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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Prospective studies on the detection of minimal residual disease (MRD) in acute leukemia patients have shown that large-scale MRD studies are feasible and that clinically relevant MRD-based risk group classification can be achieved and can now be used for designing new treatment protocols. However, multicenter international treatment protocols with MRD-based stratification of treatment need careful standardization and quality control of the MRD techniques. This was the aim of the European BIOMED-1 Concerted Action 'Investigation of minimal residual disease in acute leukemia: international standardization and clinical evaluation' with participants of 14 laboratories in eight European countries (ES, NL, PT, IT, DE, FR, SE and AT). Standardization and quality control was performed for the three main types of MRD techniques, ie flow cytometric immunophenotyping, PCR analysis of antigen receptor genes, and RT-PCR analysis of well-defined chromosomal aberrations. This study focussed on the latter MRD technique. A total of nine well-defined chromosome aberrations with fusion gene transcripts were selected: t(1;19) with E2A-PBX1, t(4;11) with MLL-AF4, t(8;21) with AML1-ETO, t(9;22) with BCR-ABL p190 and BCR-ABL p210, t(12;21) with TEL-AML1, t(15;17) with PML-RARA, inv (16) with CBFB-MYH11, and microdeletion 1p32 with SIL-TAL1. PCR primers were designed according to predefined criteria for single PCR (external primers A <--> B) and nested PCR (internal primers C <--> D) as well as for 'shifted' PCR with a primer upstream (E5' primer) or downstream (E3' primer) of the external A <--> B primers. The 'shifted' E primers were designed for performing an independent PCR together with one of the internal primers for confirmation (or exclusion) of positive results. Various local RT and PCR protocols were compared and subsequently a common protocol was designed, tested and adapted, resulting in a standardized RT-PCR protocol. After initial testing (with adaptations whenever necessary) and approval by two or three laboratories, the primers were tested by all participating laboratories, using 17 cell lines and patient samples as positive controls. This testing included comparison with local protocols and primers as well as sensitivity testing via dilution experiments. The collaborative efforts resulted in standardized primer sets with a minimal target sensitivity of 10-2 for virtually all single PCR analyses, whereas the nested PCR analyses generally reached the minimal target sensitivity of 10-4. The standardized RT-PCR protocol and primer sets can now be used for molecular classification of acute leukemia at diagnosis and for MRD detection during follow-up to evaluate treatment effectiveness.
Leukemia 1999 Dec
PMID:Standardized RT-PCR analysis of fusion gene transcripts from chromosome aberrations in acute leukemia for detection of minimal residual disease. Report of the BIOMED-1 Concerted Action: investigation of minimal residual disease in acute leukemia. 1060 11

The value of dual-color fluorescence in situ hybridization (FISH) for the detection of inv(16), using two contigs of cosmid probes mapping on both sides of the chromosome 16p breakpoint region, was evaluated in 23 acute myeloid leukemias (AML) in different phases of the disease. At diagnosis interphase FISH detected inv(16) in 19/19 (100%) cases with conventional cytogenetics (CC) evident aberration and excluded the rearrangement in two patients with CC suspected inv(16). Moreover, it also identified an associated del(16p) in two patients. At relapse, it revealed the inv(16) in 8/8 (100%) studied cases. These results were concordant with those of reverse transcriptase-polymerase chain reaction (RT-PCR). From 13 patients who obtained at least one complete remission (CR), 31 follow-up samples were analyzed using interphase FISH. Twenty-nine specimens scored negative for inv(16) and two were positive. RT-PCR detected CBFbeta/MYH11 transcripts in four of the nine CR samples analyzed, being more sensitive than interphase FISH. Eight of the 13 patients relapsed at a median time of 6.5 months (range 1-15) from the last negative FISH analysis. Of the two patients with positive FISH in CR, one relapsed soon after. At diagnosis and relapse, interphase-FISH proved to be an effective technique for detecting inv(16) appearing more sensitive than CC. Prospective studies with more frequent controls and possibly additional FISH probes are needed to assess the value of interphase FISH for minimal residual disease (MRD) and relapse prediction.
Leukemia 2000 Mar
PMID:Use of dual-color interphase FISH for the detection of inv(16) in acute myeloid leukemia at diagnosis, relapse and during follow-up: a study of 23 patients. 1072 Jan 27

We used a newly established real-time RT-PCR assay for the quantification of the leukemia-specific CBFB/MYH11 transcripts in inv(16)-positive acute myeloblastic leukemia. CBFB/MYH11 could be quantified over a five log range, with a detection limit of 10 molecules of a CBFB/MYH11 plasmid and a 1:10(5) dilution of RNA of the inv(16)-positive ME-1 cell line, respectively. The fusion transcripts were also quantified in 19 patients with acute myeloblastic leukemia and an inv(16) at initial diagnosis. The expression of CBFB/MYH11 varied over a two log range without correlation to clinical response or relapse rate. In nine patients, CBFB/MYH11 was also quantified during/after chemotherapy and autologous or allogeneic stem cell transplantation. All of these patients showed a similar decline of CBFB/MYH11 after intensive therapy. Six of these patients are in complete remission with a stable low-level or absent CBFB/MYH11 expression. Three patients relapsed, and their CBFB/MYH11 transcripts rose again to pretreatment levels. In two patients, this increase in CBFB/MYH11 could be detected by real-time PCR before hematological relapse. These data indicate that real-time RT-PCR can be used for the sensitive detection and quantification of CBFB/MYH11 transcripts in the follow-up of patients with inv(16)-positive AML.
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PMID:Detection and quantification of CBFB/MYH11 fusion transcripts in patients with inv(16)-positive acute myeloblastic leukemia by real-time RT-PCR. 1124 87

We report a case of acute myelomonocytic leukemia with eosinophilia (AML-M4eo) in a 65-year-old man following low dose methotrexate treatment for pemphigus vulgaris. Cytogenetic studies at diagnosis revealed a complex karyotype including a reciprocal translocation between 11q14.2 and 16q22, an inversion of chromosome 16(p13.1q22), and an apparently terminal deletion of 7q31. The presence of inv(16) was confirmed by reverse transcription-polymerase chain reaction which demonstrated a Type A fusion transcript derived from the core binding factor (CBF) beta and the smooth muscle myosin heavy chain (MYH11) genes. The patient was in complete hematologic and cytogenetic remission 6 months following intensive chemotherapy. Because AML-M4eo with inv(16) has a favorable prognosis, molecular studies should be performed in case the identification of inv(16) by conventional cytogenetics is difficult due to a complex karyotype.
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PMID:A novel variant three-way translocation of inversion 16 in a case of AML-M4eo following low dose methotrexate therapy. 1129 72

Amplification of the CBFbeta/MYH11 fusion transcript by a qualitative reverse transcription-polymerase chain reaction (RT-PCR) has been used to detect minimal residual disease (MRD) and assess the risk for disease relapse in inv(16)(p13q22) acute myeloid leukemia (AML). This strategy has, however, produced conflicting results and because of an uncertain predictive value, its use in the clinical setting cannot be recommended. The objective of the current study was to evaluate if quantification by Real Time RT-PCR could be useful to determine levels of CBFbeta/MYH11 fusion transcripts predictive of clinical outcome in inv(16)(p13q22) AML at diagnosis or during remission. Bone marrow (BM) samples from 16 patients with inv(16) AML enrolled on a German multicenter trial (AML HD93) were analyzed for levels of CBFbeta/MYH11 fusion transcripts by Real Time RT-PCR at diagnosis (n= 14), during remission (n= 10) and at relapse (n=6). The CBFbeta/MYH11 transcript copy number in each sample was normalized to copies of an internal control housekeeping transcript (ie 18S). The copy number measured at diagnosis or relapse were 3 to 4 log higher that those measured during remission, following completion of induction treatment. A high CBFbeta/MYH11 transcript copy number at diagnosis had a significant correlation with a high percentage of BM blasts (Spearman's coefficient = -0.66; P= 0.03), and a borderline correlation with a short complete remission (CR) duration (Spearman's coefficient = -0.51; P= 0.07). No difference in levels of CBFbeta/MYH11 fusion transcripts measured during intensification therapy was found between patients destined to relapse and those who continued in CCR (P= 0.75). Following completion of the entire chemotherapy program, patients that during CR showed a CBFbeta/MYH11 fusion transcript copy number >10 had a significantly shorter CR duration (P= 0.002) and higher risk for disease relapse (P= 0.05) than patients with a CBFbeta/MYH11 fusion transcript copy number <10. The results of the current study, therefore, suggest that it is possible to determine in remission samples a threshold of CBFbeta/MYH11 transcript copy number above which relapse occurs and below which continuous CR is likely.
Leukemia 2001 Jul
PMID:Quantification of CBFbeta/MYH11 fusion transcript by real time RT-PCR in patients with INV(16) acute myeloid leukemia. 1145 76

AML-1 is one of the most frequently translocated genes in human leukemia. AML-1 binds DNA and activates or represses transcription, while the chromosomal translocation fusion proteins in acute myeloid leukemia subvert these functions. The t(8;21) is the second most frequent translocation in acute myeloid leukemia and creates a fusion between the DNA binding domain of AML-1 and the ETO (also known as MTG8) corepressor. The t(12;21) is found in up to 25% of pediatric B cell acute lymphoblastic leukemias and fuses the ETS family transcription factor TEL to the amino terminus of AML-1. In addition, the inv(16), the most frequent translocation in acute myeloid leukemia, fuses the AML-1 cofactor CBFbeta to the smooth muscle myosin heavy chain MYH11. Both the t(8;21) and t(12;21) create transcriptional repressors that impair AML-1 target gene expression. We demonstrated that the fusion proteins encoded by these translocations contact the nuclear hormone corepressors (N-CoR/SMRT), mSin3A, and histone deacetylases. We have also found that both TEL and AML-1 interact with mSin3A. TEL also binds N-CoR and histone deacetylase-3, indicating that wild-type TEL is a transcriptional repressor. The t(12;21) fuses the mSin3A interaction domain of TEL to AML-1 to transform AML-1 from a regulated to an unregulated transcriptional repressor. The recognition that AML-1 interacts with mSin3A to repress transcription suggested that the inv(16) fusion protein might also repress the transcription of AML-1-target genes. In fact, the inv(16) encodes a protein that cooperates with AML-1 to repress transcription. The inv(16) fusion protein was found in a ternary complex with AML-1 and mSin3A, suggesting that the inv(16) also acts by recruiting transcriptional corepressors and histone deacetylases.
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PMID:Mechanisms of transcriptional repression by the t(8;21)-, t(12;21)-, and inv(16)-encoded fusion proteins. 1158 63

T(8;21) AML1(CBFA2)-ETO(MTG8) is the most common chromosomal translocation in acute myeloid leukemia (AML) in both children and adults. We sought to understand the structure and gain insight into the fusion process between AML1 and ETO by sequencing genomic fusions in 17 primary childhood AMLs and two cell lines with t(8;21). Reciprocal translocations were sequenced for seven of the 19 samples. We assumed a null hypothesis that the translocation breakpoints would be evenly distributed along the intronic breakpoint cluster regions. Testing for multimodality via smoothed bootstrap statistical methods suggested, however, the presence of two separate cluster regions within both the AML1 and ETO breakpoint cluster regions. ETObreakpoints were predominantly located in intron 1B in a defined cluster 5' of exon 1A (scan statistic P value = 0.00001). All patients with available RNA expressed an AML1-ETO mRNA fusion between exon 5 of AML1 and exon 2 of ETO. Since the structural restraints for the fusion protein of AML1-ETO exclude exon 1A, we reason that ETO intron 1B harbors a structural feature with propensity for breakage and/or recombination. Chromosomal breakpoints displayed evidence of fusion by a non-homologous end joining process, with microhomologies and nontemplate nucleotides at some fusion junctions. Breakpoints in general displayed similar complexity of duplications, deletions, and insertions to other common pediatric leukemia translocations (TEL-AML1, MLL-AF4, PML-RARA, CBFB-MYH11) that we and others have analyzed.
Leukemia 2001 Dec
PMID:Molecular characterization of genomic AML1-ETO fusions in childhood leukemia. 1175 12

PCR-based monitoring of minimal residual disease (MRD) in acute leukemias can be achieved via detection of fusion gene transcripts of chromosome aberrations or detection of immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangements. We wished to assess whether both PCR targets are complementary in acute myeloid leukemia (AML). We investigated 105 consecutive AML cases for the presence of fusion gene transcripts by reverse transcriptase polymerase chain reaction (RT-PCR): AML1-ETO associated with t(8;21), CBFB-MYH11 with inv(16), PML-RARA with t(15;17), BCR-ABL with t(9;22), and MLL-AF4 with t(4;11). In 17 out of 105 AML cases (16%), fusion gene transcripts were found. Ninety-five of these AML patients (13 with fusion gene transcripts) were also investigated for the presence of IGH, IGK, TCRG and TCRD rearrangements by Southern blot and/or PCR heteroduplex analysis and sequencing. In nine out of 95 patients (9.5%), such rearrangements were found. Combined data revealed that only one patient with a fusion gene transcript had a coexistent Ig/TCR rearrangement. The nine AML patients with Ig/TCR rearrangements, as well as five additional AML patients from a previous study were investigated in more detail, revealing that Ig/TCR rearrangements in AML are immature and unusual. The presence of Ig/TCR rearrangements in AML did not correlate with RAG gene expression levels as determined by real-time quantitative PCR. In conclusion, fusion gene transcripts and Ig/TCR rearrangements are infrequent, but complementary MRD-PCR targets in AML.
Leukemia 2002 Mar
PMID:Fusion gene transcripts and Ig/TCR gene rearrangements are complementary but infrequent targets for PCR-based detection of minimal residual disease in acute myeloid leukemia. 1189 40

The inv(16)(p13q22) chromosomal rearrangement associated with FAB M4Eo acute myeloid leukemia (AML) subtype is characterized by the presence of the CBFbeta/MYH11 fusion transcript that can be used to detect minimal residual disease (MRD). However, qualitative RT-PCR studies of MRD have so far produced conflicting results and seem of limited prognostic value. We have evaluated retrospectively MRD in a large series of CBFbeta/MYH11-positive patients employing both qualitative and quantitative (real-time PCR) approaches. 186 bone marrow samples from 36 patients were examined with a median follow-up of 27.5 months; 15 patients relapsed during follow-up. In qualitative studies, carried out by 'nested' RT-PCR assay, all patients in complete remission (CR) immediately after induction/consolidation therapy were found to be PCR positive. However, follow-up samples at later time points were persistently negative (except one case) in patients remaining in continuous CR (CCR) for more than 12 months. 16 patients were evaluated by quantitative real-time PCR assay: CBFbeta/MYH11 transcript copy number was normalized for expression of the housekeeping gene ABL, expressed as fusion gene copy number per 10(4) copies of ABL. A 2-3 log decline in leukemic transcript copy number was observed after induction/consolidation therapy. After achieving CR, the mean copy number was significantly higher in patients destined to relapse compared to patients remaining in CCR (151 vs 9, P < 0.0001 by Mann-Whitney test). Moreover, in CCR patients, the copy number dropped below the detection threshold after the treatment protocol was completed and remained undetectable in subsequent MRD analysis in accordance with results obtained by qualitative RT-PCR. On the contrary, in the seven patients who relapsed, the copy number in CR never declined below the detection threshold; thus a cut-off value discriminating these two groups of patients could be established. The findings of our study, if confirmed, might confer an important predictive value to quantitative real-time PCR determinations of MRD in patients with inv(16) leukemia.
Leukemia 2002 Jun
PMID:Assessment of minimal residual disease (MRD) in CBFbeta/MYH11-positive acute myeloid leukemias by qualitative and quantitative RT-PCR amplification of fusion transcripts. 1264 62

Multi-parameter flow cytometry, molecular genetics, and cytogenetic studies have all contributed to new classification of leukemia. In this review we discuss immunophenotypic characteristics of major genotypic leukemia categories. We describe immunophenotype of: B-lineage ALL with MLL rearrangements, TEL/AML1, BCR/ABL, E2A/PBX1 translocations, hyperdiploidy, and myc fusion genes; T-ALL with SCL gene aberrations and t(5;14) translocation; and AML with AML1/ETO, PML/RARalpha, OTT/MAL and CBFbeta/MYH11 translocations, trisomies 8 or 11 and aberrations of chromosomes 7 and 5. Whereas some genotypes associate with certain immunophenotypic features, others can present with variable immunophenotype. Single molecules (as NG2, CBFbeta/SMMHC and PML/RARalpha proteins) associated with or derived from specific translocations have been described. More often, complex immunophenotype patterns have been related to the genotype categories. Most known associations between immunophenotype and genotype have been defined empirically. Therefore, these associations should be validated in independent patient cohorts before they can be widely used for prescreening of leukemia. Progress in our knowledge on leukemia will show how the molecular-genetic changes modulate the immunophenotype as well as how the expressed protein molecules further modulate cell behavior.
Leukemia 2002 Jul
PMID:Antigen expression patterns reflecting genotype of acute leukemias. 1209 48


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