Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The 9;22 chromosomal translocation characteristic of CML results in a fused bcr/abl gene and an abnormal fusion protein, p210bcr/abl. Relative to normal c-abl, p210bc1/abl has elevated tyrosine kinase activity that is essential for its transforming activity. We recently reported a prominent 62 kDa GAP-associated P-tyr protein and five additional consistent but less prominent P-tyr proteins as well as five more minor P-tyr proteins that are constitutively tyrosine phosphorylated in primary primitive lineage negative (lin-) chronic phase CML blasts but not in comparable primary lin- normal blasts. The GAP-associated p62 protein has now been purified, sequenced and its gene has been cloned; it is a previously unidentified protein and is currently being characterized. In analyzing P-tyr proteins in primary lin- normal blasts in response to various hematopoietic cytokines, we found a striking similarity in the tyrosine phosphorylation of four major and three minor proteins after stimulation with c-kit ligand (KL) and the P-tyr proteins that are constitutively phosphorylated in primary primitive lin- chronic phase CML blasts. Other cytokines tested (ie GM-CSF, G-CSF, IL-3, FLT3 ligand, TPO, EPO) were much less active or stimulated phosphorylation of other proteins. KL/c-kit and bcr/abl have some similar activities including enhancing survival and expansion of hematopoietic progenitor cells, probably acting primarily on early progenitors at the time of lineage commitment rather than on self-renewing stem cells. Activation of growth factor receptors promote a cascade of protein phosphorylations that can ultimately result in a wide range of cellular responses. Sustained activation of discrete signaling pathways in some types of cells results in differentiation, whereas transient activation instead causes a proliferative response; in other cell types, the converse is true. It may be postulated that stem cells and primitive progenitors are at a particularly susceptible stage of development that renders them especially responsive to sustained bcr/abl-induced phorphorylation of a number of signaling proteins that are components of critical regulatory pathways, including c-kit. The affected pathways control and coordinate multiple diverse cell processes including proliferation, differentiation, maturation and apoptosis, processes that are normally tightly regulated and integrated. Perturbation of these key pathways in primitive progenitors would be expected to seriously disrupt orderly hematopoiesis and could also explain the multiple subtle pleiotropic biological abnormalities characteristically observed in later maturing CML compartments that we have collectively designated 'discordant maturation'. The true situation is undoubtedly very complex and involves interaction of multiple cytokines and signaling pathways that we are now trying to define. Constitutive downstream activation of critical pathways in susceptible early progenitors that normally require KL or other factors for activation could explain most if not all features of the disease.
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PMID:New understanding of the pathogenesis of CML: a prototype of early neoplasia. 952 44

FLT3 is a member of receptor tyrosine kinases expressed in leukemia cells, as well as in hematopoietic stem cells. Recently, a somatic alteration of the FLT3 gene was found in acute myeloid leukemia, as an internal tandem duplication (FLT3/ITD) which caused elongation of the juxtamembrane (JM) domain of FLT3. Here we characterized the FLT3/ITD and investigated its clinical significance in acute promyelocytic leukemia (APL). Seventy-four newly diagnosed patients with APL, who were treated with the same protocol in a multi-institutional study, were studied for the FLT3/ITD. Genomic and message sequences of the FLT3 gene were amplified by means of polymerase chain reaction (PCR), and elongated PCR products were sequenced. Fifteen patients (20.3%) had FLT3/ITD, all of which were transcribed in frame. Location of the duplicated fragments (six to 30 amino acids) varied from patient to patient. However, they always contained either Y591 or Y599, but the tyrosine kinase domain was not significantly affected. This finding implied that signal transduction of FLT3 is amplified by the duplication. Clinically, the presence of FLT3/ITD was related to high peripheral white blood cell counts as well as peripheral leukemia cell counts (P < 0.0001), high LDH level (P = 0.04), and low fibrinogen concentration (P = 0.04). These data suggest that FLT3/ITD plays a significant role in progression of APL.
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PMID:Internal tandem duplication of FLT3 associated with leukocytosis in acute promyelocytic leukemia. Leukemia Study Group of the Ministry of Health and Welfare (Kohseisho). 930 96

In this study, we examined a large number of patients to clarify the distribution and frequency of a recently described FLT3 tandem duplication among hematopoietic malignancies, including 112 acute myelocytic leukemia (AML), 55 acute lymphoblastic leukemia (ALL), 37 myelodysplastic syndrome (MDS), 20 chronic myelogenous leukemia (CML), 30 non-Hodgkin's lymphoma (NHL), 14 adult T cell leukemia, 15 chronic lymphocytic leukemia (CLL) and 38 multiple myeloma (MM). We also evaluated 71 cell lines derived from 11 AML, 31 ALL, two hairy cell leukemia, three acute unclassified leukemia, 10 CML, 12 NHL including six Burkitt's lymphoma, and two MM. Using genomic PCR of exon 11 coding for the juxtamembrane (JM) domain and first amino acids of the 5'-tyrosine kinase (TK) domain, this length mutation was found only in AML (22/112, 20%) and MDS (1/37). According to the FAB subclassification, they were 5/18 (28%) of M1, 4/29 (14%) of M2, 3/17 (18%) of M3, 6/24 (25%) of M4, 4/20 (20%) of M5 and 1/9 of refractory anemia with excess of blast in transformation. In the various cell lines examined, this abnormality was determined in only one derived from AML and never found in other hematological malignancies. The sequence analysis of the abnormal PCR products revealed that 23 of 24 showed internal tandem duplication with or without insertion of nucleotides. In one AML, insertion and deletion without duplication was determined. All 24 lengthened sequences were in-frame. Duplication takes place in the sequence coding for the JM domain and leaves the TK domain intact. In conclusion, we emphasize that the length mutation of FLT3 at JM/TK-I domains were restricted to AML and MDS. Since all these mutations resulted in in-frame, this abnormality might function for the proliferation of leukemic cells.
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PMID:Internal tandem duplication of the FLT3 gene is preferentially seen in acute myeloid leukemia and myelodysplastic syndrome among various hematological malignancies. A study on a large series of patients and cell lines. 932 77

There are relatively few monoclonal antibodies (mAb) that have been characterized for their applicability in studies on the immune system of various nonhuman primates. In the present study, we identified a large number of mAb that can be used in future immunological studies in three different nonhuman primates, i.e., chimpanzees, rhesus macaques, and squirrel monkeys. The reactivity of 161 anti-human mAb to T-cell antigens and cytokine receptors were tested on peripheral blood mononuclear cells (PBMC) from the three primate species by flow cytometric analysis. A total of 105 (65%), 73 (45%), and 68 (42%) antibodies reacted with PBMC from chimpanzees, rhesus macaques, and squirrel monkeys, respectively. Out of the 161 mAb, 38 reacted with all three species and 112 reacted with one or two of the species. No specific reaction was observed with mAb to receptors to GM-CSF, 4-1BB, FLT3, FLX2, common beta-chain, IL-1 (type I receptor), and IL-8.
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PMID:Flow cytometric analysis on reactivity of human T lymphocyte-specific and cytokine-receptor-specific antibodies with peripheral blood mononuclear cells of chimpanzee (Pan troglodytes), rhesus macaque (Macaca mulatta), and squirrel monkey (Saimiri sciureus) 987 63

Human hematopoiesis can be supported in beige/nude/ XID (bnx) mice by coinjection of human bone marrow stromal cells engineered to secrete human interleukin 3 (HuIL-3). The major limitation is a total absence of human B cell development in the mice, which could be due to supraphysiological levels of HuIL-3 in the circulation. In an effort to obtain human B lymphoid, as well as T lymphoid and myeloid cell development in the mice, CD34+ cells were coinjected with human marrow stromal cells engineered to secrete human IL-2, IL-7, stem cell factor or FLT3 ligand, +/- IL-3. No single factor other than IL-3 supported sustained human hematopoiesis in the mice, although cytokines were expressed for four to six months post-transplantation. Production of both HuIL-3 and IL-7 in the mice supported extrathymic development of human T lymphocytes, but no B cells, myeloid cells, or clonogenic progenitors were detected. Human B cells were not produced from CD34+ cells in the bnx mice under any condition tested. Another limitation to the bnx/Hu system is a lack of maturation of human red blood cells, although BFU-E are maintained. Stromal cells secreting human erythropoietin and IL-3 were cotransplanted into mice with HuCD34+ cells and an increase in hematocrit from 40%-45% to 80%-85% resulted, with production of human and murine red blood cells. Unfortunately, all mice (n = 9) suffered strokes, displayed paralysis and died within three weeks. The bnx/Hu cotransplantation model provides an interesting system in which to study human hematopoietic cell differentiation under the influence of various cytokines.
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PMID:Long-term cytokine production from engineered primary human stromal cells influences human hematopoiesis in an in vivo xenograft model. 940 57

Stroma-supported long-term cultures (LTC) allow estimation of stem cell quality by simultaneous enumeration of hematopoietic stem cell (HSC) frequencies in a graft using the cobblestone area forming cell (CAFC) assay, and the ability of the graft to generate progenitors in flask LTC (LTC-CFC). We have recently observed that the number and quality of mobilized peripheral blood stem cells (PBSC) was low in patients having received multiple rounds of chemotherapy. Moreover, grafts with low numbers of HSC and poor HSC quality had a high probability to cause graft failure upon their autologous infusion. Because ex vivo culture of stem cells has been suggested to present an attractive tool to improve hematological recovery or reduce graft size, we have studied the possibility that such propagation may affect stem cell quality. In order to do so, we have assessed the recovery of different stem cell subsets in CD34+ PBSC after a 7-day serum-free liquid culture using CAFC and LTC-CFC assays. A numerical expansion of stem cell subsets was observed in the presence of interleukin-3 (IL-3), stem cell factor, and IL-6, while stroma-contact, stromal soluble factors, or combined addition of FLT3-ligand and thrombopoietin improved this parameter. In contrast, ex vivo culture severely reduced the ability of the graft to produce progenitors in LTC while stromal soluble factors partly abrogated this quality loss. The best conservation of graft quality was observed when the PBSC were cultured in stroma-contact. These data suggest that ex vivo propagation of PBSC may allow numerical expansion of various stem cell subsets, however, at the expense of their quality. In addition, cytokine-driven PBSC cultures require stroma for optimal maintenance of graft quality.
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PMID:Stroma-contact prevents loss of hematopoietic stem cell quality during ex vivo expansion of CD34+ mobilized peripheral blood stem cells. 941 74

Successful treatment of melanoma and lymphoma may result from the induction of specific antitumor immunity. Dendritic cells (DCs) are powerful antigen-presenting cells and show a remarkable capacity to stimulate antigen-specific T-cell responses. Administration of FLT3 ligand (FL) results in a reversible accumulation of functionally active DCs in both lymphoid and nonlymphoid tissues. Therefore, we evaluated the possible antitumor effect of FL in murine melanoma (B16 and CL8-1) and lymphoma (EL-4) models. In all experiments, tumor growth was significantly inhibited by FL administration. Analysis by immunohistochemistry revealed an increase in the DC accumulation within B16 and EL-4 tumors after treatment with FL. No change was observed for CL8-1 melanoma. These data suggest a potential role for FL in the immunotherapy of malignant skin tumors and possible DC involvement in this effect.
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PMID:FLT3 ligand administration inhibits tumor growth in murine melanoma and lymphoma. 945 75

Proliferation and survival of hematopoietic progenitors are partially dependent on the interaction between the FLT3 receptor tyrosine kinase (RTK) and its ligand, FL. This biological function depends primarily on tyrosine phosphorylation of cellular targets that initiate several transduction cascades. These events return to their basal levels upon activation of specific phosphatases. We analyzed tyrosine phosphorylation events in response to FL, in human cell lines of different hematopoietic origins that express endogenous FLT3, namely the myelomonocytic, monocytic, pre-B and pro-B lineages. This study aimed at determining (1) the identity of FLT3 downstream substrates in physiologically relevant cells and (2) distinct substrate involvement in myeloid or early B cells. The two prominent tyrosine-phosphorylated proteins are p52SHC and p115CBL in myeloid cell lines and p52SHC and an uncharacterized p115 in early B cell lines. Following FL stimulation, a concomitant increase in both CBL phosphorylation and complex formation with p85 subunit of phosphatidylinositol 3' kinase is observed. In contrast, the GRB2/CBL association observed in unstimulated cells is not modified after stimulation, and SHC is never detected in anti-CBL immunoprecipitates. FL-inducible binding of CBL to the CRKII adaptor molecule is also demonstrated. This study presents a picture of the signaling events triggered by activation of endogenous FLT3 receptor in human hematopoietic cells, including the existence of a B cell-specific FLT3 substrate.
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PMID:FLT3 signaling in hematopoietic cells involves CBL, SHC and an unknown P115 as prominent tyrosine-phosphorylated substrates. 952 23

Epidermal Langerhans cells (LC) play a critical role in host defense. Still we know rather little about the development and functional specialization of these bone marrow-derived dendritic cells (DC) located in the most peripheral ectodermal tissue of the mammalian organism. How LC develop from their primitive progenitors in bone marrow and to what extent LC are related in their development to other lineages of the hemopoietic system is still under debate. There are currently 3 major areas of debate: 1) which are the signals required for LC development and differentiation to occur, 2) what are the (molecular) characteristics of the intermediate stages of LC differentiation, and 3) how are LC related in their development and/or function to other cells of the hemopoietic system? A better understanding of LC development and answers to these questions can be expected from recently developed technologies which allow the in vitro generation of DC with the typical molecular, morphological and functional features of LC from purified CD34+ progenitor cells under defined serum-free culture conditions. TGF-beta 1 was found to be an absolute requirement for in vitro LC development under serum-free conditions upon stimulation with the classical DC growth and differentiation factors GM-CSF, TNF-alpha and SCF. The recently identified cytokine FLT3 ligand further dramatically enhanced in vitro LC development and even allowed efficient in vitro generation of LC colonies from serum-free single cell cultures of CD34+ hemopoietic progenitor cells.
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PMID:Epidermal Langerhans cell development and differentiation. 956 75

In an effort to expand human hematopoietic progenitors and stem cells in vitro, we cultured human CD34(+)c-kitlow bone marrow cells in suspension in the presence of KIT ligand, FLK2/FLT3 ligand, interleukin-6 (IL-6), and erythropoietin with or without IL-3 and tested their engrafting capabilities by injecting them into sheep fetuses. As markers for engraftment, we analyzed CD45(+) cells and karyotypes of the colonies grown in methylcellulose culture. In three separate experiments, day-60 engraftment in the bone marrow was seen with both fresh cells and cells cultured in the presence or absence of IL-3. When fetuses were allowed to be born and analyzed for CD45(+) cells, no long-term engraftment was seen with cultured cells. We then pooled the CD45(+) cells of the fetal samples and transplanted them into secondary recipient fetuses. Day-60 engraftment in the secondary recipients was again noted when transplantation in the primary recipients was initiated with fresh cells. There were 3 cases in which cultured cells showed signs of engraftment in the secondary recipients, but the remaining 24 cases showed no signs of engraftment. These data documented that suspension culture for 2 weeks of enriched adult human bone marrow cells can maintain short-term (2 months) engrafting cells, but may not maintain longer term engrafting cells. This sheep/human xenograft model may serve as an excellent method for the evaluation of the engraftment potential of in vitro-expanded cells.
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PMID:Engraftment of cultured human hematopoietic cells in sheep. 957 5


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