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:2.7.10.2 (focal adhesion kinase)
44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cancer subtype diagnosis using microarray signatures has the potential to transform pathological diagnosis but the routine measurement of genes signatures remains difficult. Reverse transcription polymerase chain reaction (RT-PCR) measurement of Indicator genes for acute myeloid leukaemia (AML) and acute lymphoblastic leukaemia (ALL) was used to determine gene signatures. Bone marrow (BM) mononuclear cells were sorted into total, CD34(+) and CD34(-) fractions, and mRNAs globally amplified from each fraction using polyA PCR. The expression profile of the 17 top-ranked genes distinguishing AML and ALL were measured by RT-PCR in five ALL, 26 AML, 12 AML remission, four chronic myeloid leukaemia (CML) and nine morphologically normal BM samples. All but two of the genes measured showed similar expression in AML and ALL to that reported previously. Specifically, c-MYB (P </= 0.04) was significantly increased in ALL in the total fraction, whilst HOXA9 (P </= 0.19) and cystatin c (P </= 0.01) were increased in AML in the CD34(+) and CD34(-) fractions, respectively. c-MYB, hSNF2, RBAP48, HKRT-1, LYN, CD33, Adipsin and HOXA9 were increased in AML compared with remission AML, indicating an ability to determine disease activity. The method used is simple, sensitive and robust, enabling routine clinical use, and it can also be extended to other tumours types with gene signatures.
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PMID:Routine expression profiling of microarray gene signatures in acute leukaemia by real-time PCR of human bone marrow. 1602 52

We attempted to extend the lifespan of CD34+ stem/progenitor cells in human cord blood (CB) by transduction with lentiviral vectors carrying the human telomerase catalytic subunit (hTERT) and/or the human papillomavirus type 16 (HPV16) E6 and E7 oncogenes. We found that hTERT was incapable of prolonging the replicative capacity of CB cells maintained under serum-free conditions in the presence of stem cell factor, Flt3 ligand, thrombopoietin, and interleukin-3 beyond 4 months (n=3). However, transduced CB cells cultured in the same cytokine cocktail constitutively expressing HPV16 E6/E7 alone (n=2) or in concert with hTERT (n=9) continued to proliferate, giving rise to permanent (>2 years) cell lines with a CD45+ CD34- CD133+/- CD44+ CD235a+ CD71+ CD203+ CD33+ CD13+ myeloerythroid/mast cell progenitor phenotype. Notably, CB cell cultures expressing only HPV16 E6/E7 went through a crisis period, and the resulting oligoclonal cell lines were highly aneuploid. By comparison, the CB cell lines obtained by coexpression of HPV16 E6/E7 plus hTERT exhibited near-diploid karyotypes with minimal chromosomal aberrations, concomitant with stabilization of telomere length, yet were clonally derived. The immortalized E6/E7 plus hTERT-expressing CB cells were not tumorigenic when injected intravenously or subcutaneously into sublethally irradiated immunodeficient nonobese diabetic/severe combined immunodeficient mice but could be converted to a malignant state by ectopic expression of a v-H-ras or BCR-ABL oncogene. These findings provide new insights into the mechanisms governing the senescence checkpoint of primitive human hematopoietic precursors and establish a paradigm for studies of the multistep process of human leukemogenesis.
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PMID:Bypass of senescence, immortalization, and transformation of human hematopoietic progenitor cells. 1614 74

An association between an activating JAK2 mutation (JAK2(V617F)) and BCR/ABL-negative myeloproliferative disorders was recently reported in multiple simultaneous publications. In the current study, mutation analysis for JAK2(V617F) was performed in peripheral blood mononuclear cells (PBMC) from 157 patients with myelofibrosis with myeloid metaplasia (MMM) including 117 with agnogenic (AMM), 22 with postpolycythaemic (PPMM), and 18 with post-thrombocythaemic (PTMM) myeloid metaplasia. The detection rate for JAK2(V617F) was significantly higher in PPMM (91%; homozygous in 18%) compared with either AMM (45.3%; homozygous in 2.6%) or PTMM (38.9%; homozygous in 11.1%). Concomitant analysis in granulocytes (n=57) and CD34(+) cells (n=25) disclosed a higher incidence of homozygous JAK2(V617F) mutation but the overall mutation rate was similar to that obtained from PBMC. JAK2(V617F) was not detected in DNA derived from T cells (n=19). In AMM, the presence of JAK2(V617F) was associated with an older age at diagnosis and a history of thrombosis or pruritus. Multivariate analysis identified only age and the Dupriez prognostic score as independent prognostic factors; JAK2(V617F) had no prognostic significance. In conclusion, JAK2(V617F) is a myeloid lineage-specific event, its incidence in MMM is significantly higher with an antecedent history of polycythaemia vera (PV), and its presence in AMM does not affect prognosis but is associated with PV-characteristic clinical features.
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PMID:The JAK2(V617F) tyrosine kinase mutation in myelofibrosis with myeloid metaplasia: lineage specificity and clinical correlates. 1622 51

The BCR-ABL oncoprotein of chronic myelogenous leukemia (CML) localizes to the cell cytoplasm, where it activates proliferative and antiapoptotic signaling pathways. We previously reported that the combination of the ABL kinase inhibitor imatinib mesylate (IM) and the nuclear export inhibitor leptomycin B (LMB) traps BCR-ABL inside the nucleus, triggering the death of the leukemic cells. To evaluate the efficacy of the combination of IM and LMB on human cells we collected CD34-positive cells from 6 healthy donors and myeloid progenitors from 35 patients with CML. The sequential addition of IM and LMB generated the strongest reduction in the proliferative potential of the leukemic cells, with limited toxicity to normal myeloid precursors. Furthermore, nested reverse transcriptase-polymerase chain reaction (RT-PCR) analysis on colonies representative of each experimental condition demonstrated that the combination of IM and LMB was the most effective regimen in reducing the number of BCR-ABL-positive colonies. The efficacy of the 2-drug association was independent of the clinical characteristics of the patients. Our results indicate that strategies aimed at the nuclear entrapment of BCR-ABL efficiently kill human leukemic cells, suggesting that the clinical development of this approach could be of significant therapeutic value for newly diagnosed and IM-resistant CML patients.
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PMID:BCR-ABL nuclear entrapment kills human CML cells: ex vivo study on 35 patients with the combination of imatinib mesylate and leptomycin B. 1624 86

We studied the relationship between granulocyte JAK2 (V617F) mutation status, circulating CD34(+) cells, and granulocyte activation in myeloproliferative disorders. Quantitative allele-specific polymerase chain reaction (PCR) showed significant differences between various disorders with respect to either the proportion of positive patients (53%-100%) or that of mutant alleles, which overall ranged from 1% to 100%. In polycythemia vera, JAK2 (V617F) was detected in 23 of 25 subjects at diagnosis and in 16 of 16 patients whose disease had evolved into myelofibrosis; median percentages of mutant alleles in these subgroups were significantly different (32% versus 95%, P < .001). Circulating CD34(+) cell counts were variably elevated and associated with disease category and JAK2 (V617F) mutation status. Most patients had granulocyte activation patterns similar to those induced by administration of granulocyte colony-stimulating factor. A JAK2 (V617F) gene dosage effect on both CD34(+) cell counts and granulocyte activation was clearly demonstrated in polycythemia vera, where abnormal patterns were mainly found in patients carrying more than 50% mutant alleles. These observations suggest that JAK2 (V617F) may constitutively activate granulocytes and by this means mobilize CD34(+) cells. This exemplifies a novel paradigm in which a somatic gain-of-function mutation is initially responsible for clonal expansion of hematopoietic cells and later for their abnormal trafficking via an activated cell progeny.
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PMID:Relation between JAK2 (V617F) mutation status, granulocyte activation, and constitutive mobilization of CD34+ cells into peripheral blood in myeloproliferative disorders. 1637 57

We studied 25 patients with myelofibrosis with myeloid metaplasia and 19 patients with secondary myelofibrosis associated with pulmonary hypertension (PH). In these 2 groups, we compared the peripheral-blood CD34 count, the clonality of granulocytes and platelets in peripheral blood, the mutational status of the JAK2 kinase gene, and the morphology of the peripheral blood and bone marrow. We found that the following were distinctive features of myelofibrosis with myeloid metaplasia but not of secondary myelofibrosis due to PH: high circulating CD34 cell count, the presence of clonal platelets and granulocytes and of peripheral-blood dacrocytes, and a JAK2 1849G>T (V617F) mutation. We conclude that these are intrinsic features of clonal progenitors present in patients with myelofibrosis due to myeloproliferative disorders and that these features are not due to the abnormal marrow architecture seen in secondary myelofibrosis.
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PMID:High levels of circulating CD34 cells, dacrocytes, clonal hematopoiesis, and JAK2 mutation differentiate myelofibrosis with myeloid metaplasia from secondary myelofibrosis associated with pulmonary hypertension. 1692 98

Imatinib-refractory chronic myelogenous leukemia (CML) patients can experience long-term disease-free survival with myeloablative therapy and allogeneic hematopoietic cell transplantation; however, associated complications carry a significant risk of mortality. Transplantation of autologous hematopoietic cells has a reduced risk of complications, but residual tumor cells in the autograft may contribute to relapse. Development of methods for purging tumor cells that do not compromise the engraftment potential of the normal hematopoietic cells in the autograft has been a long-standing goal. Since primitive CML cells differentiate more rapidly in vitro than their normal counterparts and are also preferentially killed by mafosfamide and imatinib, we examined the purging effectiveness on CD34(+) CML cells using a strategy that combines a brief exposure to imatinib (0.5-1.0 microM for 72 h) and then mafosfamide (30-90 microg/ml for 30 min) followed by 2 weeks in culture with cytokines (100 ng/ml each of stem cell factor, granulocyte colony-stimulating factor and thrombopoietin). Treatment with 1.0 microM imatinib, 60 microg/ml mafosfamide and 14 days of culture with cytokines eliminated BCR-ABL(+) cells from chronic phase CML patient aphereses, while preserving normal progenitors. This novel purging strategy may offer a new approach to improving the effectiveness of autologous transplantation in imatinib-refractory CML patients.
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PMID:A novel triple purge strategy for eliminating chronic myelogenous leukemia (CML) cells from autografts. 1643 11

Dasatinib (BMS-354825), a novel dual SRC/BCR-ABL kinase inhibitor, exhibits greater potency than imatinib mesylate (IM) and inhibits the majority of kinase mutations in IM-resistant chronic myeloid leukemia (CML). We have previously demonstrated that IM reversibly blocks proliferation but does not induce apoptosis of primitive CML cells. Here, we have attempted to overcome this resistance with dasatinib. Primitive IM-resistant CML cells showed only single-copy BCR-ABL but expressed significantly higher BCR-ABL transcript levels and BCR-ABL protein compared with more mature CML cells (P = .031). In addition, CrKL phosphorylation was higher in the primitive CD34(+)CD38(-) than in the total CD34(+) population (P = .002). In total CD34(+) CML cells, IM inhibited phosphorylation of CrKL at 16 but not 72 hours, consistent with enrichment of an IM-resistant primitive population. CD34(+)CD38(-) CML cells proved resistant to IM-induced inhibition of CrKL phosphorylation and apoptosis, whereas dasatinib led to significant inhibition of CrKL phosphorylation. Kinase domain mutations were not detectable in either IM or dasatinib-resistant primitive CML cells. These data confirm that dasatinib is more effective than IM within the CML stem cell compartment; however, the most primitive quiescent CML cells appear to be inherently resistant to both drugs.
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PMID:Dasatinib (BMS-354825) targets an earlier progenitor population than imatinib in primary CML but does not eliminate the quiescent fraction. 1724 89

Hematopoietic stem cells (HSCs) arise, self-renew, or give rise to all hematopoietic lineages through the effects of transcription factors activated by signaling cascades. Lyl-1 encodes a transcription factor containing a basic helix-hoop-helix (bHLH) motif closely related to scl/tal, which controls numerous decisions in embryonic and adult hematopoiesis. We report here that Lyl-1 null mice are viable and display normal blood cell counts, except for a reduced number of B cells resulting from a partial block after the pro-B stage. Nevertheless, the deletion of Lyl-1 results in a diminution in the frequency of immature progenitors (Lin(-), CD34(-), sca-1(+), c-kit(+) [LSK], and LSK-side population [LSK-SP]) and in S(12) colony-forming unit (CFU-S(12)) and long-term culture-initiating cell (LTC-IC) content in embryonic day 14 fetal liver (E14 FL) and adult bone marrow (BM). More important, Lyl-1(-/-) E14 FL cells and BM are severely impaired in their competitive reconstituting abilities, especially with respect to B and T lineage reconstitution. Thus, ablation of Lyl-1 quantitatively and functionally affects HSCs, a cell population that transcribes Lyl-1 more actively than their differentiated progenies. Our results demonstrate for the first time that Lyl-1 functions are important for HSC properties and B-cell differentiation and that they are largely distinct from scl functions.
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PMID:The SCL relative LYL-1 is required for fetal and adult hematopoietic stem cell function and B-cell differentiation. 1651 64

In chronic myeloid leukaemia, CD34(+) stem/progenitor cells appear resistant to imatinib mesylate (IM) in vitro and in vivo. To investigate the underlying mechanism(s) of IM resistance, it is essential to quantify Bcr-Abl kinase status at the stem cell level. We developed a flow cytometry method to measure CrkL phosphorylation (P-CrkL) in samples with <10(4) cells. The method was first validated in wild-type (K562) and mutant (BAF3) BCR-ABL(+) as well as BCR-ABL(-) (HL60) cell lines. In response to increasing IM concentration, there was a linear reduction in P-CrkL, which was Bcr-Abl specific and correlated with known resistance. The results were comparable to those from Western blotting. The method also proved to be reproducible with small samples of normal and Ph(+) CD34(+) cells and was able to discriminate between Ph(-), sensitive and resistant Ph(+) cells. This assay should now enable investigators to unravel the mechanism(s) of IM resistance in stem cells.
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PMID:BCR-ABL activity and its response to drugs can be determined in CD34+ CML stem cells by CrkL phosphorylation status using flow cytometry. 1657 5


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