Burkhard Helmke

Universität Heidelberg, Heidelburg, Baden-Württemberg, Germany

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Publications (93)282.35 Total impact

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    ABSTRACT: Uterine fibroids rank among the most frequent symptomatic human tumors at all. Recent data suggest that mutations of the mediator subcomplex 12 gene (MED12) and rearrangements of the gene-encoding high-mobility group protein AT-hook 2 (HMGA2) characterize major genetic subtypes of these tumors, which, for example, differ by their average size. Herein, we have investigated a total of 289 fibroids from 120 patients. Of these fibroids, 256 were fully genetically analyzed. Of the latter group, 20 (7.8%) fibroids had a chromosomal rearrangement of 12q14-15 reflecting a rearranged allele of HMGA2 and 179 (69.9%) fibroids had a mutation of MED12. The remaining tumors had either another genetic abnormality or no detectable abnormality at all. We were able to demonstrate that tumors of both groups also display striking differences of their frequency in individual patients. Whereas 70.0% (14/20) HMGA2-mutated fibroids made their appearance as solitary nodules, 85.5% (153/179) MED12-mutated fibroids occurred as multiple nodules as a rule of independent clonal origin, as reflected by different MED12 mutations. These findings are likely to point to a different pathogenesis of both types of fibroids. In the predominant of these groups so far, an unknown "mutator" may cause independent mutations of MED12, resulting in an independent clonal outgrowth of nodules. Furthermore, the low but existing risk of MED12-mutated fibroids to undergo malignant transformation after a leiomyoma-STUMP (smooth muscle tumors of uncertain malignant potential)-leiomyosarcoma sequence excludes the latter mutation as a suitable stand-alone marker for benign growth.
    09/2014;
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    ABSTRACT: Epitheloid leiomyoma is a rare subtype of benign smooth muscle tumors. Herein, we present the results of classical cytogenetics, MED12 mutation analysis, and copy number variation array evaluation in one such case. Whereas cytogenetic did not show evidence for clonal chromosome abnormalities and no MED12 mutation in the "fibroid hot spot" region was detected, array hybridization revealed multiple abnormalities. Most noteworthy, almost all chromosomes showed copy-number neutral loss of heterozygosity. As examples of further abnormalities, trisomies of chromosomes 8, 12, 20, and X were noted. The data presented suggest a near-haploid karyotype of the tumor as the initial genetic alteration followed by secondary duplications of large parts of the genome. The absence of any clonal karyotypic alterations after performing classical cytogenetics is likely explained by a reduced ability of the tumor cells to proliferate in vitro. However, to the best of our knowledge this is the first report of an uterine leiomyoma showing extended uniparental disomy. It remains to be determined if this is a more common phenomenon in epithelioid leiomyomas or even subsets of "ordinary" leiomyomas.
    Molecular Cytogenetics 03/2014; 7(1):19. · 2.66 Impact Factor
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    ABSTRACT: Uterine leiomyomas (UL) are the most frequent symptomatic human tumors. Nevertheless, their molecular pathogenesis is not yet fully understood. To learn more about the biology of these common neoplasms and their response to treatment, cell cultures derived from UL are a frequently used model system, but until recently appropriate genetic markers confirming their origin from the tumor cell population were lacking for most UL, i.e., those not displaying karyotypic abnormalities. The identification of MED12 mutations in the majority of UL makes it possible to trace the tumor cell population during in vitro passaging in the absence of cytogenetic abnormalities. The present study is addressing the in vitro survival of cells carrying MED12 mutations and its association with karyotypic alterations. The results challenge numerous in vitro studies into the biology and behavior of leiomyomas. Cells of one genetic subtype of UL, i.e., those with rearrangements of the high mobility AT-hook 2 protein gene (HMGA2), seem to be able to proliferate in vitro for many passages whereas tumor cells from the much more frequent MED12-mutated lesions barely survive even the first passages. Apparently, for the most frequent type of human UL no good in vitro model seems to exist because cells do not survive culturing. On the other hand, this inability may point to an Achilles' heel of this type of UL. © 2014 Wiley Periodicals, Inc.
    Genes Chromosomes and Cancer 01/2014; · 3.55 Impact Factor
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    ABSTRACT: Glioblastoma, an aggressive brain tumor, has a poor prognosis and a high risk of recurrence. An improved chemotherapeutic approach is required to complement radiation therapy. Gold(I) complexes bearing phosphole ligands are promising agents in the treatment of cancer and disturb the redox balance and proliferation of cancer cells by inhibiting disulfide reductases. Here, we report on the antitumor properties of the gold(I) complex 1-phenyl-bis(2-pyridyl)phosphole gold chloride thio-β-D-glucose tetraacetate (GoPI-sugar), which exhibits antiproliferative effects on human (NCH82, NCH89) and rat (C6) glioma cell lines. Compared to carmustine (BCNU), an established nitrosourea compound for the treatment of glioblastomas that inhibits the proliferation of these glioma cell lines with an IC50 of 430μM, GoPI-sugar is more effective by two orders of magnitude. Moreover, GoPI-sugar inhibits malignant glioma growth in vivo in a C6 glioma rat model and significantly reduces tumor volume while being well tolerated. Both the gold(I) chloro- and thiosugar-substituted phospholes interact with DNA albeit more weakly for the latter. Furthermore, GoPI-sugar irreversibly and potently inhibits thioredoxin reductase (IC50 4.3 nM) and human glutathione reductase (IC50 88.5 nM). However, treatment with GoPI-sugar did not significantly alter redox parameters in the brain tissue of treated animals. This might be due to compensatory upregulation of redox-related enzymes but might also indicate that the antiproliferative effects of GoPI-sugar in vivo are rather based on DNA interaction and inhibition of topoisomerase I than on the disturbance of redox equilibrium. Since GoPI-sugar is highly effective against glioblastomas and well tolerated, it represents a most promising lead for drug development. This article is part of a Special Issue entitled: Thiol-Based Redox Processes.
    Biochimica et Biophysica Acta 01/2014; · 4.66 Impact Factor
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    ABSTRACT: In pleomorphic adenomas of the salivary glands (PASG) recurrent chromosomal rearrangements affecting either 8q12 or 12q14∼15 lead to an overexpression of the genes of the genuine transcription factor PLAG1 or the architectural transcription factor HMGA2, respectively. Both genes are also affected by recurrent chromosomal rearrangements in benign adipocytic tumors as e. g. lipomas and lipoblastomas. Herein, we observed a strong correlation between the expression of HMGA2 and PLAG1 in 14 benign and 23 malignant thyroid tumors. To address the question if PLAG1 can be activated by HMGA2, the expression of both genes was quantified in 32 uterine leiomyomas 17 of which exhibited an overexpression of HMGA2. All leiomyomas with HMGA2 overexpression also revealed an activation of PLAG1 in the absence of detectable chromosome 8 abnormalities affecting the PLAG1 locus. To further investigate if the overexpression of PLAG1 is inducible by HMGA2 alone, HMGA2 was transiently overexpressed in MCF-7 cells. An increased PLAG1 expression was observed 24 and 48 h after transfection. Likewise, stimulation of HMGA2 by FGF1 in adipose tissue-derived stem cells led to a simultaneous increase of PLAG1 mRNA. Altogether, these data suggest that HMGA2 is an upstream activator of PLAG1. Accordingly, this may explain the formation of tumors as similar as lipomas and lipoblastomas resulting from an activation of either of both genes by chromosomal rearrangements.
    PLoS ONE 01/2014; 9(2):e88126. · 3.53 Impact Factor
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    ABSTRACT: BACKGROUND: Deletions of the gene encoding mediator subcomplex 12 (MED12) in human smooth muscle tumors rank among the most frequent genomic alterations in human tumors at all. In a minority of these cases, small deletions are found. In an attempt to delineate key features of the deletions aimed at a better understanding of the molecular pathogenesis of uterine smooth muscle tumors we have analyzed 70 MED12 deletions including 46 cases from the literature and 24 own unpublished cases. RESULTS: The average length of the deletions was 18.7 bp ranging between 2 bp and 43 bp. While in general multitudes of 3 clearly dominated leaving the transcript in frame, deletions of 21, 24, 30, and 33 nucleotides were clearly underrepresented. Within the DNA segment affected deletion breakpoints were not randomly distributed. Most breakpoints clustered within the center of the segment where two peaks of breakpoint clusters could be distinguished. Interestingly, one of these clusters coincides with the loop of a putative folded non-B DNA structure whereas a much lower number of breaks noted in the 5' and 3' stem of the structure forming an intramolecular B-helix. The second cluster mainly consisting of 3' breaks was located in a region downstream adjacent to the stem. CONCLUSION: The present study describes for the first time main characteristics of MED12 deletions occurring in smooth muscle tumors. Interestingly, the non-random distribution of breakpoints within the deletion hotspot region may point to a role of non-canonical DNA structures for the occurrence of these mutations and the molecular pathogenesis of uterine smooth muscle tumors, respectively.
    Molecular Cytogenetics 06/2013; 6(1):23. · 2.66 Impact Factor
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    ABSTRACT: BACKGROUND: Amyloidosis is a life-threatening protein misfolding disease and affects cardiac tissue, leading to heart failure, myocardial ischemia and arrhythmia. Amyloid deposits result in oxidative stress, inflammation and apoptosis. The purpose of this study was to examine the role of innate defense components, i.e., Deleted in Malignant Brain Tumors 1 (DMBT1) and the complement system, in different types of cardiac amyloidosis. METHODS: Expression of DMBT1 and of the complement proteins C1q, C3d and C4d in cardiac specimens of patients with different types of amyloidosis were determined by immunohistochemistry and correlated with amyloid deposits stained by Congo red dye. RESULTS: Strong DMBT1 staining adjacent to amyloid deposits was detected in different amyloidosis types, depending on the extent of the deposits. DMBT1 is localized in the endomysium and perimysium, in the endocardium, in the myocytes and in endothelial cells of affected transmural vessels. C1q, C3d and C4d were detected in the amyloid deposits but also in the endomysium and perimysium, in some myocytes, in endothelial cells, in the endocardium, and around the amyloid deposits. CONCLUSIONS: Up-regulated DMBT1 and complement activation in cardiac amyloidosis may be part of the activated pathways induced by protein aggregation and the consecutive inflammatory reaction.
    Cardiovascular pathology: the official journal of the Society for Cardiovascular Pathology 12/2012; · 1.63 Impact Factor
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    ABSTRACT: MicroRNAs of the chromosome 19 microRNA cluster (C19MC) are known to be abundantly expressed in the placenta. Their genes are located on the long arm of chromosome 19 and seem to be part of a large imprinted region. Although the data available so far suggest important functions in the placenta, no data are available on their general expression patterns in cultures of placenta-derived mesenchymal stromal cells (PDMSC). Surprisingly, qRT-PCR on tissue cultures from first-trimester and term placenta mesenchymal stromal cells showed an abundant expression of the cluster members miR-517a-3p, miR-519a-3p, and miR-520c-3p. Accordingly, analyses of methylation patterns suggested that these cells had escaped methylation and epigenetic silencing, respectively, of the paternal allele. This was confirmed by the results of treatment of chorionic villous stromal cells by the demethylating agent 5-Aza-2'-deoxycytidine. Our results offer clear evidence that, in contrast to what is suggested in previous papers, members of C19MC are highly expressed in PDMSC indicating that their placenta-specific functions are not restricted to the trophoblast.
    Biochemical and Biophysical Research Communications 05/2012; 422(3):411-6. · 2.28 Impact Factor
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    ABSTRACT: The expression of high mobility group protein AT-hook2 (HMGA2) indicates a worse prognosis in many epithelial malignancies, such as colon cancer. The present study addresses methodological aspects, as well as the genetic background, of the HMGA2 expression in colon cancer. Samples of 38 colon carcinomas were studied for the expression of HMGA2 by quantitative Real-Time PCR (qRT-PCR). In selected cases, immunohistochemistry (IHC) was also performed. The overexpression of HMGA2, compared to adjacent mucosa, is not consistent among colon carcinomas: Only a minority of carcinomas strongly overexpressed HMGA2, but in no more than 50% of the tumors did the expression exceed the average value in mucosa samples. qRT-PCR clearly reveals a continuum between cases with high and low expression. For HMGA2-based risk assessment, continuous rather than discontinuous models seem to be most appropriate. However, in daily practice, IHC seems to be a suitable method to stratify for high-risk patients.
    Anticancer research 05/2012; 32(5):1589-93. · 1.71 Impact Factor
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    ABSTRACT: Recurrent chromosomal alterations are found in roughly 20% of all uterine fibroids but in the majority cytogenetic changes are lacking. Recently, mutations of the gene mediator subcomplex 12 (MED12) have been detected in a majority of fibroids but no information is available whether or not they co-occur with cytogenetic subtypes as, e.g., rearrangements of the genes encoding high mobility group AT-hook (HMGA) proteins. In a total of 80 cytogenetically characterized fibroids from 50 patients, we were not only able to confirm the frequent occurrence of MED12 mutations but also to stratify two mutually exclusive pathways of leiomyomagenesis with either rearrangements of HMGA2 reflected by clonal chromosome abnormalities affecting 12q14~15 or by mutations affecting exon 2 of MED12. On average the latter mutations were associated with a significantly smaller tumor size. However, G>A transitions of nucleotides c.130 or c.131 correlate with a significantly larger size of the fibroids compared to other MED12 mutations thus explaining the high prevalence of the former mutations among clinically detectable fibroids. Interestingly, fibroids with MED12 mutations expressed significantly higher levels of the gene encoding wingless-type MMTV integration site family, member 4 (WNT4). Based on these findings and data from the literature, we hypothesize that estrogen and the mutated MED12 cooperate in activating the Wnt pathway which in turn activates β-catenin known to cause leiomyoma-like lesions in a mouse model. The occurrence of a "fibroid-type mutation" in a rare histologic subtype of endometrial polyps suggests that this mechanism is not confined to uterine leiomyomas.
    International Journal of Cancer 01/2012; 131(7):1528-36. · 6.20 Impact Factor
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    ABSTRACT: Spontaneous cessation of growth is a frequent finding in uterine fibroids. Increasing evidence suggests an important role of cellular senescence in this growth control. Deciphering the underlying mechanisms of growth control that can be expected not only to shed light on the biology of the tumors but also to identify novel therapeutic targets. We have analyzed uterine leiomyomas and matching normal tissue for the expression of p14Arf and used explants to see if reducing the MDM2 activity using the small-molecule inhibitor nutlin-3 can induce p53 and activate genes involved in senescence and/or apoptosis. For these studies quantitative real-time RT-PCR, Western blots, and immunohistochemistry were used. Statistical analyses were performed using the student's t test. An in depth analysis of 52 fibroids along with matching myometrium from 31 patients revealed in almost all cases a higher expression of p14Arf in the tumors than in the matching normal tissue. In tissue explants, treatment with the MDM2 inhibitor nutlin-3 induced apoptosis as well as senescence as revealed by a dose-dependent increase of the expression of BAX as well as of p21, respectively. Simultaneously, the expression of the proliferation marker Ki-67 drastically decreased. Western-blot analysis identified an increase of the p53 level as the most likely reason for the increased activity of its downstream markers BAX and p21. Because as a rule fibroids express much higher levels of p14Arf, a major negative regulator of MDM2, than matching myometrium it was then analyzed if fibroids are more sensitive against nutlin-3 treatment than matching myometrium. We were able to show that in most fibroids analyzed a higher sensibility than that of matching myometrium was noted with a corresponding increase of the p53 immunopositivity of the fibroid samples compared to those from myometrium. The results show that uterine fibroids represent a cell population of advanced cellular age compared to matching myometrium. Moreover, the data point to members of the p53-network as to potential novel therapeutic targets for the treatment of uterine fibroids.
    BMC Women's Health 01/2012; 12:2. · 1.66 Impact Factor
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    ABSTRACT: BMP4 has been linked to early steps of adipocyte lineage differentiation but only little is known about its corresponding downstream pathways. Herein, we have investigated whether or not the expression of high mobility group protein HMGA2, another protein linked to proliferation and differentiation within the process of adipogenesis, may be influenced by BMP4 signaling in adipose tissue derived stem cells. Compared to FGF1, a strong inducer of HMGA2 in immortalized pre-adipocytes, BMP4 was found moderately to induce the HMGA2 mRNA expression in serum starved adipose tissue derived stem cells and myometrial cells. In contrast, no such activity was noted in canine bone marrow derived mesenchymal stem cells. As to adipocyte lineage differentiation the functions of BMP4 and HMGA2 mechanistically overlap. Thus, we propose that in adipose tissue BMP4 acts in part by activating HMGA2 making this architectural transcription factor one of the major downstream players in that system.
    Cytokine 12/2011; 56(3):811-6. · 2.52 Impact Factor
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    ABSTRACT: Cellular expression of heparanase, a degrading enzyme of the extracellular matrix, is associated with poorer prognosis in several cancers. The present analysis, has studied the role of heparanase in tumour growth and clinical outcome in patients with head and neck squamous cell carcinoma (HNSCC). We analysed the cellular expression of the active form of heparanase in 71 human HNSCCs, using immunohistochemistry. The results were compared with clinicopathological data and, in 65 cases with immunoreactivity for the proliferation marker, MIB1. Cellular heparanase expression was detected in 41 of 71 (57.74%) cases; in particular, UICC IV-stage tumours showed high heparanase levels. Heparanase was localized mainly in the cytoplasm and, to a lesser extent, at the cell membrane. High levels of heparanase were significantly correlated with an almost four-fold decrease in MIB1 labelling (P = 0.006). Comparison with clinical outcome by multivariate analysis revealed that patients with high-level heparanase expression had prolonged overall survival (P = 0.029). Although heparanase was mainly found in late-stage HNSCCs, cellular heparanase expression in HNSCCs was associated with prolonged overall survival. We propose that the proliferation-reducing effect of high heparanase levels might outweigh the tumour-promoting effects of heparanase, especially in advanced tumours.
    Histopathology 05/2011; 58(6):944-52. · 2.86 Impact Factor
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    ABSTRACT: To address the influence of genes involved in stem cell self-renewal and senescence on the growth of leiomyoma cells in vitro and to explore possible therapeutic implications of a targeted disruption of the p53-murine double minute 2 (MDM2) interaction. Gene expression studies (qRT-PCR) of fibroid tissue and cells; β-galactosidase stain and qRT-PCR after antagonizing MDM2. In fibroid cells, expression of HMGA2 decreased with passaging while that of p14(Arf) increased. Expression of these markers significantly positively, and negatively, respectively, influenced proliferation. Administration of nutlin-3, an MDM2 antagonist, induced cellular senescence and increased the expression of BAX. This, along with a significant correlation between p14(Arf) and BAX expression in native fibroids, suggests that p14(Arf) triggers senescence as well as apoptosis. p14(Arf) and HMGA2 seem to play a pivotal role in controlling the growth of fibroid cells. Antagonizing MDM2 induces senescence, as well as apoptosis, and may offer a chance to treat fibroids.
    Anticancer research 03/2011; 31(3):753-61. · 1.71 Impact Factor
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    ABSTRACT: Celiac disease (CD) is an inflammatory disorder associated with an increased risk of small bowel adenocarcinoma. Recent studies have demonstrated aberrant CpG island methylation (CIM) in chronic inflammation, aging and cancer. We hypothesized that CIM may link CD to small bowel carcinogenesis. We determined microsatellite instability (MSI), CIM, and expression of MLH1 and MGMT in 3 CD-associated small bowel carcinomas and corresponding non-neoplastic mucosa. The results were compared to those of small bowel mucosa from CD patients without carcinoma and 20 small bowel carcinomas from a non-CD origin. A high level CIM/MSI phenotype was found in all of the 3 CD-associated carcinomas and was associated with loss of MLH1 expression due to hypermethylation of the MLH1 promoter. This phenotype was noted in only 2 of the 20 investigated non-CD-associated carcinomas. Low-level CIM was already detectable in 9 of the 12 non-neoplastic mucosa samples of CD patients and in non-CD-associated carcinomas of elderly patients. In conclusion, our data reveal that the high-level CIM/MSI pathway is typical of CD-associated small bowel carcinomas and indicate that aberrant CpG island methylation links CD and carcinogenesis. The data further suggest that CD should be considered in patients with small bowel adenocarcinoma, particularly when the tumors display MSI.
    Oncology Reports 12/2010; 24(6):1535-9. · 2.30 Impact Factor
  • Fertility and sterility 11/2010; 94(6):e79; author reply e80. · 3.97 Impact Factor
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    ABSTRACT: Anorectal melanomas (AMs) are very rare and highly malignant tumors that are often diagnosed in advanced stages. After the differentiation between cutaneous melanoma (CM) and AM on the molecular level based on the presence of BRAF mutations, further modes of differentiation opened up, such as the recently discovered immunohistologically relevant protein deleted in malignant brain tumors 1 (DMBT1). Over the past several years, increasingly specific therapies have been developed on the basis of new therapy principles. Tyrosin kinase receptors such as Her2 and EGFR have been awarded a large role in this context. The goal of this study was to examine AMs for a possible expression or overexpression of these markers. Expression analyses of Her2 and EGFR were performed immunohistologically on 25 primary AMs. An overexpression of Her2 (score: 3+) was found in one AM from a 68-year-old female patient among these samples. In contrast, EGFR expression was not found in any of the AMs. The results presented here show that isolated cases of AM may benefit from an additive Her2-directed therapy, as the overexpression of Her2 was found in one of our AM patients.
    Melanoma research 10/2010; 20(5):431-4. · 2.06 Impact Factor
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    ABSTRACT: The in vitro growth of cells from uterine fibroids is characterized by an early onset of senescence. Often, an even lower growth potential than that of matching myometrial cells is noted. Also, the tremendous differences in the expression of the high mobility group protein HMGA2 seen when comparing fibroids of different genetic subtypes are surprisingly not reflected by significant differences in their growth potential in vitro. We aimed to evaluate possible changes of the HMGA2 expression level between the native tissue and cell cultures, so we performed quantitative real-time polymerase chain reaction studies that revealed a marked decrease of the HMGA2 mRNA in culture in those cases with overexpression of HMGA2. In the two cases initially showing the highest expression, it decreased by approximately 97%. Associated with the decrease of HMGA2 was a clearly increased expression of the senescence-associated p19(Arf). Together, these findings explain the similar behavior of cell cultures from fibroids of different genetic subgroups and may also offer an explanation for the early onset of in vitro senescence in these cell cultures.
    Cancer genetics and cytogenetics 10/2010; 202(1):53-7. · 1.54 Impact Factor

Publication Stats

1k Citations
282.35 Total Impact Points

Institutions

  • 2009–2014
    • Universität Heidelberg
      • • Institute of Papyrology
      • • Institute of Pathology (Mannheim)
      Heidelburg, Baden-Württemberg, Germany
  • 2010–2012
    • Universität Bremen
      • Center for Human Genetics and Genetic Counselling (ZHG)
      Bremen, Bremen, Germany
  • 2002–2009
    • German Cancer Research Center
      • Division of Molecular Genome Analysis
      Heidelberg, Baden-Wuerttemberg, Germany