associated with the formation of the BCR/ABL1 fusion gene and the BCR/ABL1 protein with increased tyrosine kinase activity. Despite the high efficiency of targeted therapy, up to 30% of patients do not respond on such therapy i.e. are primary resistant. The presence of BCR/ABL1 kinase domain mutations is considered to be one of the reasons of tyrosin kinase inhibitors resistance. To evaluate the frequency of BCR/ABL1 kinase domain mutations in Ukrainian cohort of CML patients with primary resistance to imatinib therapy, we retrospectively studied BCR/ABL1 kinase domain mutations in peripheral blood of 107 CML patients. The nucleotide sequence was determined by direct sequencing by Sanger. Mutations were reported in 45 of 107 (41.7%) CML patients. Two mutations at a time were revealed in 8 patients. So a total of 53 mutations were found out. Among them 49 were missense-mutations and 4 - deletions of different regions of the BCR/ABL1 kinase domain gene. The missense-mutations F359I/V (12 patients), T315I (8 patients) and G250E (6 patients) were most common. By localization, the mutations majority (23 of 53) was in the P-loop, 10 mutations - in the contact site, 13 mutations - in the catalytic domain and 6 – in the A-loop. Of the detected mutations, 26 (49%) resulted in a disruption of the hydrogen bond between BCR/ABL1-tyrosine kinase and imatinib. Significant reduction in overall survival was found in patients with BCR/ABL1 kinase domain mutations compared with patients with wild-type of BCR/ABL1 gene (p=0.018). The estimated 3-year overall survival was 83.4% (95% CI: 77.0%-89.8%) and 94.3% (95% CI: 91.0%-97.3%), respectively. Therefore, mutations of the BCR/ABL1 kinase domain are one of the mechanisms of primary resistance in CML patients on imatinib therapy. The occurrence of BCR/ABL1 gene mutations impairs the prognosis of imatinib therapy response.

scholarly journals Bleeding by Bruton Tyrosine Kinase-Inhibitors: Dependency on Drug Type and Disease

Cancers ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 1103
Author(s):  
Philipp von Hundelshausen ◽  
Wolfgang Siess
Keyword(s):  
Clinical Trials ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Kinase Inhibitors ◽  
Phase 1 ◽  
The Novel ◽  
B Cell Malignancies ◽  
Bruton Tyrosine Kinase ◽  
Dermatological Disorders ◽  
Reversible Covalent

Bruton tyrosine kinase (Btk) is expressed in B-lymphocytes, myeloid cells and platelets, and Btk-inhibitors (BTKi) are used to treat patients with B-cell malignancies, developed against autoimmune diseases, have been proposed as novel antithrombotic drugs, and been tested in patients with severe COVID-19. However, mild bleeding is frequent in patients with B-cell malignancies treated with the irreversible BTKi ibrutinib and the recently approved 2nd generation BTKi acalabrutinib, zanubrutinib and tirabrutinib, and also in volunteers receiving in a phase-1 study the novel irreversible BTKi BI-705564. In contrast, no bleeding has been reported in clinical trials of other BTKi. These include the brain-penetrant irreversible tolebrutinib and evobrutinib (against multiple sclerosis), the irreversible branebrutinib, the reversible BMS-986142 and fenebrutinib (targeting rheumatoid arthritis and lupus erythematodes), and the reversible covalent rilzabrutinib (against pemphigus and immune thrombocytopenia). Remibrutinib, a novel highly selective covalent BTKi, is currently in clinical studies of autoimmune dermatological disorders. This review describes twelve BTKi approved or in clinical trials. By focusing on their pharmacological properties, targeted disease, bleeding side effects and actions on platelets it attempts to clarify the mechanisms underlying bleeding. Specific platelet function tests in blood might help to estimate the probability of bleeding of newly developed BTKi.

scholarly journals Characterization of Atypical Protein Tyrosine Kinase (PTK) Genes and Their Role in Abiotic Stress Response in Rice

Plants ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 664
Author(s):  
Allimuthu Elangovan ◽  
Monika Dalal ◽  
Gopinathan Kumar Krishna ◽  
Sellathdurai Devika ◽  
Ranjeet Ranjan Kumar ◽  
...  
Keyword(s):  
Abiotic Stress ◽  
Stress Response ◽  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Rice Genome ◽  
Kinase Domain ◽  
Tyrosine Kinase Domain ◽  
Protein Tyrosine ◽  
Dual Specificity ◽  
Submergence Stress

Tyrosine phosphorylation constitutes up to 5% of the total phophoproteome. However, only limited studies are available on protein tyrosine kinases (PTKs) that catalyze protein tyrosine phosphorylation in plants. In this study, domain analysis of the 27 annotated PTK genes in rice genome led to the identification of 18 PTKs with tyrosine kinase domain. The kinase domain of rice PTKs shared high homology with that of dual specificity kinase BRASSINOSTEROID-INSENSITIVE 1 (BRI1) of Arabidopsis. In phylogenetic analysis, rice PTKs clustered with receptor-like cytoplasmic kinases-VII (RLCKs-VII) of Arabidopsis. mRNAseq analysis using Genevestigator revealed that rice PTKs except PTK9 and PTK16 express at moderate to high level in most tissues. PTK16 expression was highly abundant in panicle at flowering stage. mRNAseq data analysis led to the identification of drought, heat, salt, and submergence stress regulated PTK genes in rice. PTK14 was upregulated under all stresses. qRT-PCR analysis also showed that all PTKs except PTK10 were significantly upregulated in root under osmotic stress. Tissue specificity and abiotic stress mediated differential regulation of PTKs suggest their potential role in development and stress response of rice. The candidate dual specificity PTKs identified in this study paves way for molecular analysis of tyrosine phosphorylation in rice.

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