Degrasyn-Induced Trafficking of BCR-ABL as a Novel Mechanism of Kinase Inactivation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1003-1003
Author(s):  
Nicholas Donato ◽  
Vaibhav Kapuria ◽  
Hanshi Sun ◽  
David Maxwell ◽  
Geoffrey Bartholomeusz ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Point Mutations ◽  
Binding Pocket ◽  
Atp Binding ◽  
Wild Type ◽  
Post Translational Modification ◽  
Oncogenic Signaling ◽  
Imatinib Resistance ◽  
Unique Mechanism

Abstract Inhibitors that inactivate specific tyrosine kinases have proven to be a very effective form of therapy of many leukemias and hematopoetic disorders. Most inhibitors function by competing for the ATP-binding pocket or by preventing association with protein substrates. However, clinical and molecular studies have shown that small changes in the structure of the target kinase (point mutations, post-translational modification) affect inhibitor binding affinities, resulting in resistance to this class of inhibitor. Therefore, development of agents that reduce the activity of leukemogenic kinases through alternate mechanisms are of great interest. We previously described a novel class of compounds, termed degrasyns, which reduced BCR-ABL (and Jak2) kinase activity through a unique mechanism. Degrasyn treatment of CML or BaF3 cells expressing wild-type or mutant (T315I) BCR-ABL resulted in a reduction of cytoplasmic BCR-ABL protein levels and loss of downstream signaling without a direct effect on BCR-ABL enzymatic activity. Biochemical studies demonstrated that degrasyn induced a rapid translocation of BCR-ABL from the cytosol to the cytoskeletal fraction (complete within 60 min) and this event correlated with loss of BCR-ABL signaling and initiation of apoptosis. A GFP-fusion protein composed of wild-type or T315I mutant BCR-ABL demonstrated that degrasyn induced high density translocation to the cytoskeletal fraction, as determined by direct fluorescence imaging. Translocation was specific for BCR-ABL and not other kinases (except Jak2) or signaling proteins. Translocation correlated with degrasyn-mediated tyrosine phosphorylation of a subset of specific proteins, including the tyrosine kinase Lyn, in the insoluble cellular fraction. CML cells expressing high levels of Lyn were more sensitive to degrasyn-mediated apoptosis suggesting that Lyn plays a role in degrasyn activity. Together these results suggest that degrasyn inactivates BCR-ABL by inducing its translocation to a cellular compartment that prevents its participation in oncogenic signaling. Degrasyn-induced BCR-ABL translocation was not effected by mutations that block dasatinib and imatinib activity and shows greater activity against some forms of imatinib resistance (Lyn overexpression). Animal studies demonstrated that degrasyn has anti-leukemic activity and functions through a unique mechanism of action. Development of inhibitors with this mode of action may be of significance for CML patients that fail therapy with ATP-binding pocket-directed tyrosine kinase inhibitors.

Targeting the KIT Activating Switch Control Pocket: A Novel Mechanism to Inhibit Mast Cell Activation and KIT D816V Neoplastic Mast Cell Proliferation

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1740-1740
Author(s):  
Geethani Bandara ◽  
Yun Bai ◽  
Eunice Ching Chan ◽  
Irina Maric ◽  
Olga Simakova ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Mast Cell ◽  
Mast Cells ◽  
Tyrosine Kinase ◽  
Cell Activation ◽  
Binding Pocket ◽  
Mast Cell Activation ◽  
Atp Binding ◽  
Wild Type ◽  
Kit D816v

Abstract Abstract 1740 KIT is a receptor protein tyrosine kinase which undergoes dimerization, autophysophyralation, and activation upon binding of its ligand, stem cell factor (SCF). Such activation is critical for the growth, differentiation and survival of mast cells. By enhancing Fcƒ'RI-mediated degranulation and inducing chemotaxis, KIT also contributes to mast cell functional responses. Targeting KIT is thus an attractive approach for the management of proliferative mast cell disorders. Activating mutations in the KIT tyrosine kinase domain, most notably KIT D816V, are commonly observed in the myeloproliferative disorder, systemic mastocytosis. The D816 mutation is located within the activating-switch component of KIT. These D816 mutations render the activating-switch of KIT constitutively active and capable of binding into the cognate switch pocket of KIT. Such aberrant switch function aggressively fluxes KIT to a catalytically active conformation. Switch pocket inhibitors block access to this switch pocket, thus inhibiting the ability of the mutationally activated D816 KIT mutants to adopt active conformations. Most tyrosine kinase inhibitors in clinical development target the KIT ATP binding pocket whose structure is often conserved among kinases resulting in off-target inhibition. In contrast, switch pockets are more unique among kinases. Utilizing rational drug design, the diversity in switch pockets that KIT uses to switch between the inactive and active conformations may be targeted to fine-tune selectivity of an inhibitor against the rest of the kinome. In this study, two such KIT “switch pocket” (SP) inhibitors, DP-2976 and DCC-2618, were examined for their effects on KIT activation, mast cell activation and mast cell proliferation; in comparison to the known ATP binding pocket inhibitors, imatinib and PKC412. To explore the effects of the SP inhibitors on KIT activity, wild type KIT and KIT D816V were expressed in 293T cells and their KIT autophosphorylation status assessed. DP-2976 and DCC-2618 blocked the inherent autophosphorylation of both wild type and KIT D816V in the transfected cells in the nanomolar range. Similar results were observed on constiutively active KIT in HMC 1.1 and 1.2 cells; and in CD34+ derived human mast cells in which KIT phosphorylation was induced by exposure to SCF (10 ng/ml). SP inhibitors had minimal effects on FcεRI mediated mast cell degranulation. They did completely block the SCF potentiation of FceRI-mediated degranulation. In contrast, PKC 412 inhibited degrangulation in an SCF independent manner suggesting a more generalized kinase targeting. To assess the anti-neoplastic activity of the SP inhibitors, they were next examined in HMC1.1 and HMC1.2 cell proliferation assays. Both DP-2976 and DCC-2618 significantly reduced cell numbers through induction of apoptosis with low nanomolar IC50 values. To examine whether such observations translate into clinical efficacy, ex vivo studies were performed. The survival of primary bone marrow mast cells from mastocytosis patients was significantly reduced when exposed to the SP inhibitors; a reduction comparable to PKC412. Overall, DP-2976 and DCC-2618 “switch pocket” inhibitors offer a novel and potent KIT inhibition profile whose selectivity and dual suppression of SCF enhanced mast cell activation and KIT D816V neoplastic proliferation may provide significant therapeutic benefits. Disclosures: Wise: Deciphera Pharmaceuticals LLC: Employment. Flynn:Deciphera Pharmaceuticals LLC: Employment, Equity Ownership.

Activation of pp60(src) is critical for stretch-induced orienting response in fibroblasts

1999 ◽  
Vol 112 (9) ◽  
pp. 1365-1373 ◽  
Author(s):  
X. Sai ◽  
K. Naruse ◽  
M. Sokabe
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Time Course ◽  
Kinase Inhibitor ◽  
Orienting Response ◽  
Cyclic Stretch ◽  
Wild Type ◽  
Herbimycin A ◽  
Molecular Masses

When subjected to uni-axial cyclic stretch (120% in length, 1 Hz), fibroblasts (3Y1) aligned perpendicular to the stretch axis in a couple of hours. Concomitantly with this orienting response, protein tyrosine phosphorylation of cellular proteins (molecular masses of approximately 70 kDa and 120–130 kDa) increased and peaked at 30 minutes. Immuno-precipitation experiments revealed that paxillin, pp125(FAK), and pp130(CAS) were included in the 70 kDa, and 120–130 kDa bands, respectively. Treatment of the cells with herbimycin A, a tyrosine kinase inhibitor, suppressed the stretch induced tyrosine phosphorylation and the orienting response suggesting that certain tyrosine kinases are activated by stretch. We focused on pp60(src), the most abundant tyrosine kinase in fibroblasts. The kinase activity of pp60(src) increased and peaked at 20 minutes after the onset of cyclic stretch. Treatment of the cells with an anti-sense S-oligodeoxynucleotide (S-ODN) against pp60(src), but not the sense S-ODN, inhibited the stretch induced tyrosine phosphorylation and the orienting response. To further confirm the involvement of pp60(src), we performed the same sets of experiments using c-src-transformed 3Y1 (c-src-3Y1) fibroblasts. Cyclic stretch induced a similar orienting response in c-src-3Y1 to that in wild-type 3Y1, but with a significantly faster rate. The time course of the stretch-induced tyrosine phosphorylation was also much faster in c-src-3Y1 than in 3Y1 fibroblasts. These results strongly suggest that cyclic stretch induces the activation of pp60(src) and that pp60(src) is indispensable for the tyrosine phosphorylation of pp130(CAS), pp125(FAK) and paxillin followed by the orienting response in 3Y1 fibroblasts.

Anaplastic Lymphoma Kinase Is Activated through the Pleiotrophin/Receptor Protein Tyrosine Phosphatase (RPTP)β/ζ Signaling Pathway: A Unique Mechanism of Activation of Receptor Protein Tyrosine Kinases.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4355-4355
Author(s):  
Pablo Perez-Pinera ◽  
Wei Zhang ◽  
Zhaoyi Wang ◽  
James R. Berenson ◽  
Thomas F. Deuel
Keyword(s):  
Tyrosine Kinase ◽  
Signaling Pathway ◽  
Tyrosine Phosphorylation ◽  
Protein Tyrosine Phosphatase ◽  
Tyrosine Kinases ◽  
Tyrosine Phosphatase ◽  
Receptor Protein ◽  
Protein Tyrosine ◽  
Anaplastic Lymphoma ◽  
Unique Mechanism

Abstract Anaplastic Lymphoma Kinase (ALK) is a receptor-type transmembrane tyrosine kinase (RTK) of the insulin receptor superfamily that structurally is most closely related to leukocyte tyrosine kinase. It was first discovered as a chimeric protein (NPM-ALK) of nucleophosmin and the C-terminal (kinase) domain of ALK in anaplastic large cell lymphomas (ALCL). NPM-ALK is constitutively active and generates the oncogenic signals that are the pathogenic mechanisms of these highly malignant cancers. The full-length ALK also is believed to have an important role in the pathogenesis of other human malignancies, since its expression is found in rhabdomyosarcomas, neuroblastomas, neuroectodermal tumors, glioblastomas, breast carcinomas, and melanomas. Recently it was proposed that pleiotrophin (PTN the protein, Ptn the gene) is the ligand that stimulates ALK to transduce signals to activate downstream targets. However, this proposal contrasted with earlier studies that demonstrated Receptor Protein Tyrosine Phosphatase (RPTP)β/ζ is the functional receptor for PTN. PTN was shown to inactivate RPTPβ/ζ and thereby permit the activity of different tyrosine kinases to increase tyrosine phosphorylation of the substrates of RPTPβ/ζ at the sites that are dephosphorylated by RPTPβ/ζ in cells not stimulated by PTN. Subsequent studies identified β-catenin, β-adducin, Fyn, GIT1/Cat-1, P190RhoGAP, and histone deacetylase 2 (HDAC-2) as downstream targets of the PTN/RPTPβ/ζ signaling pathway and demonstrated that their levels of tyrosine phosphorylation increase in PTN-stimulated cells. This diversity of PTN-regulated targets is one basis for the pleiotrophic activities of PTN. We now demonstrate that tyrosine phosphorylation of ALK is increased in PTN-stimulated cells through the PTN/RPTPβ/ζ signaling pathway. It is furthermore shown that ALK is activated in PTN-stimulated cells when it is expressed in cells without its extracellular domain, that β-catenin is a substrate of ALK, that the tyrosine phosphorylation site in β-catenin phosphorylated by ALK is the same site dephosphorylated by RPTPβ/ζ, and that PTN-stimulated tyrosine phosphorylation of β-catenin requires expression of ALK. The data suggest a unique mechanism to activate ALK; the data support a mechanism in which β-catenin is phosphorylated in tyrosine through the coordinated inactivation of RPTPβ/ζ, the activation of the tyrosine kinase activity of ALK, and the phosphorylation of β-catenin by ALK at the same site regulated by RPTPβ/ζ in PTN-stimulated cells. Since PTN often is inappropriately expressed in the same malignancies that express ALK, the data suggest a mechanism through which ALK signaling may contribute to those malignancies that express full length ALK through the activity of PTN to signal constitutively the same pathways as NPM-ALK in ALCL.

The Novel Inhibitor PLX3397 Effectively Inhibits FLT3-Mutant AML,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3632-3632 ◽  
Author(s):  
Elizabeth Burton ◽  
Bernice Wong ◽  
Jiazhong Zhang ◽  
Brian West ◽  
Gideon Bollag ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Tumor Regression ◽  
Wild Type ◽  
Gut Peptides ◽  
Cellular Assays ◽  
Juxtamembrane Region ◽  
Clear Dose Response ◽  
Major Subset ◽  
Subcutaneous Xenograft

Abstract Abstract 3632 Mutations of FLT3 (FMS-like tyrosine kinase 3) are frequently present in acute myeloid leukemias (AML) and associated with poor prognosis. Such mutations alter the conformation and constitutively activate the FLT3 tyrosine kinase. PLX3397 is a novel, orally active, selective small molecule inhibitor of the FLT3-mutant as well as FMS and KIT kinases. The high selectivity of PLX3397 has been demonstrated in biochemical assays against a panel of over 200 recombinant kinases. FLT3, FMS and KIT are significantly inhibited at low nanomolar concentrations. The majority of kinases screened were not inhibited by PLX3397 (IC50=>10 μM) and the only other kinase with significant sub-micromolar inhibition was KDR. PLX3397 exhibits favorable pharmaceutical properties and demonstrated minimal off-target activity when tested in a broad array of 71 targets in 8 families (Neurotransmitter-related, Steroids, Ion Channels, Nitric Oxide, Prostaglandins, Growth Factors, Brain/Gut Peptides, and Enzymes). In cellular assays PLX3397 effectively inhibited ligand-stimulated autophosphorylation of the endogenous receptor tyrosine kinases FMS (IC50=20 nM) and KIT (IC50=120 nM), but not ligand-stimulated wild-type FLT3 (IC50=1.7 μM). In SEMK2 cells, which over-express wild type FLT3, PLX3397 inhibited FLT3 autophosphorylation with an IC50 of 240 nM. The phosphorylation of the activated FLT3-ITD protein in MV-4-11 and MOLM-14 cells harboring mutations (FLT3-ITD) of the juxtamembrane region of FLT3 was efficiently inhibited (IC50 26 nM and 30 nM respectively), indicating a possible application in this major subset of FLT3 mutated AML. In parallel with inhibition of FLT3-mutant autophosphorylation, the phosphorylation of known downstream effectors (STAT5, AKT, MEK, and ERK) is also inhibited. However, no inhibition was observed for KDR phosphorylation. PLX3397 potently inhibited of mutant-FLT3 driven proliferation in both MV-4-11 and MOLM-14 AML cells in culture (IC50=100–200 nM), but was less effective against HL60 cells expressing wild-type FLT3. MV-4-11 cells grown as subcutaneous xenograft tumors in mice were also highly sensitive to PLX3397 dosed orally at 10 or 30 mg/kg qd, with tumor regression observed at the higher dose. Primary samples collected from AML patients at relapse were tested in culture. While no significant effects were seen in FLT3 wild type samples below 1 μM, a clear dose response to PLX3397 was observed in samples with FLT3-ITD mutations. Because the estimated protein binding of PLX3397 in plasma is >99%, we also tested the drug against the MV-4-11 cell line in plasma and found the IC50 for inhibition of autophosphorylation to be 2.8 μM. In humans trough levels of PLX3397 (10 μM) capable of inhibiting FLT3-mutants can be safely achieved. A Phase I/II study is planned to evaluate the safety and explore the efficacy of PLX3397 in patients with FLT3-mutant- AML. Disclosures: Burton: Plexxikon Inc.: Employment. Wong:Plexxikon Inc.: Employment. Zhang:Plexxikon Inc.: Employment. West:Plexxikon Inc.: Employment. Bollag:Plexxikon Inc.: Employment. Habets:Plexxikon Inc.: Employment. Nguyen:Plexxikon Inc.: Employment. Levis:Plexxikon Inc.: Honoraria.

Induction of autophagy across multiple tumor types through irreversible HER tyrosine kinase blockade.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e13517-e13517
Author(s):  
William Rayford Gwin ◽  
Leihua Liu ◽  
Sumin Zhao ◽  
Wenle Xia ◽  
Neil Spector
Keyword(s):  
Prostate Cancer ◽  
Cell Cycle ◽  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Cellular Proliferation ◽  
Kinase Inhibitor ◽  
Significant Inhibition ◽  
Wild Type ◽  
Multiple Tumor ◽  
Mtorc1 Signaling

e13517 Background: Human epidermal growth factor receptor (HER) receptor tyrosine kinases play a key role in solid tumor oncogenesis. Despite broad expression of HER receptors in solid tumors, HER targeted therapies have not shown significant improvement in survival, calling into question the value of wild-type HER receptors as therapeutic targets. Here we found that an irreversible pan-HER tyrosine kinase inhibitor (TKI), neratinib, but not similar HER TKIs, induced morphologic changes in ovarian, TNBC, and prostate cancer cell lines consistent with induction of autophagy. Methods: SKOV3 (ovarian), OVCAR8 (ovarian), HBL-100 (TNBC), and LAPC4 (prostate) cancer cells were treated with lapatinib, gefitinib, CI-1033, afatinib, and neratinib (0.5mM-2.5mM). The activation state of HER2, EGFR, HER3, Akt, Erk, p70S6, 4EBP1, and Ulk1 was determined by Western blot analysis (WB) at various time points of neratinib treatment. LC3 was analyzed by immunofluorescence (IF) microscopy and WB. Analysis of proliferation, apoptosis, and cell cycle were performed using WST-1, annexin V, and PI staining, respectively. Results: Neratinib, but not similar HER TKIs, induced marked cytoplasmic vacuolization in tumors. The conversion of LC3-I to LC3-II in neratinib-treated cells was consistent with induction of autophagy. Moreover, PI3K/Akt, MAPK/Erk1/2 and mTORC1 signaling cascades were inhibited in neratinib-treated cells, and were associated with the inhibition of phospho-Ulk1, a key step in autophagy initiation. Treatment with neratinib alone resulted in G1 cell cycle arrest. Importantly, the combination of neratinib and chloroquine, an autophagy inhibitor, induced a statistically significant inhibition of cellular proliferation (p <0.01) and increased apoptosis compared to treatment with either drug alone. Conclusions: Our data suggest that more effective inhibition of wild-type HER receptors, can lead to mTORC1 inhibition, which in turn triggers autophagy. Here, autophagy appears to protect cells rather than inducing apoptosis. Consequently, targeting both HER receptors and autophagy represents an attractive therapeutic strategy to treat tumors expressing wild-type HER receptors.

scholarly journals Point Mutations in Human β Cardiac Myosin Heavy Chain Have Differential Effects on Sarcomeric Structure and Assembly: An ATP Binding Site Change Disrupts Both Thick and Thin Filaments, Whereas Hypertrophic Cardiomyopathy Mutations Display Normal Assembly

1997 ◽  
Vol 137 (1) ◽  
pp. 131-140 ◽  
Author(s):  
K. David Becker ◽  
Kim R. Gottshall ◽  
Reed Hickey ◽  
Jean-Claude Perriard ◽  
Kenneth R. Chien
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Subcellular Localization ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Point Mutations ◽  
Neonatal Rat ◽  
Atp Binding ◽  
Cardiac Myosin ◽  
Wild Type ◽  
Thick Filaments

Hypertrophic cardiomyopathy is a human heart disease characterized by increased ventricular mass, focal areas of fibrosis, myocyte, and myofibrillar disorganization. This genetically dominant disease can be caused by mutations in any one of several contractile proteins, including β cardiac myosin heavy chain (βMHC). To determine whether point mutations in human βMHC have direct effects on interfering with filament assembly and sarcomeric structure, full-length wild-type and mutant human βMHC cDNAs were cloned and expressed in primary cultures of neonatal rat ventricular cardiomyocytes (NRC) under conditions that promote myofibrillogenesis. A lysine to arginine change at amino acid 184 in the consensus ATP binding sequence of human βMHC resulted in abnormal subcellular localization and disrupted both thick and thin filament structure in transfected NRC. Diffuse βMHC K184R protein appeared to colocalize with actin throughout the myocyte, suggesting a tight interaction of these two proteins. Human βMHC with S472V mutation assembled normally into thick filaments and did not affect sarcomeric structure. Two mutant myosins previously described as causing human hypertrophic cardiomyopathy, R249Q and R403Q, were competent to assemble into thick filaments producing myofibrils with well defined I bands, A bands, and H zones. Coexpression and detection of wild-type βMHC and either R249Q or R403Q proteins in the same myocyte showed these proteins are equally able to assemble into the sarcomere and provided no discernible differences in subcellular localization. Thus, human βMHC R249Q and R403Q mutant proteins were readily incorporated into NRC sarcomeres and did not disrupt myofilament formation. This study indicates that the phenotype of myofibrillar disarray seen in HCM patients which harbor either of these two mutations may not be directly due to the failure of the mutant myosin heavy chain protein to assemble and form normal sarcomeres, but may rather be a secondary effect possibly resulting from the chronic stress of decreased βMHC function.

scholarly journals CFTR: the nucleotide binding folds regulate the accessibility and stability of the activated state.

1996 ◽  
Vol 107 (1) ◽  
pp. 103-119 ◽  
Author(s):  
D J Wilkinson ◽  
M K Mansoura ◽  
P Y Watson ◽  
L S Smit ◽  
F S Collins ◽  
...  
Keyword(s):  
Aspartic Acid ◽  
Atp Hydrolysis ◽  
Nucleotide Binding ◽  
Binding Pocket ◽  
Active State ◽  
Slow Phase ◽  
Atp Binding ◽  
Wild Type ◽  
Rate Analysis ◽  
Activated State

The functional roles of the two nucleotide binding folds, NBF1 and NBF2, in the activation of the cystic fibrosis transmembrane conductance regulator (CFTR) were investigated by measuring the rates of activation and deactivation of CFTR Cl- conductance in Xenopus oocytes. Activation of wild-type CFTR in response to application of forskolin and 3-isobutyl-1-methylxanthine (IBMX) was described by a single exponential. Deactivation after washout of the cocktail consisted of two phases: an initial slow phase, described by a latency, and an exponential decline. Rate analysis of CFTR variants bearing analogous mutations in NBF1 and NBF2 permitted us to characterize amino acid substitutions according to their effects on the accessibility and stability of the active state. Access to the active state was very sensitive to substitutions for the invariant glycine (G551) in NBF1, where mutations to alanine (A), serine (S), or aspartic acid (D) reduced the apparent on rate by more than tenfold. The analogous substitutions in NBF2 (G1349) also reduced the on rate, by twofold to 10-fold, but substantially destabilized the active state as well, as judged by increased deactivation rates. In the putative ATP-binding pocket of either NBF, substitution of alanine, glutamine (Q), or arginine (R) for the invariant lysine (K464 or K1250) reduced the on rate similarly, by two- to fourfold. In contrast, these analogous substitutions produced opposite effects on the deactivation rate. NBF1 mutations destabilized the active state, whereas the analogous substitutions in NBF2 stabilized the active state such that activation was prolonged compared with that seen with wild-type CFTR. Substitution of asparagine (N) for a highly conserved aspartic acid (D572) in the ATP-binding pocket of NBF1 dramatically slowed the on rate and destabilized the active state. In contrast, the analogous substitution in NBF2 (D1370N) did not appreciably affect the on rate and markedly stabilized the active state. These results are consistent with a hypothesis for CFTR activation that invokes the binding and hydrolysis of ATP at NBF1 as a crucial step in activation, while at NBF2, ATP binding enhances access to the active state, but the rate of ATP hydrolysis controls the duration of the active state. The relatively slow time courses for activation and deactivation suggest that slow processes modulate ATP-dependent gating.

New insights into the mechanisms of hematopoietic cell transformation by activated receptor tyrosine kinases

Blood ◽  
2010 ◽  
Vol 116 (14) ◽  
pp. 2429-2437 ◽  
Author(s):  
Federica Toffalini ◽  
Jean-Baptiste Demoulin
Keyword(s):  
Growth Factor ◽  
Tyrosine Kinase ◽  
Tyrosine Kinases ◽  
Kinase Inhibitors ◽  
Cell Transformation ◽  
Chromosomal Rearrangements ◽  
Point Mutations ◽  
Large Cell ◽  
Kinase Domain ◽  
Leukemic Cells

Abstract A large number of alterations in genes encoding receptor tyrosine kinase (RTK), namely FLT3, c-KIT, platelet-derived growth factor (PDGF) receptors, fibroblast growth factor (FGF) receptors, and the anaplastic large cell lymphoma kinase (ALK), have been found in hematopoietic malignancies. They have drawn much attention after the development of tyrosine kinase inhibitors. RTK gene alterations include point mutations and gene fusions that result from chromosomal rearrangements. In both cases, they activate the kinase domain in the absence of ligand, producing a permanent signal for cell proliferation. Recently, this simple model has been refined. First, by contrast to wild-type RTK, many mutated RTK do not seem to signal from the plasma membrane, but from various locations inside the cell. Second, their signal transduction properties are altered: the pathways that are crucial for cell transformation, such as signal transducer and activator of transcription (STAT) factors, do not necessarily contribute to the physiologic functions of these receptors. Finally, different mechanisms prevent the termination of the signal, which normally occurs through receptor ubiquitination and degradation. Several mutations inactivating CBL, a key RTK E3 ubiquitin ligase, have been recently described. In this review, we discuss the possible links among RTK trafficking, signaling, and degradation in leukemic cells.

High frequency of point mutations clustered within the adenosine triphosphate–binding region of BCR/ABL in patients with chronic myeloid leukemia or Ph-positive acute lymphoblastic leukemia who develop imatinib (STI571) resistance

Blood ◽  
2002 ◽  
Vol 99 (9) ◽  
pp. 3472-3475 ◽  
Author(s):  
Susan Branford ◽  
Zbigniew Rudzki ◽  
Sonya Walsh ◽  
Andrew Grigg ◽  
Chris Arthur ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Chronic Myeloid Leukemia ◽  
Adenosine Triphosphate ◽  
High Frequency ◽  
Myeloid Leukemia ◽  
Lymphoblastic Leukemia ◽  
Point Mutations ◽  
Atp Binding ◽  
Binding Region ◽  
Imatinib Resistance

Abstract Point mutations were found in the adenosine triphosphate (ATP) binding region of BCR/ABL in 12 of 18 patients with chronic myeloid leukemia (CML) or Ph-positive acute lymphoblastic leukemia (Ph+ ALL) and imatinib resistance (defined as loss of established hematologic response), but they were found in only 1 of 10 patients with CML with imatinib refractoriness (failure to achieve cytogenetic response). In 10 of 10 patients for whom samples were available, the mutation was not detected before the initiation of imatinib therapy. Three mutations (T315I, Y253H, and F317L present in 3, 1, and 1 patients, respectively) have a predicted role in abrogating imatinib binding to BCR/ABL, whereas 3 other mutations (E255K, G250E, and M351T, present in 4, 2, and 2 patients, respectively) do not. Thus we confirm a high frequency of mutations clustered within the ATP-binding region of BCR/ABL in resistant patients. Screening may allow intervention before relapse by identifying emerging mutations with defined impacts on imatinib binding. Certain mutations may respond to higher doses of imatinib, whereas other mutations may mandate switching to another therapeutic strategy.

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