Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.10.2 (focal adhesion kinase)
 44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In recent years, the prognosis of chronic myeloid leukemia (CML) has been greatly improved either with interferon-alpha (IFN-alpha) therapy or allogeneic bone marrow transplantation (BMT). In the present study, minimal residual disease was evaluated in 21 patients in complete cytogenetic response (CCR) after such treatments. Samples from bone marrow aspirates or peripheral blood or both were analyzed by conventional cytogenetics, Southern blot, interphase fluorescent in situ hybridization (FISH), and quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR). In all patients, FISH detected 1% to 12% nuclei with a BCR-ABL fusion gene, whereas Q-RT-PCR experiments were negative or weakly positive. Based on these results, we hypothesize that the BCR-ABL genomic rearrangement persists unexpressed in nonproliferating cells whatever the treatment (IFN-alpha or BMT). These data point to the need for follow-up of CML patients in CCR over an extensive period at the DNA level (FISH) to evaluate the residual disease and at the RNA level (Q-RT-PCR) to estimate the risk of relapse. (Blood. 2000;95:404-408)
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PMID:Persistence of BCR-ABL genomic rearrangement in chronic myeloid leukemia patients in complete and sustained cytogenetic remission after interferon-alpha therapy or allogeneic bone marrow transplantation. 1062 42

We studied lineage-specific chimerism and minimal residual disease (MRD) in sequential posttransplant samples from 55 patients who underwent unmanipulated (n = 44) or partially T-cell-depleted (n = 11) allogeneic bone marrow transplantation (BMT) for chronic myeloid leukemia (CML). Chimerism was assessed by polymerase chain reaction (VNTR [variable number of tandem repeats]-PCR) analysis in highly purified CD19+, CD3+, CD15+, and CD56+ cell fractions, whereas MRD was investigated in whole blood by reverse transcriptase-PCR (RT-PCR) of both p210(BCR-ABL) and p190(BCR-ABL) hybrid transcripts. Of 55 patients, 14 (including 6 T-cell-depleted patients) had cytogenetic relapse at 5-80 months and progressed to hematologic relapse, while 41 patients remained in prolonged cytogenetic remission 12-107 months post-BMT. Before leukemia recurrence, patients in the relapse group showed a consistent evolution pattern sequentially featured by persistent p210(BCR-ABL) positivity, increasing mixed chimerism (MC) in myeloid cells, p190(BCR-ABL) positivity, and, finally, cytogenetic relapse. Myeloid MC preceded cytogenetic relapse by 2-12 months, whereas p190(BCR/ABL) was detected 1-6 months prior to cytogenetic relapse in 11 patients and concomitant with cytogenetic relapse in 3 patients. In the remission group, all patients invariably tested negative for p190(BCR-ABL); 10 patients tested positive for p210(BCR-ABL) at variable time-points but showed persistent full donor chimerism (DC), whereas 31 patients tested p210(BCR-ABL) negative and displayed full DC or transient MC due to the persistence of recipient T cells. Two patients in the relapse group were successfully reinduced into molecular remission with donor lymphocyte infusion. Sequential molecular analysis after such treatment showed the inverse pattern to that observed prior to relapse, ie, progressive disappearance of p190(BCR-ABL) transcripts, conversion of myeloid chimerism to donor type, and, finally, p210(BCR-ABL) negativity. We conclude that lineage-specific chimerism and p190(BCR-ABL) messenger RNA (mRNA) analyses contribute a better characterization of CML evolution after BMT and enable early identification of patients at the highest risk of relapse. (Blood. 2000;95:2659-2665)
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PMID:Molecular analysis of lineage-specific chimerism and minimal residual disease by RT-PCR of p210(BCR-ABL) and p190(BCR-ABL) after allogeneic bone marrow transplantation for chronic myeloid leukemia: increasing mixed myeloid chimerism and p190(BCR-ABL) detection precede cytogenetic relapse. 1075 48

Fluorescence in situ hybridization (FISH) is increasingly used as an adjunct to conventional cytogenetic analysis in the diagnosis of haematological malignancies and in monitoring minimal residual disease. FISH, however, is generally performed on slides prepared after short-term sample incubation and therefore, whilst faster than conventional cytogenetics, still requires a minimum of 2 days for a result to be obtained. A simplification of the FISH procedure is reported using uncultured cytospin preparations of bone marrow or peripheral blood for the rapid diagnosis of the BCR-ABL and PML-RARa gene rearrangements. It demonstrates that culturing has no effect on the ratio of normal to abnormal cells in the nondividing population. Data is presented from an analysis of 24 cases in whom unequivocal results were obtained in less than 12 h and in complete concordance with results obtained by conventional cytogenetics and/or interphase FISH.
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PMID:Rapid detection of BCR/ABL and PML/RARA using fluorescence in situ hybridization in cytospin preparations. 1079 99

The degree of tumor load reduction after therapy is an important prognostic factor for patients with CML. Conventional metaphase analysis has been considered to be the 'gold standard' for evaluating patient response to treatment but this technique normally requires bone marrow aspiration and is therefore invasive. The frequency of cytogenetic analyses can be considerably reduced if patients are also monitored by molecular methods, which can be performed on peripheral blood specimens. Of the various techniques available, most attention has been paid to RT-PCR for BCR-ABL mRNA since this is by far the most sensitive. Simple, non-quantitative RT-PCR analysis gives only limited information on patients after treatment. Quantitative RT-PCR assays have been developed to monitor the kinetics of residual BCR-ABL transcripts over time. Variables in the quantitative PCR assay may be controlled for by quantification of transcripts of a normal gene (eg ABL or glucose-6-phosphate dehydrogenase, G6PD) as an internal standard. After allogeneic stem cell transplantation, most patients become RT-PCR negative, often after a period of low level positivity that may persist for several months. Those patients destined to relapse are characterized by the reappearance and/or rising levels of BCR-ABL transcripts. In contrast, for patients treated with interferon-alpha (IFN) residual disease is rarely, if ever, eliminated. The actual level of minimal residual disease in complete cytogenetic responders to IFN correlates with the probability of relapse. New quantitative real time procedures promise to simplify the protocols that are currently in use, but standardization and the introduction of rigorous, internationally accepted controls are required to enable RT-PCR to become a robust and routine basis for therapeutic decisions.
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PMID:Detection and quantification of residual disease in chronic myelogenous leukemia. 1086 64

Our laboratory has been involved in the study of glutathione-sulfhydryl-transferase-pi (GST-pi) for several years. We have recently observed that during haematopoiesis in BMSC liquid cultures from CML patients who were candidates for transplant GST-pi was expressed in presumably malignant cells during different stages of cellular maturation. To confirm this finding, in the present work we are detecting GST-pi expression by immunofluorescence in BCR-ABL+ and BCR-ABL- cells done by FISH of PB from 30 CML patients during different clinical status: treatment (T), hematological relapse (R), blastic crisis (BC) or post-allotrasplant (PT). As well as in PB from 30 Blood-Bank donors. The results were %BCR-ABL+ GST-pi+ cells: T = 1-67, R = 33-69, BC = 90-100 and PT = 1-2; %BCR-ABL- GST-pi+ cells: T = 2-31, R = 5-18, BC = 0-10 and PT = 2-5; %BCR-ABL- GST-pi- cells: T = 2-97, R = 13-62, BC = 0 and PT = 93-96; %BCR-ABL+ GST-pi- cells: T = 0, R = 0, BC = 0 and PT = 0. GST-pi was not expressed in donor cells. The results obtained confirm our previous observations and suggest that GST-pi expression might be used for the evaluation of the minimal residual disease in CML patients.
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PMID:GST-pi expression in BCR-ABL+ and BCR-ABL- cells from CML patients. 1095 10

We designed a novel multiplex in-cell reverse transcription-polymerase chain reaction method for the simultaneous detection and differentiation of p190 and p210 BCR-ABL mRNAs within single cells from the human chronic myeloid leukemia and Philadelphia positive acute lymphoblastic leukemia. Human K562 chronic myeloid leukemia and SUP B-15 Ph+ acute lymphoblastic leukemia cell lines were used as positive controls for p210 and p190 BCR-ABL mRNAs, respectively. HL60 cell line was used as a negative control. After the leukemia cells were fixed and permeabilized, without extracting nucleic acids, the mRNAs were reverse transcribed to cDNAs, and the cDNAs were amplified by multiplex polymerase chain reaction with fluorescent primers specific for p190 and p210 BCR-ABL mRNAs. After transfer onto glass slides by cytospin, the amplified cells were detected by fluorescence microscopy. Fluorescence microscopy after propidium iodide or 4',6-diamidino-2-phenylindone counterstaining showed that the positive K562 cells exhibited a yellow-green fluorescent cytoplasm around a red nucleus, and that the positive SUP B-15 cells exhibited an orange cytoplasm around a blue nucleus. Only the red or blue nucleus was visible in respective negative HL60 cells. The specificity of amplification was confirmed by the absence of a signal when control experiments were performed either with RNase digestion of mRNA or without reverse transcriptase/Taq polymerase. We conclude that the multiplex in-cell reverse transcription-polymerase chain reaction method is capable of simultaneously detecting and differentiating the p210 and p190 BCR-ABL mRNAs of chronic myeloid leukemia and Philadelphia-positive acute lymphoblastic leukemia cells, and that it may be useful in quantitatively monitoring the minimal residual disease during therapy.
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PMID:Multiplex in-cell reverse transcription-polymerase chain reaction for the simultaneous detection of p210 and p190 BCR-ABL mRNAs in chronic myeloid leukemia and Philadelphia-positive acute lymphoblastic leukemia cell lines. 1109 54

The reverse transcriptase-polymerase chain reaction (RT-PCR) has become widely used for monitoring minimal residual disease after allogeneic stem cell transplantation (SCT) for chronic myeloid leukemia (CML). However, most of these studies were performed using qualitative RT-PCR, and the interpretation of the results obtained has been conflicting. The correlation of a quantitative RT-PCR test performed early after SCT (at 3 to 5 months) and long-term outcome of CML patients surviving for more than 6 months was studied. Between January 1991 and June 1999, data from 138 CML patients who received allografts were evaluated. Early RT-PCR results were classified as (1) negative if there were no BCR-ABL transcripts detected (n = 61), (2) positive at low level if the total number of BCR-ABL transcripts was less than 100 per microg RNA and/or the BCR-ABL/ABL ratio was less than 0.02% (n = 14), or (3) positive at high level if transcript levels exceeded the thresholds defined above (n = 63). Three years after SCT the cumulative incidence of relapse was 16.7%, 42.9%, and 86.4%, respectively (P =.0001). The relationship between BCR-ABL transcript level and probability of relapse was apparent whether patients had received sibling or unrelated donor SCT and also whether or not the transplantation was T cell depleted. The results suggest that quantitative RT-PCR performed early after SCT is useful for predicting outcome and may help to define the need for further treatment.
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PMID:Early detection of BCR-ABL transcripts by quantitative reverse transcriptase-polymerase chain reaction predicts outcome after allogeneic stem cell transplantation for chronic myeloid leukemia. 1123 91

Molecular genetic and cytoimmunological markers have been applied for the detection of minimal residual disease (MRD) in hematological malignancies. These markers include surface markers or rearranged T-cell receptor and immunoglobulin genes in the lymphoid malignancies and fused genes associated with chromosomal translocations such as BCR-ABL in t(9;22) or PML-RAR alpha in t(15;17) in myeloid malignancies. The expression of the WT1 gene is recognized as the universal tumor marker for a wide variety of hematological malignancies. Using these sensitive markers, a tumor cell in 10,000 to 1,000,000 normal cells can be detected. By examining a large number of patients, it has been shown that the MRD in the early phase of chemotherapy has a correlation with the prognosis of childhood ALL. Based on these observations, a new strategy of chemotherapy in which the post-remission therapy is modified based on the MRD results has begun. The amount of tumor cells contaminated in the autologous stem cell grafts in ALL patients might be related to the prognosis. The diagnosis of MRD will be used as an important routine examination in chemotherapy for leukemia/lymphoma patients.
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PMID:[The minimal residual disease (MRD) in hematological malignancies]. 1143 42

Qualitative RT-PCR methods used for monitoring minimal residual disease (MRD) in APL patients fail to predict relapse in up to 25% of patients in remission. We report here the development and evaluation of a highly sensitive (10(-5) and 10(-6) with one round and two rounds of PCR, respectively) competitive RT-PCR method to quantitate the PML-RARalpha fusion transcripts. PML-RARalpha transcript's levels were normalised to 10(5) copies of ABL transcript. Serial BM and PB samples from 16 patients with APL and t(15;17) were examined. Presentation samples from three patients (three BM, one PB) showed levels in the range of 0.7 x 10(6)-3.5 x 10(6) and 1.2 x 10(5) molecules in BM and PB samples respectively. Serial quantitation of MRD in both BM and PB samples showed significantly lower levels of PML-RARalpha transcripts in remission, although the majority of samples remain positive for the PML-RARalpha transcripts even those in long-term remission (up to 94 months). Levels of PML-RARalpha in remission samples were up to 2 x 10(2) and up to 5.2 x 10(1) molecules in BM and PB respectively. BM and PB samples taken from two patients 2-4 months before relapse showed significantly higher levels of PML-RARalpha transcripts (1.2 x 10(4) molecules in BM; 3.5 x 102, 1.2 x 10(2) and 1.2 x 10(3) in PB). The same samples, when tested with a standard qualitative RT-PCR for the amplification of PML-RARalpha (with a sensitivity of 10(-4)) produced negative results. This indicates that the qualitative methods would not have predicted relapse in these patients. Our data show that quantitating PML-RARalpha transcripts with a sensitive method may provide a superior approach for monitoring MRD in APL and identifying patients at high risk of relapse.
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PMID:Monitoring minimal residual disease and predicting relapse in APL by quantitating PML-RARalpha transcripts with a sensitive competitive RT-PCR method. 1145 74

The detection of BCR-ABL specific RNA by RT-PCR has been shown to predict relapse when positive 6 months after allogeneic stem cell transplantation (SCT) for chronic myelogenous leukemia (CML). In the present study, the focus was on evaluation of residual disease during the first weeks following SCT. In this study, 177 blood or marrow samples were obtained from 33 patients who received allogeneic (20 patients) or autologous (13 patients) SCT on day 0, day 30 and every 3 months for 1 year. T-cell depletion (TCD) was performed in 4 cases. On day 0 (day of graft infusion), 10/30 evaluable patients had negative RT-PCR (33%) regardless of pretransplant characteristics. On day 30, 14/18 patients (77%) from the allogeneic group had negative RT-PCR versus 0% in the autologous group. 2/4 patients who received TCD allogeneic grafts had day 30-positive PCR. Five patients in the allogeneic group had at least one positive RT-PCR sample between day 30 and day 90: 3 of them subsequently relapsed suggesting possible correlation between early positivity and relapse. Our results show that disappearance of MRD can be achieved within 3 months after transplantation in the majority of patients treated with allogeneic but not after autologous SCT. This suggests that the GVL effect might be operational early during the first weeks following transplantation.
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PMID:Survey of early disapearance of BCR/ABL fusion transcript after allogeneic or autologous stem cell transplantation for chronic myelogenous leukemia. 1169 49


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