Susanne Viehmann

High expression of precursor microRNA-155/BIC RNA in children with Burkitt lymphoma.

In a recent issue of Genes, Chromosomes & Cancer, van den Berg et al. (2003) reported on the high expression of human BIC RNA in Hodgkin lymphoma by means of the serial analysis of gene expression (SAGE) technique. In addition, using a combination of RNA in situ hybridization and immunostaining, they found that the expression of BIC is specific for Reed–Sternberg (RS) cells and that the BIC transcripts were in the nuclei of the RS cells. In contrast to their findings in Hodgkin lymphoma, an analysis of 43 cases of non-Hodgkin lymphoma (NHL; 15 follicular lymphomas, 7 diffuse large B-cell lymphomas, 9 Burkitt lymphomas, 7 anaplastic large cell lymphomas, and 5 T-cell-rich B-cell lymphomas) did not reveal any remarkable up-regulation of BIC expression, although one case of Burkitt lymphoma (BL) showed weak expression of BIC in a minority of its tumor cells. The BIC locus was originally identified as a common retroviral integration site in avian-leukosis virus–induced B–cell lymphomas (Clurman and Hayward, 1989; Tam et al., 1997). It should be stressed that the human BIC gene encodes a microRNA, miR-155. This microRNA is encoded by nucleotides 241–262 of BIC, which spans 1,421 bp in total and is on chromosome 21 (GenBank accession number: AF402776). MicroRNAs (miRNAs) are an abundant class of noncoding RNAs that interact with coding mRNA and trigger either translation repression or direct RNA cleavage via RNA interference, depending on the degree of complementarity with the specific target mRNA (Ambros, 2001; Lagos-Quintana et al., 2001; Ruvkun, 2001; Lai, 2002; Pasquinelli, 2002; Ambros et al., 2003; Lagos-Quintana et al., 2003). Mature miRNAs are 21–23 nt long and are excised from an approximately 60to 80-nt double-stranded RNA hairpin by Dicer RNase III (Hutvagner et al., 2001). In the last 2 years, more than 200 human microRNA genes have been described, but the prediction and validation of their target mRNAs by computerized means and experimental approaches is a tantalizing and still unresolved task (Ambros et al., 2003). Apart from this open question, it was recently hypothesized that microRNA genes might play an important role in oncogenesis (McManus, 2003). We support the idea advanced by van den Berg et al. that BIC might play a role in the selection of B cells, but here we also extend their data by demonstrating that miR-155/BIC is highly expressed in pediatric BL but not in other hematologic malignancies, for example, pre-B/common or T-cell leukemia. The conditio sine qua non for the development of BL is activation of the MYC oncogene, mostly by chromosomal translocations in which MYC is juxtaposed to an immunoglobulin enhancer. On the other hand, there is a body of evidence that the activation of MYC alone is not sufficient for full malignancy. Providing support for this contention was the demonstration that MYC cooperates with BMI1, PIM1, RAF, BCL2, or, as shown in a very recent report, with the Werner syndrome protein WRN (Grandori et al., 2003) We analyzed tumor cells from 21 children (ages 2–13 years, with a median age of 6 years) with BL (n 11), common/pre-B acute lymphoblastic leukemia (ALL; n 6), or T-cell ALL (n 4). In all cases selected, the proportion of tumor cells was 80% or greater. The presence of an IGH/MYC rearrangement was demonstrated by long-distance polymerase chain reaction (PCR) in all BL cases, whereas neither the common/pre-B nor T-cell ALLs had such a recombination (Busch et al., unpublished data). All patients were treated according to the NHL-BFM 90, 95, or the ALL-BFM 90, 95 multicenter therapy study (Reiter et al., 1999; Schrappe et al., 2000). Immunophenotyping was done according to standard procedures (Ludwig et al., 1994). Informed parental consent was obtained in all cases. RNA isolated from peripheral blood from 11 healthy volunteers served as an additional control.

Monitoring of minimal residual disease (MRD) by real-time quantitative reverse transcription PCR (RQ-RT-PCR) in childhood acute myeloid leukemia with AML1/ETO rearrangement.

The fusion transcript AML1/ETO corresponding to translocation t(8;21)(q22;q22) can be found in approximately 7–12% of childhood de novo AML. Despite the favorable prognosis, some of these patients relapse. Most of MRD studies so far were performed on adults treated not uniformly. Therefore, we analyzed the follow-up of 15 AML1/ETO-positive children using real-time quantitative reverse transcription PCR (RQ-RT-PCR), all enrolled in the multicenter therapy trial AML-BFM 98. AML1/ETO copy numbers were normalized to the control gene ABL and the results were expressed in copy numbers AML1/ETO per 10 000 copies ABL. At diagnosis, a median of 10 789 copies AML1/ETO was found. A linear decrease to about 10 copies (2–4 log) could be seen in most of the children by the start of consolidation. In the majority of cases they remained positive at this low level during the ongoing therapy. Four children relapsed and two of them had a decrease of less than 2 log before starting consolidation. Three of the relapsed children showed, prior to relapse, an increase of the AML1/ETO fusion transcript at 6, 9, and 11 weeks, respectively. These results suggest that monitoring of minimal residual disease using RQ-RT-PCR could be helpful in detecting patients with a higher risk of relapse.

Trisomy 21 is a recurrent secondary aberration in childhood acute lymphoblastic leukemia with TEL/AML1 gene fusion.

TEL/AML1 gene fusion is the most frequent genetic lesion in pediatric acute lymphoblastic leukemia (ALL). It occurs as a consequence of the cryptic chromosomal translocation t(12;21)(p13;q22). In a cohort of 50 RT‐PCR–positive TEL/AML1 patients, karyotype examination by GTG banding and fluorescence in situ hybridization (FISH) allowed us to identify chromosome anomalies in addition to the already existing t(12;21). Secondary aberrations were found in 29 out of 41 patients (71%) at initial diagnosis and in all 9 patients with relapse. Structural rearrangements affected chromosome arms 2p, 2q, 5q, 9p, 12p (n = 2), 6q, 11p (n = 3), and 21q (n = 4). An extra chromosome 21 was found to be the most frequent anomaly. It was detected in 6 out of 41 patients at initial diagnosis (15%) and in 7 out of the 9 patients at relapse. No karyotype with trisomy 21 exceeded 47 chromosomes. Gain of chromosome 21 was the sole anomaly in GTG‐banding analysis in 2/41 patients at initial diagnosis and in 4/9 at relapse. Notably, chromosome painting analysis performed in 11 out of the 13 patients with an extra chromosome 21 revealed duplication of the normal chromosome 21 in 8, and duplication of der(21)t(12;21) in 3 patients. Furthermore, gain of der(21)t(12;21) chromosome was confined exclusively to the relapse patients. Genes Chromosomes Cancer 24:272–277, 1999.

Infant acute lymphoblastic leukemia - combined cytogenetic, immunophenotypical and molecular analysis of 77 cases.

We used karyotyping, fluorescence in situ hybridization (FISH), Southern blotting, and RT-PCR in order to analyze prospectively 77 infants (less than 1 year of age) with acute lymphoblastic leukemia for the occurrence of 11q23/MLL rearrangements and/or other cytogenetic abnormalities. Out of the 69 informative samples we found an 11q23/MLL rearrangement in 42 cases (61%). Regarding only pro-B ALL cases, the incidence of 11q23/MLL rearranged cases, however, reached more than 90% The infants were treated within the therapy studies ALL-BFM90, ALL-BFM95 and CoALL-05–92. For patients with an adequate follow-up of 4 years the event-free survival of the 11q23/MLL-positive and 11q23/MLL-negative group was 0.2 or 0.64, respectively (P = 0.024). The monoclonal antibody 7.1. (moab 7.1) does not react with normal hematopoetic precursors or mature blood cells but was shown to specifically react with leukemic cells bearing a rearrangement of chromosome 11q23 or the MLL gene, respectively. We, therefore, specifically addressed the question whether the reactivity of moab 7.1, as determined by flow cytometry, may substitute for molecular testing of an 11q23/MLL rearrangement in this cohort of infant ALLs. Reactivity of moab 7.1 indicated a 11q23/MLL rearrangement with a specificity of 100%. However, five of the 11q23/MLL-positive cases did not react with moab 7.1 indicating a sensitivity of 84% only. Three of these five moab 7.1-negative but 11q23/MLL-positive cases could be identified by their unique expression pattern of CD65s and/or CD15. Thus, 95% of all 11q23/MLL-positive ALL cases in infancy may be identified by flow cytometry based on their expression of CD15, CD65s and/or moab 7.1.

ORIGIN AND EVOLUTION OF MITOCHONDRIA : WHAT HAVE WE LEARNT FROM RED ALGAE?

Abstract The purpose of this review is to present an account of our current understanding of the structure, organization and evolution of mitochondrial genomes, and to discuss the origin and evolution of mitochondria from the perspective recently provided by the extensive sequenc-ing of various mitochondrial genomes. Mitochondrial-en-coded protein phylogenies are congruent with nuclear phylogenies and strongly support a monophyletic origin of mitochondria. The newly available data from red-algal mitochondrial genomes, in particular, show that the structural and functional diversity of mitochondrial genomes can be accounted for by paralogous evolution. We also discuss the influence of other constraints, such as uniparental inheritance, on the evolution of genome organization in mitochondria.

Cryptic chromosomal aberrations leading to an AML1/ETO rearrangement are frequently caused by small insertions

The translocation t(8;21)(q22;q22), which results in the fusion of the AML1 (RUNX1) and ETO (CBFA2T1) genes, is a recurrent aberration in acute myeloid leukemia (AML), preferentially correlated with FAB M2, and has the highest incidence in childhood AML. Because of the favorable prognosis, the evidence of the t(8;21) or the AML1/ETO fusion gene is mandatory in most of the therapy trials, allowing the stratification of the patients to the correct risk group in terms of treatment. Here we present six out of 59 children with AML who were positive for AML1/ETO by RT‐PCR, but showed no evidence of the classical t(8;21)(q22;q22) by conventional cytogenetics. Because of the discrepancies between molecular and cytogenetic analyses, these six patients were further investigated by fluorescence in situ hybridization analysis. Small hidden interstitial insertions resulting in an AML1/ETO rearrangement were detected in five (8.5%) of the 59 patients, whereas the sixth patient showed a cryptic three‐way translocation. The insertions could be characterized as ins(21;8) in three patients and ins(8;21) in the remaining two. Additionally, three of the patients showed secondary chromosome aberrations leading to a higher complexity of the karyotype. In conclusion, the combination of more than one standard technique in the analysis of AML1/ETO is useful to reveal the overall frequency of cryptic chromosome rearrangements and permits a better understanding of the mechanisms involved in the generation of this fusion gene.

Genes for two subunits of succinate dehydrogenase form a cluster on the mitochondrial genome of Rhodophyta

Mitochondrial DNA from the unicellular rhodophyteCyanidium caldarium RK-1 and the multicellularChondrus crispus were isolated, cloned, and sequenced. Two genes,sdhB andsdhC, that encode subunits of the succinate dehydrogenase, were identified by similarity. These genes form a cluster (titsdhCB) in both red algae.

Correlation of BCR/ABL transcript variants with patients' characteristics in childhood chronic myeloid leukaemia.

Background and objective:  The characteristic chromosomal translocation t(9;22)(q34;q11) in chronic myeloid leukaemia (CML) mainly results in the two different BCR/ABL fusion transcripts b2a2 or b3a2. Both transcript variants can occur simultaneously due to alternative splicing of the b3a2 transcript. Conflicting results have been reported on the influence of the transcripts on haematological findings at diagnosis and the course of the disease in adults while data concerning these topics on childhood CML are still missing. This paper reports on a correlation of BCR/ABL transcript variants with patients’ characteristics in childhood CML.

Treatment response and residual-disease monitoring in initial and relapsed TEL-AML1 positive childhood ALL.

Treatment response and residual-disease monitoring in initial and relapsed TEL-AML1 positive childhood ALL

Granulocyte colony-stimulating factor receptor expression and 11q23/ MLL genotype in childhood acute lymphoblastic leukemia developing during the first 18 months of life

Granulocyte colony-stimulating factor receptor expression and 11q23/ MLL genotype in childhood acute lymphoblastic leukemia developing during the first 18 months of life