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Biomedical subjects

Reiner Siebert

Publications and source records attributed to Reiner Siebert.

At least 19 recordsLinked to original sources
PubMed2006

Five members of the CEBP transcription factor family are targeted by recurrent IGH translocations in B-cell precursor acute lymphoblastic leukemia (BCP-ALL).

CCAAT enhancer-binding protein (CEBP) transcription factors play pivotal roles in proliferation and differentiation, including suppression of myeloid leukemogenesis. Mutations of CEBPA are found in a subset of acute myeloid leukemia (AML) and in some cases of familial AML. Here, using cytogenetics, fluorescence in situ hybridization (FISH), and molecular cloning, we show that 5 CEBP gene family members are targeted by recurrent IGH chromosomal translocations in BCP-ALL. Ten patients with t(8;14)(q11;q32) involved CEBPD on chromosome 8, and 9 patients with t(14;19)(q32;q13) involved CEBPA, while a further patient involved CEBPG, located 71 kb telomeric of CEBPA in chromosome band 19q13; 4 patients with inv(14)(q11q32)/t(14;14)(q11;q32) involved CEBPE and 3 patients with t(14;20)(q32;q13) involved CEBPB. In 16 patients the translocation breakpoints were cloned using long-distance inverse-polymerase chain reaction (LDI-PCR). With the exception of CEBPD breakpoints, which were scattered within a 43-kb region centromeric of CEBPD, translocation breakpoints were clustered immediately 5' or 3' of the involved CEBP gene. Except in 1 patient with t(14;14)(q11;q32), the involved CEBP genes retained germ-line sequences. Quantitative reverse transcription (RT)-PCR showed overexpression of the translocated CEBP gene. Our findings implicate the CEBP gene family as novel oncogenes in BCP-ALL, and suggest opposing functions of CEBP dysregulation in myeloid and lymphoid leukemogenesis.

Burkitt Lymphoma↗
PubMed2006

Chromosomal breakpoints affecting immunoglobulin loci are recurrent in Hodgkin and Reed-Sternberg cells of classical Hodgkin lymphoma.

Chromosomal breakpoints affecting immunoglobulin (IG) loci are recurrent in many subtypes of B-cell lymphomas. However, despite the predominant B-cell origin of the Hodgkin and Reed-Sternberg (HRS) cells in classical Hodgkin lymphoma (cHL), the presence of chromosomal translocations in IG loci has not yet been systematically explored. Therefore, we have investigated a series of cHL for chromosomal breakpoints in the IGH (n = 230), IGL (n = 139), and IGK (n = 138) loci by interphase cytogenetics. Breakpoints in the IGH, IGL, or IGK locus were observed in the HRS cells of 26 of 149 (17%), 2 of 70, and 1 of 77 evaluable cHLs, respectively. The IG partners could be identified in eight cHLs and involved chromosomal bands 2p16 (REL), 3q27 (BCL6, two cases), 8q24.1 (MYC), 14q24.3, 16p13.1, 17q12, and 19q13.2 (BCL3/RELB). In 65 of 85 (76%) cHLs evaluable for an IGH triple-color probe, the HRS cells showed evidence for a (partial) deletion of the IGH constant region, suggesting the presence of class switch recombination (CSR). Furthermore, analyses with this probe in cases with IGH breakpoints indicated that at least part of them seem to be derived from CSR defects. Our results show that chromosomal breakpoints affecting the IG loci are recurrent in cHL.

Adolescent↗
PubMed2006

Homozygous deletions localize novel tumor suppressor genes in B-cell lymphomas.

Integrative genomic and gene-expression analyses have identified amplified oncogenes in B-cell non-Hodgkin lymphoma (B-NHL), but the capability of such technologies to localize tumor suppressor genes within homozygous deletions remains unexplored. Array-based comparative genomic hybridization (CGH) and gene-expression microarray analysis of 48 cell lines derived from patients with different B-NHLs delineated 20 homozygous deletions at 7 chromosome areas, all of which contained tumor suppressor gene targets. Further investigation revealed that only a fraction of primary biopsies presented inactivation of these genes by point mutation or intragenic deletion, but instead some of them were frequently silenced by epigenetic mechanisms. Notably, the pattern of genetic and epigenetic inactivation differed among B-NHL subtypes. Thus, the P53-inducible PIG7/LITAF was silenced by homozygous deletion in primary mediastinal B-cell lymphoma and by promoter hypermethylation in germinal center lymphoma, the proapoptotic BIM gene presented homozygous deletion in mantle cell lymphoma and promoter hypermethylation in Burkitt lymphoma, the proapoptotic BH3-only NOXA was mutated and preferentially silenced in diffuse large B-cell lymphoma, and INK4c/P18 was silenced by biallelic mutation in mantle-cell lymphoma. Our microarray strategy has identified novel candidate tumor suppressor genes inactivated by genetic and epigenetic mechanisms that substantially vary among the B-NHL subtypes.

Adaptor Proteins, Signal Transducing↗
PubMed2006

A biologic definition of Burkitt's lymphoma from transcriptional and genomic profiling.

BACKGROUND: The distinction between Burkitt's lymphoma and diffuse large-B-cell lymphoma is unclear. We used transcriptional and genomic profiling to define Burkitt's lymphoma more precisely and to distinguish subgroups in other types of mature aggressive B-cell lymphomas. METHODS: We performed gene-expression profiling using Affymetrix U133A GeneChips with RNA from 220 mature aggressive B-cell lymphomas, including a core group of 8 Burkitt's lymphomas that met all World Health Organization (WHO) criteria. A molecular signature for Burkitt's lymphoma was generated, and chromosomal abnormalities were detected with interphase fluorescence in situ hybridization and array-based comparative genomic hybridization. RESULTS: We used the molecular signature for Burkitt's lymphoma to identify 44 cases: 11 had the morphologic features of diffuse large-B-cell lymphomas, 4 were unclassifiable mature aggressive B-cell lymphomas, and 29 had a classic or atypical Burkitt's morphologic appearance. Also, five did not have a detectable IG-myc Burkitt's translocation, whereas the others contained an IG-myc fusion, mostly in simple karyotypes. Of the 176 lymphomas without the molecular signature for Burkitt's lymphoma, 155 were diffuse large-B-cell lymphomas. Of these 155 cases, 21 percent had a chromosomal breakpoint at the myc locus associated with complex chromosomal changes and an unfavorable clinical course. CONCLUSIONS: Our molecular definition of Burkitt's lymphoma clarifies and extends the spectrum of the WHO criteria for Burkitt's lymphoma. In mature aggressive B-cell lymphomas without a gene signature for Burkitt's lymphoma, chromosomal breakpoints at the myc locus were associated with an adverse clinical outcome.

Algorithms↗
PubMed2006

Breakpoint cloning and haplotype analysis indicate a single origin of the common Inv(10)(p11.2q21.2) mutation among northern Europeans.

The pericentric inv(10)(p11.2q21.2) mutation has been frequently identified in cytogenetic laboratories, is phenotypically silent, and is considered to be a polymorphic variant. Cloning and sequencing of the junction fragments on 10p11 and 10q21 revealed that neither inversion breakpoint directly involved any genes or repetitive sequences, although both breakpoint regions contain a number of repeats. All 20 apparently unrelated inv(10) families in our study had identical breakpoints, and detailed haplotype analysis showed that the inversions were identical by descent. Thus, although considered a common variant, inv(10)(p11.2q21.2) has a single ancestral founder among northern Europeans.

Base Sequence↗
PubMed2006

Characterization of two supernumerary marker chromosomes in a patient with signs of Klinefelter syndrome, mild facial anomalies, and severe speech delay.

A boy with signs of Klinefelter syndrome, mild facial dysmorphic features, and severely retarded speech development displayed a female karyotype with mosaicism for two marker chromosomes 48,XX,+mar1,+mar2[68]/47,XX,+mar1[19]/47,XX,+mar2[6]/46,XX[8]. Using chromosomal microdissection, locus-specific fluorescence in situ hybridization (FISH), and PCR with several Y-chromosome markers, the larger supernumerary marker chromosome (SMC) was characterized as a ring Y-chromosome. Detection of the SRY-region explained the male phenotype. The smaller second marker chromosome contained the pericentromeric region of chromosome 8. We suggest that the co-occurrence of a partial Y-chromosome and partial trisomy 8 explain the severe speech delay and the facial dysmorphic features.

Abnormalities, Multiple↗
PubMed2006

Diffuse large B-cell lymphoma in pediatric patients belongs predominantly to the germinal-center type B-cell lymphomas: a clinicopathologic analysis of cases included in the German BFM (Berlin-Frankfurt-Munster) Multicenter Trial.

Diffuse large B-cell lymphoma (DLBCL) in adults is a heterogeneous disease. Biologic subgroups of DLBCL with a favorable prognosis (germinal center B-cell-like, GCB) and with a poor prognosis (activated B-cell-like, ABC) have been defined by gene expression profiling and can be distinguished by immunohistochemistry. In contrast to their adult counterparts, children with DLBCL have an excellent prognosis. We analyzed 63 cases of DLBCL in pediatric patients by immunohistochemistry and fluorescence in situ hybridization (FISH) and found a striking predominance of a GCB subtype, which might explain the good clinical outcome in these lymphomas. Interestingly, FISH applied to 50 of these cases, as well as conventional cytogenetics available in 3 cases, revealed absence of the translocation t(14;18) involving the BCL2 gene, which is present in about 15% of adult GCB subtype DLBCL. Our data indicate that pediatric DLBCL differs from adult DLBCL and might comprise a biologically unique subgroup of DLBCL from which important insights into the pathogenesis and biology of this disease might be gained.

Child↗
PubMed2006

A novel fusion of the MALT1 gene and the microtubule-associated protein 4 (MAP4) gene occurs in diffuse large B-cell lymphoma.

Rearrangements of the MALT1 gene by the t(11;18)(q21;q21) and t(14;18)(q32;q21) are the most frequent structural chromosomal abnormalities in MALT lymphomas. These translocations lead to fusions of BIRC3-MALT1 and IGH-MALT1 respectively, and activate the NF-kappaB pathway. Among 122 diffuse large B-cell lymphomas and 28 Burkitt's lymphomas screened by interphase FISH, we found two cases with a break within MALT1, but without a t(11;18) or a t(14;18). Molecular genetic analyses in one of these cases revealed a novel "in frame" fusion of exon 9 of MALT1 and exon 9 of the microtubule-associated protein 4 (MAP4) gene. The translocation was accompanied by a deletion of MALT1 sequences distal to the breakpoint including the caspase-like domain, which is essential for activation of NF-kappaB. As a result of the deletion, the reciprocal 5'MAP4-3'MALT1 transcript was not present, demonstrating that the 5'MALT1-3'MAP4 fusion represents the pathogenetically relevant transcript. Immunohistochemistry with amino-terminal and carboxy-terminal MALT1 antibodies, indicated a strong expression of the chimeric MALT1-MAP4 protein. Moreover, NF-kappaB activation was not increased in this case as shown by the levels of IkappaBalpha phosphorylation and NEMO ubiquitination. Our data demonstrate that the pathogenetic consequences of the novel MALT1-MAP4 fusion are different from those of the known MALT1-associated chromosomal rearrangements and do not involve NF-kappaB activation.

Aged↗
PubMed2006

Non-linkage of familial rhabdoid tumors to SMARCB1 implies a second locus for the rhabdoid tumor predisposition syndrome.

BACKGROUND: Rhabdoid tumors represent an independent entity among embryonal neoplasms. These tumors affect the kidney (RTK, rhabdoid tumor of kidney) and central nervous system (AT/RT, atypical teratoid, rhabdoid tumor), but may also be found in peripheral soft tissue. Unifying features include immunohistochemical characteristics and inactivation of the putative tumor suppressor gene SMARCB1 (hSNF5/INI1) in chromosome 22q11.2. Several familial cases have been published and summarized under the term rhabdoid tumor predisposition syndrome. In all of the published familial cases, inactivation of SMARCB1 was detected in tumor tissues. PROCEDURE AND RESULTS: We report on a family with three children, two of which were affected by rhabdoid tumors, one RTK, the other an AT/RT. While both children demonstrated typical morphological and clinical features neither the RTK nor the AT/RT showed evidence for inactivation of SMARCB1 in molecular studies including CGH and array CGH, FISH, gene dosage analysis by dHPLC, and DNA-sequencing. Immunohistochemistry for SMARCB1 showed normal expression within the nuclei of tumor cells. Furthermore, both children inherited different paternal and maternal SMARCB1 alleles evidenced by haplotype analysis. Conventional cytogenetic, FISH, and mutation analyses lacked evidence for SMARCB1 aberrations or gross chromosomal changes in the parents. CONCLUSIONS: We thus demonstrate a family with rhabdoid tumor predisposition syndrome without linkage to SMARCB1. This finding indicates that other loci than SMARCB1 below the resolution of array CGH are involved in the origin of these tumors. Our data impact on the clinical counseling of affected families and warrant further studies in the molecular biology of these enigmatic tumors.

Chromosomal Proteins, Non-Histone↗
PubMed2006

Identification of recurrent chromosomal breakpoints in multiple myeloma with complex karyotypes by combined G-banding, spectral karyotyping, and fluorescence in situ hybridization analyses.

The description of novel chromosomal aberrations in multiple myeloma (MM) remains necessary to fully understand the pathogenesis of this heterogeneous disease. Therefore, we have used spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH) with locus-specific probes to characterize the chromosomal abnormalities in 11 MM cases in which G-banding revealed a complex karyotype. SKY refined G-banding karyotypes in all cases. Recurrent breakpoints involved bands Xp11, 8q24, 11q13, 12q13, 13q21, and 14q32. In addition, combined SKY and FISH analyses permitted us to identify a subset of patients harboring 22q11.2 rearrangements not involving the IGL locus. This finding suggests the presence of other gene(s) in band 22q11 that might be implicated in MM pathogenesis. Moreover, band 1p13 was identified as a novel partner of immunoglobulin (IG) translocations in MM. Finally, using interphase FISH, we have detected interstitial deletions in 13q14 and 17p13, as well as cryptic translocations affecting IGH, which were neither detected by G-banding nor by SKY. The results of the present study suggest the existence of hitherto unknown nonrandom chromosomal changes that may play a role in the pathogenesis of MM. Our findings underline the importance of the combination of banding, SKY, and FISH analyses to increase the accuracy of karyotype interpretation in plasma cell neoplasias.

Chromosome Aberrations↗
PubMed2006

BCL6 alternative breakpoint region break and homozygous deletion of 17q24 in the nodular lymphocyte predominance type of Hodgkin's lymphoma-derived cell line DEV.

DEV is the only cell line derived from nodular lymphocyte predominance type of Hodgkin's lymphoma (NLPHL); however, a comprehensive report about the genetic and immunophenotypic profile of this unique cell line is lacking. We analyzed DEV with respect to immunophenotype and genetic aberrations. The immunostaining revealed positivity for CD45, CD20, CD22, CD79a, IgA2, CD80, CD86, CD74, and BCL6. Cytogenetically, DEV has complex chromosome 3 translocations involving chromosomes 7, 14, and 22. A detailed analysis of the 3q27 breakpoint of the der(3)t(3;14)(p14;q32)t(3;22)(q27;q11.2) revealed a break in the BCL6 alternative breakpoint region. Using array comparative genomic hybridization, a 3-megabase homozygous deletion at 17q24.1-24.2 was identified. Fluorescence in situ hybridization indicated the presence of 2 chromosome 17 homologues, each of which carried a small interstitial deletion. Eight microsatellite markers flanking the homozygously deleted region all showed a homozygous pattern suggesting loss of one of the parental alleles. D17S1809 and D17S1816 could not be amplified using DEV DNA, in keeping with a location within the homozygously deleted segment. In conclusion, DEV has an immunophenotype that is consistent with the neoplastic cells of NLPHL cases, the lymphocytic and histiocytic cells. We demonstrated involvement of the BCL6 gene based on the presence of a breakpoint in the alternative breakpoint region and nuclear staining for BCL6 protein and identified a homozygously deleted region at 17q24.

Cell Line, Tumor↗
PubMed2006

Chromosomal translocations fusing the BCL6 gene to different partner loci are recurrent in primary central nervous system lymphoma and may be associated with aberrant somatic hypermutation or defective class switch recombination.

Primary central nervous system lymphomas (PCNSLs) are diffuse large B cell lymphomas confined to the brain. Only minimal data exist on chromosomal aberrations underlying PCNSLs. We studied 41 PCNSLs by fluorescence in situ hybridization for breakpoints affecting the BCL6 locus in chromosomal band 3q27. Of 37 cases evaluable, 14 (38%) carried a breakpoint in the BCL6 locus. Two of these showed juxtaposition of BCL6 to the IGH locus. In 4 cases, the BCL6 breakpoints were cloned using long-distance inverse polymerase chain reaction. All breakpoints were located within the BCL6 major translocation cluster. The translocation partners were the IGH gene in 14q32.33, the IGL gene in 22q11.22, and the histone 1 H4I gene in 6p22.1. In the fourth case, a deletion in 3q leads to loss of an 837-kb fragment extending from the first intron of BCL6 to the third intron of the lipoma-preferred partner (LPP) gene. This deletion may bring the BCL6 gene under the control of regulatory elements of the LPP gene or the miRNA-28 gene located in intron 4 of LPP. DNA sequence analysis of the junctional sequences provided evidence that aberrant class switch recombination or somatic hypermutation may be involved in the generation of BCL6 translocations.

Base Sequence↗
PubMed2006

Distinct comparative genomic hybridisation profiles in gastric mucosa-associated lymphoid tissue lymphomas with and without t(11;18)(q21;q21).

t(11;18)(q21;q21) occurs specifically in mucosa-associated lymphoid tissue (MALT) lymphoma and the translocation generates a functional API2-MALT1 fusion product that activates nuclear factor (NF)kappaB. t(11;18) positive lymphomas usually lack the chromosomal aberrations and microsatellite alterations frequently seen in the translocation-negative MALT lymphomas. To further understand their genetic differences, we investigated gastric MALT lymphomas with and without t(11;18) by comparative genomic hybridisation. In general, both chromosomal gains and losses were far more frequent in t(11;18)-negative (median = 3.4 imbalances) than t(11;18)-positive cases (median = 1.6 imbalances), with gains being more frequent than losses. Recurrent chromosomal gains involving whole or major parts of a chromosome were seen for chromosomes 3, 12, 18 and 22 (23%, 19%, 19% and 27% respectively). Discrete recurrent chromosomal gains were found at 9q34 (11/26 = 42%). Bioinformatic analysis of genes mapping to 9q34 revealed potential targets. Among them, TRAF2 and CARD9 are known interaction partners of BCL10, playing a role in NFkappaB activation. Interphase fluorescent in situ hybridisation confirmed genomic gain of the TRAF2, CARD9 and MALT1 loci in 5/6 and 2/2 cases showing chromosomal gains at 9q34 and 18q21 respectively. The results further highlight the genetic difference between MALT lymphomas with and without t(11;18). Moreover, our findings suggest that genomic gain of genes that modulate NFkappaB activation, such as MALT1, TRAF2 and CARD9, may play a role in the pathogenesis of the translocation-negative MALT lymphoma.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Aberrant expression of ID2, a suppressor of B-cell-specific gene expression, in Hodgkin's lymphoma.

The global loss of B-cell-specific gene expression is a distinctive feature of the Hodgkin-Reed/Sternberg (HRS) cells of classical Hodgkin's lymphoma (HL). The reasons for this loss remained largely unknown as transcription factors with pleiotropic effects on B-cell-specific gene expression, namely E2A, EBF, and PAX5, are present in primary HRS cells. We show here that ID2, which can inactivate E2A and perhaps PAX5, is not detectable in normal B cells but is strongly and uniformly expressed in HRS cells of all cases of classical HL. Recurrent chromosomal gains of the ID2 gene might contribute to this aberrant expression. Co-immunoprecipitation of E2A with ID2 from HRS-derived cell lines together with the high amount of ID2 relative to the B-cell transcription factors E2A and PAX5 in HRS-derived cell lines and primary HRS cells indicated that aberrant ID2 expression contributes significantly to the loss of the B-cell-specific gene expression in HRS cells. ID2 was also expressed in lymphocyte-predominance HL, mediastinal large B-cell, diffuse large B-cell, and Burkitt's lymphoma, where lower amounts of ID2 relative to E2A and PAX5 compared with HRS cells might prevent a global down-regulation of B-cell-specific genes and ID2 may contribute to lymphomagenesis in other ways.

B-Lymphocytes↗