AFDN

Gene Summary

Gene:AFDN; afadin, adherens junction formation factor
Aliases: AF6, MLLT4, MLL-AF6, l-afadin
Location:6q27
Summary:This gene encodes a multi-domain protein involved in signaling and organization of cell junctions during embryogenesis. It has also been identified as the fusion partner of acute lymphoblastic leukemia (ALL-1) gene, involved in acute myeloid leukemias with t(6;11)(q27;q23) translocation. Alternatively spliced transcript variants encoding different isoforms have been described for this gene, however, not all have been fully characterized.[provided by RefSeq, May 2011]
Databases:VEGA, OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:afadin
Source:NCBIAccessed: 16 March, 2017

Ontology:

What does this gene/protein do?
Show (17)
Pathways:What pathways are this gene/protein implicaed in?
Show (3)

Cancer Overview

Research Indicators

Publications Per Year (1992-2017)
Graph generated 16 March 2017 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Western Blotting
  • Kinesin
  • Cancer Gene Expression Regulation
  • Cloning, Molecular
  • Leukaemia
  • Polymerase Chain Reaction
  • Young Adult
  • Translocation
  • Signal Transduction
  • MLL
  • Molecular Sequence Data
  • Neoplasm Invasiveness
  • Mutation
  • Transcription Factors
  • Adolescents
  • Chromosome Mapping
  • MLLT4
  • Myosins
  • Oncogene Fusion Proteins
  • Nuclear Proteins
  • Gene Rearrangement
  • ras Proteins
  • Tumor Suppressor Gene
  • Neoplasm Proteins
  • Chromosome 6
  • DNA-Binding Proteins
  • Childhood Cancer
  • Histone-Lysine N-Methyltransferase
  • Ubiquitin-Protein Ligases
  • Tight Junctions
  • KMT2A
  • DNA Sequence Analysis
  • FISH
  • Acute Myeloid Leukaemia
  • Myeloid Leukemia
  • Proto-Oncogenes
  • Base Sequence
  • Chromosome 11
  • Infant
  • Sequence Tagged Sites
  • Chromosome Breakage
Tag cloud generated 16 March, 2017 using data from PubMed, MeSH and CancerIndex

Specific Cancers (1)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Entity Topic PubMed Papers
Acute Myeloid Leukaemia (AML)t(6;11)(q27;q23) in Acute Myeloid Leukemia

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: MLLT4 (cancer-related)

Xu Y, Chang R, Peng Z, et al.
Loss of polarity protein AF6 promotes pancreatic cancer metastasis by inducing Snail expression.
Nat Commun. 2015; 6:7184 [PubMed] Related Publications
Pancreatic cancer (PC) is a particularly lethal form of cancer with high potential for metastasis to distant organs. Disruption of cell polarity is a hallmark of advanced epithelial tumours. Here we show that the polarity protein AF6 (afadin and MLLT4) is expressed at low levels in PC. We demonstrate that depletion of AF6 markedly promotes proliferation and metastasis of PC cells through upregulation of the expression of Snail protein, and this requires the nuclear localization of AF6. Furthermore, AF6 deficiency in PC cells leads to increased formation of a Dishevelled 2 (Dvl2)-FOXE1 complex on the promoter region of Snail gene, and activation of Snail expression. Altogether, our data established AF6 as a potential inhibitor of metastasis in PC cells. Targeting the Dvl2-FOXE1-Snail signalling axis may thus represent a promising therapeutic strategy.

Yamamoto T, Mori T, Sawada M, et al.
Loss of AF-6/afadin induces cell invasion, suppresses the formation of glandular structures and might be a predictive marker of resistance to chemotherapy in endometrial cancer.
BMC Cancer. 2015; 15:275 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: AF-6/afadin plays an important role in the formation of adherence junctions. In breast and colon cancer, loss of AF-6/afadin induces cell migration and cell invasion. We aimed to elucidate the role of AF-6/afadin in human endometrial cancer.
METHODS: Morphology and AF-6/afadin expression in endometrial cancer cell lines was investigated by 3-dimensional culture. We used Matrigel invasion assay to demonstrate AF-6/afadin knockdown induced invasive capability. Cell proliferation assay was performed to estimate chemoresistance to doxorubicin, paclitaxel and cisplatin induced by AF-6/afadin knockdown. The associations between AF-6/afadin expression and clinicopathological status were determined by immunohistochemical analysis in endometrial cancer tissues. Informed consent was obtained from all patients before the study.
RESULTS: The majority of cell clumps in 3-dimensional cultures of Ishikawa cells that strongly expressed AF-6/afadin showed round gland-like structures. In contrast, the cell clumps in 3-dimensional cultures of HEC1A and AN3CA cells-both weakly expressing AF-6/afadin-showed irregular gland-like structures and disorganized colonies with no gland-like structures, respectively. AF-6/afadin knockdown resulted in reduced number of gland-like structures in 3-dimensional cultures and enhancement of cell invasion and phosphorylation of ERK1/2 and Src in the highly AF-6/afadin-expressing endometrial cancer cell line. Inhibitors of MAPK/ERK kinase (MEK) (U0126) and Src (SU6656) suppressed the AF-6/afadin knockdown-induced invasive capability. AF-6/afadin knockdown induced chemoresistance to doxorubicin, paclitaxel and cisplatin in Ishikawa cells, not in HEC1A. Immunohistochemical analysis showed that AF-6/afadin expression was significantly associated with myometrial invasion and high histological grade.
CONCLUSIONS: AF-6/afadin regulates cell morphology and invasiveness. Invasive capability is partly regulated through the ERK and Src pathway. The inhibitors to these pathways might be molecular-targeted drugs which suppress myometrial invasion in endometrial cancer. AF-6/afadin could be a useful selection marker for fertility-sparing therapy for patients with atypical hyperplasia or grade 1 endometrioid adenocarcinoma with no myometrial invasion. AF-6/afadin knockdown induced chemoresistance especially to cisplatin. Therefore, loss of AF-6/afadin might be a predictive marker of chemoresistance to cisplatin.

Deshpande AJ, Deshpande A, Sinha AU, et al.
AF10 regulates progressive H3K79 methylation and HOX gene expression in diverse AML subtypes.
Cancer Cell. 2014; 26(6):896-908 [PubMed] Free Access to Full Article Related Publications
Homeotic (HOX) genes are dysregulated in multiple malignancies, including several AML subtypes. We demonstrate that H3K79 dimethylation (H3K79me2) is converted to monomethylation (H3K79me1) at HOX loci as hematopoietic cells mature, thus coinciding with a decrease in HOX gene expression. We show that H3K79 methyltransferase activity as well as H3K79me1-to-H3K79me2 conversion is regulated by the DOT1L cofactor AF10. AF10 inactivation reverses leukemia-associated epigenetic profiles, precludes abnormal HOXA gene expression, and impairs the transforming ability of MLL-AF9, MLL-AF6, and NUP98-NSD1 fusions-mechanistically distinct HOX-activating oncogenes. Furthermore, NUP98-NSD1-transformed cells are sensitive to small-molecule inhibition of DOT1L. Our findings demonstrate that pharmacological inhibition of the DOT1L/AF10 complex may provide therapeutic benefits in an array of malignancies with abnormal HOXA gene expression.

Manara E, Baron E, Tregnago C, et al.
MLL-AF6 fusion oncogene sequesters AF6 into the nucleus to trigger RAS activation in myeloid leukemia.
Blood. 2014; 124(2):263-72 [PubMed] Related Publications
A rare location, t(6;11)(q27;q23) (MLL-AF6), is associated with poor outcome in childhood acute myeloid leukemia (AML). The described mechanism by which MLL-AF6, through constitutive self-association and in cooperation with DOT-1L, activates aberrant gene expression does not explain the biological differences existing between t(6;11)-rearranged and other MLL-positive patients nor their different clinical outcome. Here, we show that AF6 is expressed in the cytoplasm of healthy bone marrow cells and controls rat sarcoma viral oncogene (RAS)-guanosine triphosphate (GTP) levels. By contrast, in MLL-AF6-rearranged cells, AF6 is found localized in the nucleus, leading to aberrant activation of RAS and of its downstream targets. Silencing MLL-AF6, we restored AF6 localization in the cytoplasm, thus mediating significant reduction of RAS-GTP levels and of cell clonogenic potential. The rescue of RAS-GTP levels after MLL-AF6 and AF6 co-silencing confirmed that MLL-AF6 oncoprotein potentiates the activity of the RAS pathway through retention of AF6 within the nucleus. Exposure of MLL-AF6-rearranged AML blasts to tipifarnib, a RAS inhibitor, leads to cell autophagy and apoptosis, thus supporting RAS targeting as a novel potential therapeutic strategy in patients carrying t(6;11). Altogether, these data point to a novel role of the MLL-AF6 chimera and show that its gene partner, AF6, is crucial in AML development.

Lim JH, Jang S, Park CJ, et al.
FISH analysis of MLL gene rearrangements: detection of the concurrent loss or gain of the 3' signal and its prognostic significance.
Int J Lab Hematol. 2014; 36(5):571-9 [PubMed] Related Publications
INTRODUCTION: The rearrangement of the mixed-lineage leukemia (MLL) gene occurs through translocations and insertions involving a variety of partner chromosome genes. However, there are few studies on aberrant MLL signal patterns such as concurrent 3' MLL deletion.
METHODS: A total of 84 patients with acute leukemia (AL) who had MLL rearrangements detected by florescence in situ hybridization (FISH) were enrolled in the study. The distribution of MLL fusion partner genes was analyzed, and aberrant MLL signals were evaluated.
RESULTS: Seventy-seven (91.7%) patients had MLL rearrangements, involving previously described translocation partner genes (TPGs). Among these TPGs, the frequencies of MLLT3, AFF1, MLLT4, and ELL were 29.8%, 17.9%, 15.5%, and 13.1%, respectively. A high frequency of MLLT4 in our study was due to the high proportion of acute myeloid leukemia cases in pediatric and adult patients. Aberrant MLL signals were found in 18 patients: 11 (61.1%) with 3' MLL signal loss and 7 with 3' MLL signal gain. All cases with 3' MLL signal gain were due to an extra derivative partner chromosome. The median overall survival period of patients with 3' MLL gain was shorter than that in patients without aberrant MLL signal patterns.
CONCLUSION: Aberrant MLL signals were frequently detected by FISH analysis. The 3' MLL gain was associated with poor prognosis in patients with AL. Therefore, it is important to detect aberrant MLL signal patterns using FISH analysis.

Shiba N, Ichikawa H, Taki T, et al.
NUP98-NSD1 gene fusion and its related gene expression signature are strongly associated with a poor prognosis in pediatric acute myeloid leukemia.
Genes Chromosomes Cancer. 2013; 52(7):683-93 [PubMed] Related Publications
The cryptic t(5;11)(q35;p15.5) creates a fusion gene between the NUP98 and NSD1 genes. To ascertain the significance of this gene fusion, we explored its frequency, clinical impact, and gene expression pattern using DNA microarray in pediatric acute myeloid leukemia (AML) patients. NUP98-NSD1 fusion transcripts were detected in 6 (4.8%) of 124 pediatric AML patients. Supervised hierarchical clustering analyses using probe sets that were differentially expressed in these patients detected a characteristic gene expression pattern, including 18 NUP98-NSD1-negative patients (NUP98-NSD1-like patients). In total, a NUP98-NSD1-related gene expression signature (NUP98-NSD1 signature) was found in 19% (24/124) and in 58% (15/26) of cytogenetically normal cases. Their 4-year overall survival (OS) and event-free survival (EFS) were poor (33.3% in NUP98-NSD1-positive and 38.9% in NUP98-NSD1-like patients) compared with 100 NUP98-NSD1 signature-negative patients (4-year OS: 86.0%, 4-year EFS: 72.0%). Interestingly, t(7;11)(p15;p15)/NUP98-HOXA13, t(6;11)(q27;q23)/MLL-MLLT4 and t(6;9)(p22;q34)/DEK-NUP214, which are known as poor prognostic markers, were found in NUP98-NSD1-like patients. Furthermore, another type of NUP98-NSD1 fusion transcript was identified by additional RT-PCR analyses using other primers in a NUP98-NSD1-like patient, revealing the significance of this signature to detect NUP98-NSD1 gene fusions and to identify a new poor prognostic subgroup in AML.

Meyer C, Hofmann J, Burmeister T, et al.
The MLL recombinome of acute leukemias in 2013.
Leukemia. 2013; 27(11):2165-76 [PubMed] Free Access to Full Article Related Publications
Chromosomal rearrangements of the human MLL (mixed lineage leukemia) gene are associated with high-risk infant, pediatric, adult and therapy-induced acute leukemias. We used long-distance inverse-polymerase chain reaction to characterize the chromosomal rearrangement of individual acute leukemia patients. We present data of the molecular characterization of 1590 MLL-rearranged biopsy samples obtained from acute leukemia patients. The precise localization of genomic breakpoints within the MLL gene and the involved translocation partner genes (TPGs) were determined and novel TPGs identified. All patients were classified according to their gender (852 females and 745 males), age at diagnosis (558 infant, 416 pediatric and 616 adult leukemia patients) and other clinical criteria. Combined data of our study and recently published data revealed a total of 121 different MLL rearrangements, of which 79 TPGs are now characterized at the molecular level. However, only seven rearrangements seem to be predominantly associated with illegitimate recombinations of the MLL gene (≈ 90%): AFF1/AF4, MLLT3/AF9, MLLT1/ENL, MLLT10/AF10, ELL, partial tandem duplications (MLL PTDs) and MLLT4/AF6, respectively. The MLL breakpoint distributions for all clinical relevant subtypes (gender, disease type, age at diagnosis, reciprocal, complex and therapy-induced translocations) are presented. Finally, we present the extending network of reciprocal MLL fusions deriving from complex rearrangements.

Wang C, Cai X, Chen B, et al.
Up-regulation of tissue inhibitor of metalloproteinase-2 promotes SHI-1 cell invasion in nude mice.
Leuk Lymphoma. 2013; 54(12):2707-11 [PubMed] Related Publications
The role of tissue inhibitor of metalloproteinase-2 (TIMP-2) in extramedullary infiltration of acute leukemia is unclear. We demonstrated in our previous study that the up-regulation of TIMP-2 promoted SHI-1 cell invasion in vitro. We investigated in the present study whether TIMP-2 would have the same effect in vivo. A retroviral vector carrying human TIMP-2 cDNA was constructed and transfected into SHI-1 cells. Three subclone cells (S1, S2 and S3) that highly expressed TIMP-2 were selected to establish nude mouse models of acute leukemia. Times of leukemic onset in mice of S1, S2 and S3 groups were all earlier than that of the SHI-1 group, whereas the survival times of S1, S2 and S3 groups were all shorter than that of the SHI-1 group (p < 0.05). Histopathological results demonstrated severe leukemic infiltration in numerous organs in each group. Reverse transcription polymerase chain reaction (RT-PCR) assay showed that several organs expressed the MLL/F6 fusion gene. Moreover, the numbers of organs infiltrated by leukemic cells in S1, S2 and S3 groups were more than those in the SHI-1 group (p < 0.05). Up-regulating TIMP-2 expression enhanced SHI-1 cell invasion in nude mice and resulted in more severe leukemia infiltration. This phenomenon suggests that targeted therapy with TIMP-2 for acute leukemia should be performed with prudence.

Türkmen S, Timmermann B, Bartels G, et al.
Involvement of the MLL gene in adult T-lymphoblastic leukemia.
Genes Chromosomes Cancer. 2012; 51(12):1114-24 [PubMed] Related Publications
While the MLL "recombinome" is relatively well characterized in B-cell precursor acute lymphoblastic leukemia (BCP ALL), available data for adult acute T-lymphoblastic leukemia (T-ALL) are scarce. We performed fluorescence in situ hybridization (FISH) for an MLL split signal on 223 adult T-ALL samples obtained within the framework of the German Multicenter ALL 07/2003 therapy trial. Three biphenotypic leukemias (T-ALL/AML) were also included in the analysis. Samples showing any alteration by FISH were further investigated to characterize the MLL aberration. In addition, they were investigated for common genetic lesions known in T-ALL. Twenty-two cases (9.5%) showed an abnormal MLL signal by FISH analysis. Most of these appeared to be deletions or gains but in five cases (2.1%) a chromosomal translocation involving the MLL gene was identified. The translocation partners and chromosomal breakpoints were molecularly characterized. Three T-ALLs had an MLL-AF6/t(6;11) and two biphenotypic leukemias had an MLL-ELL/t(11;19). The chromosomal breakpoints in two of the MLL-AF6-positive cases were located outside the classical MLL major breakpoint cluster known from BCP ALL. In conclusion, the spectrum of MLL translocation partners in adult T-ALL much more resembles that of AML than that of BCP ALL and thus the mechanisms by which MLL contributes to leukemogenesis in adult T-ALL appear to differ from those in BCP ALL. Proposals are made for the diagnostic assessment of MLL fusion genes in adult T-ALL.

Cerveira N, Lisboa S, Correia C, et al.
Genetic and clinical characterization of 45 acute leukemia patients with MLL gene rearrangements from a single institution.
Mol Oncol. 2012; 6(5):553-64 [PubMed] Related Publications
Chromosomal rearrangements affecting the MLL gene are associated with high-risk pediatric, adult and therapy-associated acute leukemia. In this study, conventional cytogenetic, fluorescence in situ hybridization, and molecular genetic studies were used to characterize the type and frequency of MLL rearrangements in a consecutive series of 45 Portuguese patients with MLL-related leukemia treated in a single institution between 1998 and 2011. In the group of patients with acute lymphoblastic leukemia and an identified MLL fusion partner, 47% showed the presence of an MLL-AFF1 fusion, as a result of a t(4;11). In the remaining cases, a MLL-MLLT3 (27%), a MLL-MLLT1 (20%), or MLL-MLLT4 (7%) rearrangement was found. The most frequent rearrangement found in patients with acute myeloid leukemia was the MLL-MLLT3 fusion (42%), followed by MLL-MLLT10 (23%), MLL-MLLT1 (8%), MLL-ELL (8%), MLL-MLLT4 (4%), and MLL-MLLT11 (4%). In three patients, fusions involving MLL and a septin family gene (SEPT2, SEPT6, and SEPT9), were identified. The most frequently identified chromosomal rearrangements were reciprocal translocations, but insertions and deletions, some cryptic, were also observed. In our series, patients with MLL rearrangements were shown to have a poor prognosis, regardless of leukemia subtype. Interestingly, children with 1 year or less showed a statistically significant better overall survival when compared with both older children and adults. The use of a combined strategy in the initial genetic evaluation of acute leukemia patients allowed us to characterize the pattern of MLL rearrangements in our institution, including our previous discovery of two novel MLL fusion partners, the SEPT2 and CT45A2 genes, and a very rare MLL-MLLT4 fusion variant.

Bindels EM, Havermans M, Lugthart S, et al.
EVI1 is critical for the pathogenesis of a subset of MLL-AF9-rearranged AMLs.
Blood. 2012; 119(24):5838-49 [PubMed] Free Access to Full Article Related Publications
The proto-oncogene EVI1 (ecotropic viral integration site-1), located on chromosome band 3q26, is aberrantly expressed in human acute myeloid leukemia (AML) with 3q26 rearrangements. In the current study, we showed, in a large AML cohort carrying 11q23 translocations, that ∼ 43% of all mixed lineage leukemia (MLL)-rearranged leukemias are EVI1(pos). High EVI1 expression occurs in AMLs expressing the MLL-AF6, -AF9, -AF10, -ENL, or -ELL fusion genes. In addition, we present evidence that EVI1(pos) MLL-rearranged AMLs differ molecularly, morphologically, and immunophenotypically from EVI1(neg) MLL-rearranged leukemias. In mouse bone marrow cells transduced with MLL-AF9, we show that MLL-AF9 fusion protein maintains Evi1 expression on transformation of Evi1(pos) HSCs. MLL-AF9 does not activate Evi1 expression in MLL-AF9-transformed granulocyte macrophage progenitors (GMPs) that were initially Evi1(neg). Moreover, shRNA-mediated knockdown of Evi1 in an Evi1(pos) MLL-AF9 mouse model inhibits leukemia growth both in vitro and in vivo, suggesting that Evi1 provides a growth-promoting signal. Using the Evi1(pos) MLL-AF9 mouse leukemia model, we demonstrate increased sensitivity to chemotherapeutic agents on reduction of Evi1 expression. We conclude that EVI1 is a critical player in tumor growth in a subset of MLL-rearranged AMLs.

Fournier G, Cabaud O, Josselin E, et al.
Loss of AF6/afadin, a marker of poor outcome in breast cancer, induces cell migration, invasiveness and tumor growth.
Oncogene. 2011; 30(36):3862-74 [PubMed] Related Publications
Afadin/AF6, an F-actin-binding protein, is ubiquitously expressed in epithelia and has a key role during development, through its regulatory role in cell-cell junction organization. Afadin loss of expression in 15% of breast carcinoma is associated with adverse prognosis and increased risk of metastatic relapse. To determine the role of afadin in breast cancer, we studied the functional consequences of afadin protein extinction using in vitro and in vivo models. Three different breast cancer cell lines representative of the major molecular subtypes were stably repressed for afadin expression (knockdown of afadin (afadin KD)) using RNA interference. Collective and individual migrations as well as Matrigel invasion were markedly increased in afadin KD cells. Heregulin-β1 (HRG-β1)-induced migration and invasion were increased by twofold in afadin KD cells. Conversely, ectopic expression of afadin in the afadin-negative T47D cell line inhibited spontaneous and HRG-β1-induced migrations. RAS/MAPK and SRC kinase pathways were activated in afadin KD cells. Activation levels positively correlated with migration and invasion strength. Use of MEK1/2 (U0126) and SRC kinases (SU6656) inhibitors reduced afadin-dependent migration and invasion. Afadin extinction in the SK-BR-3 cell line markedly accelerated tumor growth development in mouse mammary gland and lung metastasis formation. These results may explain why the loss of afadin expression in tumors correlates with high tumor size and poor metastasis-free survival in patients.

Liedtke M, Ayton PM, Somervaille TC, et al.
Self-association mediated by the Ras association 1 domain of AF6 activates the oncogenic potential of MLL-AF6.
Blood. 2010; 116(1):63-70 [PubMed] Free Access to Full Article Related Publications
MLL is a common target for chromosomal translocations associated with acute leukemia resulting in its fusion with a large variety of nuclear or cytoplasmic proteins that may activate its oncogenic properties by distinct but poorly understood mechanisms. The MLL-AF6 fusion gene represents the most common leukemogenic fusion of mixed lineage leukemia (MLL) to a cytoplasmic partner protein. Here, we identified a highly conserved Ras association (RA1) domain at the amino-terminus of AF6 as the minimal region sufficient for MLL-AF6 mediated myeloid progenitor immortalization in vitro and short latency leukemogenesis in vivo. Moreover, the ability of RA1 to activate MLL oncogenesis is conserved with its Drosophila ortholog, Canoe. Although the AF6 RA1 domain has previously been defined as an interaction surface for guanosine triphosphate-bound Ras, single amino acid substitutions known to abolish the AF6-Ras interaction did not abrogate MLL-AF6-mediated oncogenesis. Furthermore, fusion of MLL to heterologous RA domains of c-Raf1 or RalGDS, or direct fusion of MLL to constitutively active K-RAS, H-RAS, or RAP1 was not sufficient for oncogenic activation of MLL. Rather, the AF6 RA1 domain efficiently mediated self-association, suggesting that constitutive MLL self-association is a more common pathogenic mechanism for MLL oncogenesis than indicated by previous studies of rare MLL fusion partners.

Banga SS, Peng L, Dasgupta T, et al.
PHF10 is required for cell proliferation in normal and SV40-immortalized human fibroblast cells.
Cytogenet Genome Res. 2009; 126(3):227-42 [PubMed] Free Access to Full Article Related Publications
Normal human diploid fibroblasts have limited life span in culture and undergo replicative senescence after 50-60 population doublings. On the contrary, cancer cells typically divide indefinitely and are immortal. Expression of SV40 large T and small t antigens in human fibroblasts transiently extends their life span by 20-30 population doublings and facilitates immortalization. We have identified a rearrangement in chromosome 6 shared by SV40-transformed human fibroblasts. Rearrangements involving chromosome 6 are among the most frequent in human carcinogenesis. In this paper, we extend analysis of the 6q26-q27 region, a putative site for a growth suppressor gene designated SEN6 involved in immortalization of SV40-transformed cells. Detailed molecular characterization of the rearranged chromosomes (6q*, normal appearing; and 6q(t), translocated) in the SV40-immortalized cell line HALneo by isolating each of these 2 chromosomes in mouse/HAL somatic cell hybrids is presented. Analysis of these mouse/HAL somatic cell hybrids with polymorphic and nonpolymorphic markers revealed that the 6q* has undergone a chromosomal break in the MLLT4 gene (alias AF6). This result in conjunction with previous published observations leads us to conclude that SEN6 lies between MLLT4 and TBP at chromosomal region 6q27. Examination of different genes (MLLT4, DLL1, FAM120B, PHF10) located within this interval that are expressed in HS74 normal fibroblast cells reveals that overexpression of epitope-tagged truncated PHF10 cDNAs resulted in reduced cell proliferation in multiple cell lines. Paradoxically, down-regulation of PHF10 by RNAi also resulted in loss of cell proliferation in normal fibroblast cells, indicating PHF10 function is required for cell growth. Taken together, these observations suggest that decreased cell proliferation with epitope-tagged truncated PHF10 proteins may be due to dominant negative effects or due to unregulated expression of these mutant proteins. Hence we conclude that PHF10 is not SEN6 but is required for cell growth.

Kobayashi S, Obata M, Hagihara M, et al.
The presence of mature granulocytes/monocytes derived from leukemic cells in MLL-associated leukemia.
Int J Hematol. 2009; 90(5):591-6 [PubMed] Related Publications
We observed the mature granulocytes/monocytes derived from leukemic cells in patients with acute myeloid leukemia who present mixed lineage leukemia gene (MLL). Morphologic observation and fluorescence in situ hybridization analysis (FISH) for chromosome 11q23 abnormality were studied, and a multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was done to identify the fusion partners with MLL. The bone marrow cells with FISH signals of MLL showed the cell differentiation of the myeloid and/or monocytic lineages in 4 of 6 AML patients. MLL partner genes were AF6, AF9, ELL, and ENL, respectively. There was no correlation between the fusion partner and the appearance of mature cells derived from MLL clones. RT-PCR showed the fusion between MLL exon 9 or 10 and the partner genes in mature granulocytes/monocytes. These findings suggest that subgroup of leukemia cells with MLL rearrangement has the differentiation potential of leukemic cells and mature granulocytes/monocytes derived from MLL clones may be biologically different from normal mature cells.

Giusiano S, Formisano-Tréziny C, Benziane A, et al.
Development of a biochip-based assay integrated in a global strategy for identification of fusion transcripts in acute myeloid leukemia: a work flow for acute myeloid leukemia diagnosis.
Int J Lab Hematol. 2010; 32(4):398-409 [PubMed] Related Publications
Three major types of rearrangements are involved in acute myeloid leukemias (AML): t(8;21)(q22;q22), inv(16)(p13q22), and 11q23/MLL abnormalities. Their precise identification becomes essential for diagnosis, prognosis, and therapeutic choices. Resulting fusion transcripts (FT) are also powerful markers for monitoring the efficacy of treatment, the minimal residual disease (MRD) and could become therapeutic targets. Today, the challenge is to propose an individual follow-up for each patient even for those with a rare fusion event. In this study, we propose a biochip-based assay integrated in a global strategy for identification of rare FT in AML, after fluorescence in situ hybridization detection, as described by the World Health Organization classification. Using cell lines, we developed and validated a biochip-based assay called the AMLFusionChip that identifies every FT of AML1-ETO, CBFbeta-MYH11 as well as MLL-AF9, MLL-ENL, MLL-AF6, and MLL-AF10. The original design of our AMLFusionChip.v01 enables the identification of these FT wherever the breakpoint on the partner gene may be. In case of biochip negative result, our 3'RACE amplification strategy enables to clone and then sequence the new translocation partner. This AMLFusionChip strategy fits into the concept of personalized medicine for the largest number of patients.

Radtke I, Mullighan CG, Ishii M, et al.
Genomic analysis reveals few genetic alterations in pediatric acute myeloid leukemia.
Proc Natl Acad Sci U S A. 2009; 106(31):12944-9 [PubMed] Free Access to Full Article Related Publications
Pediatric de novo acute myeloid leukemia (AML) is an aggressive malignancy with current therapy resulting in cure rates of only 60%. To better understand the cause of the marked heterogeneity in therapeutic response and to identify new prognostic markers and therapeutic targets a comprehensive list of the genetic mutations that underlie the pathogenesis of AML is needed. To approach this goal, we examined diagnostic leukemic samples from a cohort of 111 children with de novo AML using single-nucleotide-polymorphism microarrays and candidate gene resequencing. Our data demonstrate that, in contrast to pediatric acute lymphoblastic leukemia (ALL), de novo AML is characterized by a very low burden of genomic alterations, with a mean of only 2.38 somatic copy-number alterations per leukemia, and less than 1 nonsynonymous point mutation per leukemia in the 25 genes analyzed. Even more surprising was the observation that 34% of the leukemias lacked any identifiable copy-number alterations, and 28% of the leukemias with recurrent translocations lacked any identifiable sequence or numerical abnormalities. The only exception to the presence of few mutations was acute megakaryocytic leukemias, with the majority of these leukemias being characterized by a high number of copy-number alterations but rare point mutations. Despite the low overall number of lesions across the patient cohort, novel recurring regions of genetic alteration were identified that harbor known, and potential new cancer genes. These data reflect a remarkably low burden of genomic alterations within pediatric de novo AML, which is in stark contrast to most other human malignancies.

Meyer C, Kowarz E, Hofmann J, et al.
New insights to the MLL recombinome of acute leukemias.
Leukemia. 2009; 23(8):1490-9 [PubMed] Related Publications
Chromosomal rearrangements of the human MLL gene are associated with high-risk pediatric, adult and therapy-associated acute leukemias. These patients need to be identified, treated appropriately and minimal residual disease was monitored by quantitative PCR techniques. Genomic DNA was isolated from individual acute leukemia patients to identify and characterize chromosomal rearrangements involving the human MLL gene. A total of 760 MLL-rearranged biopsy samples obtained from 384 pediatric and 376 adult leukemia patients were characterized at the molecular level. The distribution of MLL breakpoints for clinical subtypes (acute lymphoblastic leukemia, acute myeloid leukemia, pediatric and adult) and fused translocation partner genes (TPGs) will be presented, including novel MLL fusion genes. Combined data of our study and recently published data revealed 104 different MLL rearrangements of which 64 TPGs are now characterized on the molecular level. Nine TPGs seem to be predominantly involved in genetic recombinations of MLL: AFF1/AF4, MLLT3/AF9, MLLT1/ENL, MLLT10/AF10, MLLT4/AF6, ELL, EPS15/AF1P, MLLT6/AF17 and SEPT6, respectively. Moreover, we describe for the first time the genetic network of reciprocal MLL gene fusions deriving from complex rearrangements.

van der Weyden L, Adams DJ
The Ras-association domain family (RASSF) members and their role in human tumourigenesis.
Biochim Biophys Acta. 2007; 1776(1):58-85 [PubMed] Free Access to Full Article Related Publications
Ras proteins play a direct causal role in human cancer with activating mutations in Ras occurring in approximately 30% of tumours. Ras effectors also contribute to cancer, as mutations occur in Ras effectors, notably B-Raf and PI3-K, and drugs blocking elements of these pathways are in clinical development. In 2000, a new Ras effector was identified, RAS-association domain family 1 (RASSF1), and expression of the RASSF1A isoform of this gene is silenced in tumours by methylation of its promoter. Since methylation is reversible and demethylating agents are currently being used in clinical trials, detection of RASSF1A silencing by promoter hypermethylation has potential clinical uses in cancer diagnosis, prognosis and treatment. RASSF1A belongs to a new family of RAS effectors, of which there are currently 8 members (RASSF1-8). RASSF1-6 each contain a variable N-terminal segment followed by a Ras-association (RA) domain of the Ral-GDS/AF6 type, and a specialised coiled-coil structure known as a SARAH domain extending to the C-terminus. RASSF7-8 contain an N-terminal RA domain and a variable C-terminus. Members of the RASSF family are thought to function as tumour suppressors by regulating the cell cycle and apoptosis. This review will summarise our current knowledge of each member of the RASSF family and in particular what role they play in tumourigenesis, with a special focus on RASSF1A, whose promoter methylation is one of the most frequent alterations found in human tumours.

Letessier A, Garrido-Urbani S, Ginestier C, et al.
Correlated break at PARK2/FRA6E and loss of AF-6/Afadin protein expression are associated with poor outcome in breast cancer.
Oncogene. 2007; 26(2):298-307 [PubMed] Related Publications
Common fragile sites (CFSs) are regions of chromosomal break that may play a role in oncogenesis. The most frequent alteration occurs at FRA3B, within the FHIT gene, at chromosomal region 3p14. We studied a series of breast carcinomas for break of a CFS at 6q26, FRA6E, and its associated gene PARK2, using fluorescence in situ hybridization on tissue microarrays (TMA). We found break of PARK2 in 6% of cases. We studied the PARK2-encoded protein Parkin by using immunohistochemistry on the same TMA. Loss of Parkin was found in 13% of samples but was not correlated with PARK2 break. PARK2 break but not Parkin expression was correlated with prognosis. Alteration of PARK2/FRA6E may cause haplo-insufficiency of one or several telomeric potential tumor suppressor genes (TSG). The AF-6/MLLT4 gene, telomeric of PARK2, encodes the Afadin scaffold protein, which is essential for epithelial integrity. Loss of Afadin was found in 14.5% of cases, and 36% of these cases showed PARK2 break. Loss of Afadin had prognostic impact, suggesting that AF-6 may be a TSG. Loss of Afadin was correlated with loss of FHIT expression, suggesting fragility of FRA6E and FRA3B in a certain proportion of breast tumors.

Robinson BW, Slater DJ, Felix CA
BglII-based panhandle and reverse panhandle PCR approaches increase capability for cloning der(II) and der(other) genomic breakpoint junctions of MLL translocations.
Genes Chromosomes Cancer. 2006; 45(8):740-53 [PubMed] Related Publications
Panhandle PCR techniques to amplify known sequence flanked by unknown sequence have been useful for MLL genomic breakpoint junctions and fusion transcripts because MLL has a large number of partner genes. However, genomic panhandle PCR approaches are impeded when the restriction fragment that contains the breakpoint junction is too large to amplify. We devised new panhandle PCR approaches for MLL genomic breakpoint junctions that create the template from BglII restriction fragments by attaching MLL sequence to a BglII site in the partner gene. This leads to the annealing of MLL and its complement in the handle and creates an intrastrand loop containing the breakpoint junction sequence for amplification with primers all from MLL. BglII panhandle PCR for der(11) breakpoint junctions was accomplished by ligating a phosphorylated oligonucleotide containing a BglII overhang and sequence complementary to MLL exon 7 to the 3' ends of BglII digested DNA, and forming the template from the sense strand of DNA. In BglII reverse panhandle PCR for der(other) breakpoint junctions, a phosphorylated oligonucleotide containing a BglII overhang and the complement of antisense sequence in MLL exon 10 was ligated to the 3' ends of BglII digested DNA, and the template was formed from the antisense strand of DNA. These approaches amplified 5'-MLL-MLLT4-3' and 5'-AFF1-MLL-3' breakpoint junctions. The former is significant because few t(6;11) genomic breakpoint junctions have been sequenced. BglII panhandle PCR approaches increase the possibilities for cloning MLL genomic breakpoint junctions where there is heterogeneity in partner genes and breakpoint locations.

Burjanivova T, Madzo J, Muzikova K, et al.
Prenatal origin of childhood AML occurs less frequently than in childhood ALL.
BMC Cancer. 2006; 6:100 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: While there is enough convincing evidence in childhood acute lymphoblastic leukemia (ALL), the data on the pre-natal origin in childhood acute myeloid leukemia (AML) are less comprehensive. Our study aimed to screen Guthrie cards (neonatal blood spots) of non-infant childhood AML and ALL patients for the presence of their respective leukemic markers.
METHODS: We analysed Guthrie cards of 12 ALL patients aged 2-6 years using immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements (n = 15) and/or intronic breakpoints of TEL/AML1 fusion gene (n = 3). In AML patients (n = 13, age 1-14 years) PML/RARalpha (n = 4), CBFbeta/MYH11 (n = 3), AML1/ETO (n = 2), MLL/AF6 (n = 1), MLL/AF9 (n = 1) and MLL/AF10 (n = 1) fusion genes and/or internal tandem duplication of FLT3 gene (FLT3/ITD) (n = 2) were used as clonotypic markers. Assay sensitivity determined using serial dilutions of patient DNA into the DNA of a healthy donor allowed us to detect the pre-leukemic clone in Guthrie card providing 1-3 positive cells were present in the neonatal blood spot.
RESULTS: In 3 patients with ALL (25%) we reproducibly detected their leukemic markers (Ig/TCR n = 2; TEL/AML1 n = 1) in the Guthrie card. We did not find patient-specific molecular markers in any patient with AML.
CONCLUSION: In the largest cohort examined so far we used identical approach for the backtracking of non-infant childhood ALL and AML. Our data suggest that either the prenatal origin of AML is less frequent or the load of pre-leukemic cells is significantly lower at birth in AML compared to ALL cases.

Shih LY, Liang DC, Fu JF, et al.
Characterization of fusion partner genes in 114 patients with de novo acute myeloid leukemia and MLL rearrangement.
Leukemia. 2006; 20(2):218-23 [PubMed] Related Publications
The fusion transcripts of MLL rearrangement [MLL(+)] in acute myeloid leukemia (AML) and their clinicohematologic correlation have not be well characterized in the previous studies. We used Southern blot analysis to screen MLL(+) in de novo AML. Reverse transcriptase-polymerase chain reaction was used to detect the common MLL fusion transcripts. cDNA panhandle PCR was used to identify infrequent or unknown MLL partner genes. MLL(+) was identified in 114 (98 adults) of 988 AML patients. MLL fusion transcripts comprised of 63 partial tandem duplication of MLL (MLL-PTD), 14 MLL-AF9, 9 MLL-AF10, 9 MLL-ELL, 8 MLL-AF6, 4 MLL-ENL and one each of MLL-AF1, MLL-AF4, MLL-MSF, MLL-LCX, MLL-LARG, MLL-SEPT6 and MLL-CBL. The frequency of MLL-PTD was 7.1% in adults and 0.9% in children (P<0.001). 11q23 abnormalities were detected in 64% of MLL/t11q23 and in none of MLL-PTD by conventional cytogenetics. There were no differences in remission rate, event-free survival and overall survival between adult MLL-PTD and MLL/t11q23 groups. Adult patients had a significantly poorer outcome than children. The present study showed that cDNA panhandle PCR can identify all rare or novel MLL partner genes. MLL-PTD was rare in childhood AML. MLL(+) adults had a poor outcome with no difference in survival between MLL-PTD and MLL/t11q23 groups.

Chen S, Xue Y, Zhang X, et al.
A new human acute monocytic leukemia cell line SHI-1 with t(6;11)(q27;q23), p53 gene alterations and high tumorigenicity in nude mice.
Haematologica. 2005; 90(6):766-75 [PubMed] Related Publications
BACKGROUND AND OBJECTIVES: Human leukemia cell lines are of great value in leukemia research. Thus far 36 leukemia cell lines carrying the 11q23 translocation and MLL rearrangements, including two cell lines with t(6;11)(q27;q23) and an MLL-AF6 fusion gene have been described. We have established a new monocytic cell line with t(6;11), designated SHI-1, and herein describe its biological characteristics.
DESIGN AND METHODS: Mononuclear cells isolated from the bone marrow of a patient with acute monocytic leukemia (AML-M5b) at relapse were inoculated and passaged by liquid culture. The biological features of the cell line were characterized by morphological assays, flow cytometry, cytogenetic analysis, reverse transcription polymerase chain reaction (RT-PCR), direct sequencing, fluorescence in situ hybridization (FISH), clonogenic culture, quantitative fluorescent PCR, zymography, short tandem repeating sequences-PCR (STR-PCR), multiplex-FISH (M-FISH), and tumorigenic capacity in nude mice.
RESULTS: The SHI-1 cell line has been maintained in continuous culture without any external cytokines for three years. The morphology and immunoprofile of the cells show typical features of monocytic lineage. Karyotypic analysis demonstrated a t(6;11)(q27;q23) translocation accompanied by a deletion of 17p, which are the same abnormalities as were seen in the leukemia cells of this patient in relapse. The MLL-AF6 fusion transcript and the loss of one p53 allele were proven by chromosome painting, FISH and RT-PCR analysis in both SHI-1 cells and the primary leukemia cells. A point mutation of ATC-->ACC at codon 195 of exon 6 in another p53 allele was found by direct sequencing of DNA in SHI-1 cells as well as in the primary leukemia cells. Neither Epstein-Barr virus nor mycoplasma was detected in SHI-1 cells. Tumor masses were found in all sixteen mice 9-19 days after subcutaneous injection of SHI-1 cells. DNA fingerprinting confirmed the authenticity of the cell line.
INTERPRETATION AND CONCLUSIONS: SHI-1 is a new monocytic leukemia cell line with the t(6;11) translocation, p53 gene alterations, and high tumorigenicity in nude mice. It could be a valuable tool in the study of leukemogenesis.

Maroc N, Morel A, Beillard E, et al.
A diagnostic biochip for the comprehensive analysis of MLL translocations in acute leukemia.
Leukemia. 2004; 18(9):1522-30 [PubMed] Related Publications
Reciprocal rearrangements of the MLL gene are among the most common chromosomal abnormalities in both Acute Lymphoblastic and Myeloid Leukemia. The MLL gene, located on the 11q23 chromosomal band, is involved in more than 40 recurrent translocations. In the present study, we describe the development and validation of a biochip-based assay designed to provide a comprehensive molecular analysis of MLL rearrangements when used in a standard clinical pathology laboratory. A retrospective blind study was run with cell lines (n=5), and MLL positive and negative patient samples (n=31), to evaluate assay performance. The limits of detection determined on cell line data were 10(-1), and the precision studies yielded 100% repeatability and 98% reproducibility. The study shows that the device can detect frequent (AF4, AF6, AF10, ELL or ENL) as well as rare partner genes (AF17, MSF). The identified fusion transcripts can then be used as molecular phenotypic markers of disease for the precise evaluation of minimal residual disease by RQ-PCR. This biochip-based molecular diagnostic tool allows, in a single experiment, rapid and accurate identification of MLL gene rearrangements among 32 different fusion gene (FG) partners, precise breakpoint positioning and comprehensive screening of all currently characterized MLL FGs.

Mitterbauer-Hohendanner G, Mannhalter C
The biological and clinical significance of MLL abnormalities in haematological malignancies.
Eur J Clin Invest. 2004; 34 Suppl 2:12-24 [PubMed] Related Publications
The MLL (Mixed Lineage Leukaemia or Myeloid/Lymphoid Leukaemia) gene on chromosome 11q23 is frequently involved in chromosomal translocations associated with human acute leukaemias. These translocations lead to fusion genes generally resulting in novel chimeric proteins containing the amino terminus of MLL fused in-frame to one of about 30 distinct partner proteins. Abnormalities involving the MLL gene are observed in leukaemias of either lymphoid or myeloid lineage derivation, as well as in poorly differentiated or biphenotypic leukaemias. They are frequently seen in infant patients, and patients with therapy-related secondary AML following treatment with inhibitors of topoisomerase II (epipodophyllotoxins). In the majority of cases, abnormalities involving the MLL gene are associated with a very poor prognostic outcome. In this review, we will discuss some of the recent advances in MLL research resulting from biological as well as clinical studies.

Li CY, Chow C, Chan WY
Allelic imbalance mapped to 6q14.1 is associated with loss of expression of 5-HT receptor 1B in non-Hodgkin lymphomas.
Diagn Mol Pathol. 2004; 13(2):69-74 [PubMed] Related Publications
Previous studies on lymphomas suggested that the long arm of chromosome 6 harbors 1 or more tumor suppressor genes. This study analyzed the status of 25 microsatellite markers in 39 cases, including 9 nodal and 30 extranodal, of non-Hodgkin lymphomas. Thirty of the 39 cases (77%) showed abnormality in at least 1 of the markers. Of the 655 informative results, 135 (20%) were abnormal. These included 5 homozygous deletions, 91 allelic imbalances (AI), and 38 microsatellite instability. The 2 commonest regions of abnormality were mapped to 6q14.1 and 6q27. There was no significant difference in the frequency of these regional losses between nodal and extranodal lymphomas, B-or T-cell lineage, and association with Epstein-Barr virus. The first common deletion region at 6q14.1 is flanked by the HTR1B (5-hydroxytryptamine receptor 1B) gene proximally and a novel unknown gene. AI in the region was found associated with loss of expression HTR1B by RT-PCR. The deletion region at 6q27 was narrowed to approximately 3Mb and maximal at marker D6S386. This locus includes the recently identified SMOC2 (secreted modular calcium-binding protein 2), AF6, and DLL1 (human delta-like 1 protein) genes. RT-PCR analyses of AF6 and DLL1 expression showed poor correlation with the AI results.

Ono R, Taki T, Taketani T, et al.
LCX, leukemia-associated protein with a CXXC domain, is fused to MLL in acute myeloid leukemia with trilineage dysplasia having t(10;11)(q22;q23).
Cancer Res. 2002; 62(14):4075-80 [PubMed] Related Publications
There are a limited number of reports of acute myeloid leukemia (AML) with t(10;11)(q22;q23). We showed that the MLL gene on 11q23 was fused to the LCX (leukemia-associated protein with a CXXC domain) gene on 10q22 in a de novoadult AML-M2 with trilineage dysplasia having t(10;11)(q22;q23). LCX consisted of at least 12 exons and was predicted to encode a 2136-amino-acid protein with an estimated molecular mass of 235.3 kDa. The LCX protein had a zinc-binding CXXC domain that MLL also contains within a methyltransferase domain, three nuclear localization signals, an alpha-helical coiled-coil region, and two homologous regions to CG2083 proteins of Drosophila melanogaster. We found approximately 12-, 9.5-, and 7.5-kb transcripts of LCX. Expression of the 7.5-kb transcript was detected in fetal heart, lung, and brain, and in adult skeletal muscle, thymus, and ovary. Expression of the 9.5-kb transcript was detected in fetal lung and brain and in adult ovary. Expression of the 12-kb transcript was detected in fetal heart and brain and in adult thymus and ovary. LCX was expressed in 8 of 22 leukemic cell lines, but not in EBV-induced normal B-cell lines. The MLL-LCX fusion protein lacked a CXXC domain of LCX, but retained an alpha-helical coiled-coil region at the COOH terminus, similar to MLL-SEPTING, MLL-CDCREL1, MLL-AF1p/Eps15, and MLL-AF6, which suggests that these fusion proteins are involved in the pathogenesis of 11q23-associated leukemia through similar mechanisms.

Andersson A, Höglund M, Johansson B, et al.
Paired multiplex reverse-transcriptase polymerase chain reaction (PMRT-PCR) analysis as a rapid and accurate diagnostic tool for the detection of MLL fusion genes in hematologic malignancies.
Leukemia. 2001; 15(8):1293-300 [PubMed] Related Publications
The MLL gene in chromosome band 11q23 is frequently rearranged in acute lymphoblastic and acute myeloid leukemias. To date, more than 50 different chromosomal regions are known to participate in translocations involving 11q23, many of which affect MLL. The pathogenetically important outcome of these rearrangements is most likely the creation of a fusion gene consisting of the 5' part of the MLL gene and the 3' end of the partner gene. Although abnormalities of the MLL gene as such are generally associated with poor survival, recent data suggest that the prognostic impact varies among the different fusion genes generated. Hence, detection of the specific chimeric gene produced is important for proper prognostication and clinical decision making. We have developed a paired multiplex reverse-transcriptase polymerase chain reaction analysis to facilitate a rapid and accurate detection of the most frequent MLL fusion genes in adult and childhood acute leukemias. To increase the specificity, two sets of primers were designed for each fusion gene, and these paired primer sets were run in parallel in two separate multiplex one-step PCR reactions. Using the described protocol, we were able to amplify successfully, in one single assay, the six clinically relevant fusion genes generated by the t(4;11)(q21;q23) [MLL/AF4], t(6;11)(q27;q23) [MLL/AF6], t(9;11)(p21-22;q23) [MLL/AF9], t(10;11)(p11-13;q23) [MLL/AF10], t(11;19)(q23;p13.1) [MLL/ELL], and t(11;19)(q23; p13.3) [MLL/ENL] in cell lines, as well as in patient material.

Strehl S, König M, Mann G, Haas OA
Multiplex reverse transcriptase-polymerase chain reaction screening in childhood acute myeloblastic leukemia.
Blood. 2001; 97(3):805-8 [PubMed] Related Publications
To determine the incidence of leukemia-specific rearrangements, 60 cases of childhood acute myeloblastic leukemia and transient myeloproliferative disorder were screened with a novel multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) assay, and the results were correlated with the cytogenetic findings. The RT-PCR assay detects 28 different fusion genes and more than 80 different fusion transcript variants. RNA was isolated from methanol/acetic acid-fixed cells that had been routinely prepared for cytogenetic analysis. Nine different fusion transcripts were found in 40% of the cases, whereas 78.3% of the cases had abnormal karyotypes. Two cases with a t(6;11) and an MLL/AF6 gene fusion were missed cytogenetically. Conversely, cytogenetic analysis revealed 10 other well-defined chromosome rearrangements. Although cytogenetic analysis reveals a much broader range of abnormalities, multiplex RT-PCR serves as quality control and provides the essential information for minimal residual disease studies. Moreover, discrepant findings lead to the detection of new rearrangements on the molecular genetic level.

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