German Cancer Research Center Research Articles

Induction and transport of Wnt 5a during macrophage-induced malignant invasion is mediated by two types of extracellular vesicles

Recently, we have shown that macrophage (MΦ)-induced invasion of breast cancer cells requires upregulation of Wnt 5a in MΦ leading to activation of β-Catenin-independent Wnt signaling in the tumor cells. However, it remained unclear, how malignant cells induce Wnt 5a in MΦ and how it is transferred back to the cancer cells. Here we identify two types of extracellular particles as essential for this intercellular interaction in both directions. Plasma membrane-derived microvesicles (MV) as well as exosomes from breast cancer cells, although biologically distinct populations, both induce Wnt 5a in MΦ. In contrast, the particle-free supernatant and vesicles from benign cells, such as platelets, have no such effect. Induction is antagonized by the Wnt inhibitor Dickkopf-1. Subsequently, Wnt 5a is shuttled via responding MΦ-MV and exosomes to the tumor cells enhancing their invasion. Wnt 5a export on both vesicle fractions depends at least partially on the cargo protein Evenness interrupted (Evi). Its knockdown leads to Wnt 5a depletion of both particle populations and reduced vesicle-mediated invasion. In conclusion, MV and exosomes are critical for MΦ-induced invasion of cancer cells since they are responsible for upregulation of MΦ-Wnt 5a as well as for its delivery to the recipient cells via a reciprocal loop. Although of different biogenesis, both populations share common features regarding function and Evi-dependent secretion of non-canonical Wnts.

Critical role of zinc finger protein 521 in the control of growth, clonogenicity and tumorigenic potential of medulloblastoma cells

The stem cell-associated transcription co-factor ZNF521 has been implicated in the control of hematopoietic, osteo-adipogenic and neural progenitor cells. ZNF521 is highly expressed in cerebellum and in particular in the neonatal external granule layer that contains candidate medulloblastoma cells-of-origin, and in the majority of human medulloblastomas. Here we have explored its involvement in the control of human and murine medulloblastoma cells. The effect of ZNF521 on growth and tumorigenic potential of human medulloblastoma cell lines as well as primary Ptc1-/+ mouse medulloblastoma cells was investigated in a variety of in vitro and in vivo assays, by modulating its expression using lentiviral vectors carrying the ZNF521 cDNA, or shRNAs that silence its expression. Enforced overexpression of ZNF521 in DAOY medulloblastoma cells significantly increased their proliferation, growth as spheroids and ability to generate clones in single-cell cultures and semisolid media, and enhanced their migratory ability in wound-healing assays. Importantly, ZNF521-expressing cells displayed a greatly enhanced tumorigenic potential in nude mice. All these activities required the ZNF521 N-terminal motif that recruits the nucleosome remodeling and histone deacetylase complex, which might therefore represent an appealing therapeutic target. Conversely, silencing of ZNF521 in human UW228 medulloblastoma cells that display high baseline expression decreased their proliferation, clonogenicity, sphere formation and wound-healing ability. Similarly, Zfp521 silencing in mouse Ptc1-/+ medulloblastoma cells drastically reduced their growth and tumorigenic potential. Our data strongly support the notion that ZNF521, through the recruitment of the NuRD complex, contributes to the clonogenic growth, migration and tumorigenicity of medulloblastoma cells.

MO‐E‐500‐01: Point and Counter Point Debate: The More Important Heavy Charged Particle Radiotherapy of the Future Is More Likely to Be with Heavy Ions Rather Than Protons

Over the past decade there has been considerable interest and progress in the use of heavy ions such as protons and carbon in an attempt to improve the effectiveness of radiotherapy. Both have a physical advantage over photons because of their Bragg peaks, but only heavier ions such as carbon have a potential biological advantage. This has led to the claim that the most important heavy charged particle radiotherapy of the future is more likely to be with heavy ions rather than protons, and this is the premise argued in this debate.Arguments FOR the Proposition include:• relative to protons, heavier ions exhibit reduced lateral scattering, reduced range straggling, and increased dose to the tumor compared to the entrance dose• accurate positioning of the Bragg peak can be verified by in vivo measurements with heavier ions but not with protons• with heavier ions there is increased RBE in the tumor compared to normal tissues• with heavier ions there is increased effectiveness in hypoxic tumorsArguments AGAINST the Proposition include:• proton facilities cost 2–3 times less than those for heavier ions• cost and size of isocentric gantries for protons are much less• there are large uncertainties in the RBE in the heavier ion Bragg peak• with protons, there is the potential for reoxygenation during a course of therapy• due to fragmentation, heavier ion beams exhibit a tail of high‐LET radiation beyond the Bragg peakArguing for the Proposition is Oliver Jakel, Ph.D. Dr. Jakel completed his Ph.D. in Theoretical Physics at the University Erlangen, Germany in 1994. He then moved to the Department for Medical Physics, German Cancer Research Center (DKFZ), Heidelberg, where he is currently Full Professor and Director of the Heidelberg Ion Beam Therapy Facility of the University Hospital. Arguing against the Proposition is Alfred R. Smith, Ph.D. Dr. Smith obtained his Ph.D. in Physics from Texas Tech University, Lubbock, TX, in 1970. After completing a Postdoctoral Fellowship in Medical Physics at The University of Texas, M. D. Anderson Hospital and Tumor Institute, he held faculty positions at several institutions until he moved back to M. D. Anderson in 2002 as a Full Professor and Director of Proton Therapy Development, a position he held until 2010.Learning Objectives:1. To understand why heavy ions might be more suitable than photons for the treatment of some cancers2. To understand the relative benefits of therapy with heavier ions compared with protons3. To understand the relative benefits of therapy with protons compared with heavier ions

Methylation of the TERT promoter and risk stratification of childhood brain tumours: an integrative genomic and molecular study

Identification of robust biomarkers of malignancy and methods to establish disease progression is a major goal in paediatric neuro-oncology. We investigated whether methylation of the TERT promoter can be a biomarker for malignancy and patient outcome in paediatric brain tumours. For the discovery cohort, we used samples obtained from patients with paediatric brain tumours and individuals with normal brain tissues stored at the German Cancer Research Center (Heidelberg, Germany). We used methylation arrays for genome-wide assessment of DNA. For the validation cohort, we used samples obtained from several tissues for which full clinical and follow-up data were available from two hospitals in Toronto (ON, Canada). We did methylation analysis using quantitative Sequenom and pyrosequencing of an identified region of the TERT promoter. We assessed TERT expression by real-time PCR. To establish whether the biomarker could be used to assess and predict progression, we analysed methylation in paired samples of tumours that transformed from low to high grade and from localised to metastatic, and in choroid plexus tumours of different grades. Finally, we investigated overall survival in patients with posterior fossa ependymomas in which the identified region was hypermethylated or not. All individuals responsible for assays were masked to the outcome of the patients. Analysis of 280 samples in the discovery cohort identified one CpG site (cg11625005) in which 78 (99%) of 79 samples from normal brain tissues and low-grade tumours were not hypermethylated, but 145 (72%) of 201 samples from malignant tumours were hypermethylated (>15% methylated; p<0.0001). Analysis of 68 samples in the validation cohort identified a subset of five CpG sites (henceforth, upstream of the transcription start site [UTSS]) that was hypermethylated in all malignant paediatric brain tumours that expressed TERT but not in normal tissues that did not express TERT (p<0.0001). UTSS had a positive predictive value of 1.00 (95% CI 0.95-1.00) and a negative predictive value of 0.95 (0.87-0.99). In two paired samples of paediatric gliomas, UTSS methylation increased during transformation from low to high grade; it also increased in two paired samples that progressed from localised to metastatic disease. Two of eight atypical papillomas that had high UTSS methylation progressed to carcinomas, while the other six assessed did not progress or require additional treatment. 5-year overall survival was 51% (95% CI 31-71) for 25 patients with hypermethylated UTSS posterior fossa ependymomas and 95% (86-100) for 20 with non-hypermethylated tumours (p=0.0008). 5-year progression-free survival was 86% (68-100) for the 25 patients with non-hypermethylated UTSS tumours and 30% (10-50) for those with hypermethylated tumours (p=0.0008). Hypermethylation of the UTSS region in the TERT promoter is associated with TERT expression in cancers. In paediatric brain tumours, UTSS hypermethylation is associated with tumour progression and poor prognosis. This region is easy to amplify, and the assay to establish hypermethylation can be done on most tissues in most clinical laboratories. Therefore the UTSS region is a potentially accessible biomarker for various cancers. The Canadian Institute of Health Research and the Terry Fox Foundation.

Images as tools. On visual epistemic practices in the biological sciences

Contemporary visual epistemic practices in the biological sciences raise new questions of how to transform an iconic data measurements into images, and how the process of an imaging technique may change the material it is ‘depicting’. This case-oriented study investigates microscopic imagery, which is used by system and synthetic biologists alike. The core argument is developed around the analysis of two recent methods, developed between 2003 and 2006: localization microscopy and photo-induced cell death. Far from functioning merely as illustrations of work done by other means, images can be determined as tools for discovery in their own right and as objects of investigation. Both methods deploy different constellations of intended and unintended interactions between visual appearance and underlying biological materiality. To characterize these new ways of interaction, the article introduces the notions of ‘operational images’ and ‘operational agency’. Despite all their novelty, operational images are still subject to conventions of seeing and depicting: Phenomena emerging with the new method of localization microscopy have to be designed according to image traditions of older, conventional fluorescence microscopy to function properly as devices for communication between physicists and biologists. The article emerged from a laboratory study based on interviews conducted with researchers from the Kirchhoff-Institute for Physics and German Cancer Research Center (DKFZ) at Bioquant, Heidelberg, in 2011.

NK cells – Versatile tools for viral defense and cancer treatment

About 150 NK cell researchers met at the German Cancer Research Center (DKFZ) in Heidelberg, Germany from 26–28 September 2012 for the Natural Killer Cell Symposium which was organized by the NK cell study group of the German Society for Immunology (DGfI) and sponsored by the European Journal of Immunology (EJI), the European Federation of Immunological Societies (EFIS) and the DGfI. The meeting was a forum for the discussion of the function and regulation of these fascinating innate immune cells and the opportunities for the transfer of this knowledge to cancer immunotherapy.

The challenges of understanding cancer of unknown primary

1 Division of Molecular Genetic Epidemiology, German Cancer Research Centre (DKFZ), Heidelberg, Germany 2 Center for Primary Health Care Research, Lund University, Malm€ o, Sweden 3 Stanford Prevention Research Center, Stanford University School of Medicine, Palo Alto, CA 4 Cancer Gene Therapy Group, Transplantation Laboratory and Department of Pathology, Haartman Institute, University of Helsinki, Helsinki, Finland

Pregnancy hormone levels linked to HR‐negative breast cancer risk

Higher amounts of the pregnancy hormones estradiol and progesterone were associated with a greater risk of hormone receptor (HR)-negative breast cancer before age 50 years, according to a study by German researchers. The nested, casecontrol study was presented at the American Association for Cancer Research (AACR) International Conference on Frontiers in Cancer Prevention Research held in October 2012. Investigators assessed the effects of hormone exposure during early pregnancy and its possible effect on maternal breast cancer. Annekatrin Lukanova, MD, PhD, associate professor at the German Cancer Research Center in Heidelberg, says that the association between pregnancy and breast cancer risk is complex and that researchers do not fully understand the biological mechanisms for the protective effect of childbearing on cancer risk. She and colleagues used the Northern Sweden Maternity cohort to conduct the study, which compared 417 controls with 223 women who had donated blood samples during their first trimester of pregnancy and who later developed breast cancer. About three-quarters of the women with diagnosed cancer had HR-positive cancer. The 2 groups of hormones that researchers examined were: estradiol, estrone, and progesterone, which increase substantially with pregnancy progression; and testosterone and insulin growth factor-1 (IGF-1). Researchers found that circulating concentrations of IGF-1 are directly associated with risk for HRpositive breast cancer, which is in line with studies of women who were not pregnant. At the same time, they found a higher risk of HR-negative breast cancer diagnosed before age 50 years affiliated with higher levels of estradiol and progesterone. Because the study was small and the hormones were only measured during the first trimester of pregnancy, further and larger studies are needed to better understand the association of pregnancy hormones with the risk for hormone-defined maternal breast cancer, says Dr. Lukanova.

The Response of 2D TL Foils After Doses of Co-60 Gamma-ray, 6 MV X-ray and 60 MeV Proton Beams Applied in Radiotherapy

A Two-dimensional Thermoluminescence Dosimetry (2D TLD) system has been developed at the Institute of Nuclear Physics (IFJ PAN) in Krakow, Poland. The system consists of TLD foils and of a TL reader equipped with a 200 200 mm 2 heater and a CCD camera. The foils contain LiF:Mg,Cu,P powder mixed with ETFE polymer pressed at high temperature. The CCD camera is able to read out 2D images of TL light in real time, and the re-usable foils are flexible, mechanically stable and waterresistant. We have determined the response of our 2D TL foils after doses of Co-60 gamma-ray, 6MV X-ray and 60MeV proton radiotherapy beams, over the range between 1 and 30Gy, as required in clinical dosimetry. In measurements, performed at the German Cancer Research Center (DKFZ), at the Heidelberg Ion-Beam Therapy Center (HIT) and at the IFJ PAN, we compared the dose response of our foils with those of Kodak R EDR2 and Gafchromic R EBT films. We find both the sensitivity (signal per dose) and the characteristic (saturation) dose of our 2D TL foils to be higher than those of the EDR2 and EBT films, for all beam modalities tested.

BiotecVisions 2013, January

BiotecVisions 2013, January

Population Level Survival of Patients with Chronic Myeloid Leukemia in Germany in the Early 21st Century

AbstractAbstract 759 Background:Chronic myeloid leukemia (CML) is a rare but highly treatable form of leukemia. Recent advances in the treatment of CML have dramatically improved survival in clinical trials. Prior work using the Surveillance, Epidemiology, and End Results (SEER) database in the United States (US) suggest that survival on the population level has increased, but still lags behind that seen in clinical trials1. In the past, evaluation of population level survival in other countries has been hampered by the lack of high quality data from a large enough population to evaluate trends in survival in this rare disease. Recently, a collaborative effort between the German Cancer Research Center and several German cancer registries has provided the chance to evaluate population level survival for rare cancers. Here, we examine survival for patients with CML in Germany in the early 21st century, comparing it to survival in the US. Methods:Data was extracted from the SEER13 database in the US and 13 epidemiologic cancer registries in Germany. Patients diagnosed with CML between 1997 and 2006 were included in the analysis. Data quality analysis showed an excessive level of cases diagnosed by death certificate only in older patients (age 70+) so analysis was restricted to patients age 15–69. Patients diagnosed by death certificate only were excluded from the analysis. Period analysis was used to provide the most up-to-date possible estimates of five year relative survival. Results:1848 cases of CML were identified in the German database and 2910 cases in the SEER database. More cases were seen in younger patients in the US, whereas each age group was roughly equal in size in Germany (see table). Five year relative survival was 68.7% overall in Germany and 72.7% in the US. Survival was higher in the US for all age groups except for age 15–39, but the difference was only significant for ages 50–59, where a difference of 10.2 percentage units was seen (see table). Survival decreased with age, ranging from 83.1% and 81.9%, respectively, in Germany and the US for patients age 15–39 to 54.2% and 54.5%, respectively, in patients age 65–69. Conclusions:Estimated five year survival increased in the US compared to previous estimates of survival in the 2000–2004 period1. Five year survival estimates were higher in the US than in Germany overall, but the difference was only significant for age 50–59. These results are the first detailing survival in CML on the population level in Germany and providing an international comparison. Survival did not equal that seen in clinical trials of tyrosine kinase inhibitors2 for either country, but comparison with previous estimates in earlier time periods suggests that improvement in survival is ongoing. Greater concentration on treatment of CML in older patients may help improve survival for this population, for which survival lags.Table:Five year relative survival of patients diagnosed with CML in Germany and the US by age.GermanyUSGermany - USAgeNRSSENRSSEDiffP (Model)15-3932483.13.081881.92.0+1.20.940340-4932878.03.468082.82.2−4.80.177750-5942367.53.569777.72.6−10.20.001860-6436165.33.834070.34.1−5.00.206565-6941254.24.037554.54.2−0.30.5093Overalla184868.71.6291072.71.5−4.00.0036a=age adjusted using the percentage of cases in each age group in GermanyN=Number of casesRS=Relative SurvivalSE=Standard ErrorDiff=Difference between survival in Germany and the US Disclosures:No relevant conflicts of interest to declare.

Development and validation of a digital Gleason score biomarker signature for risk stratification of patients with prostate cancer.

40 Background: Gleason score (GS) is the most widely used grading system of cell differentiation in prostate cancer and one of the best pathological predictors of disease progression. Patients with high GS (&gt; 7) are the most likely to experience metastasis, whereas patients with low GS (&lt; 7) are expected to have favorable long-term outcomes. In addition to the subjective nature of GS assessment, patients with GS 7 represent a heterogeneous group in terms of patient outcomes. In this study a biomarker signature is developed and validated which could improve the prediction of high risk disease among GS 7 radical prostatectomy (RP) patients. Methods: Patient specimens from the Mayo Clinic RP Registry (n = 764) were used for feature selection and training of a 392-feature K-nearest neighbor (KNN, k = 11) classifier. These 392 genomic features were identified by assessing differential expression between patients with GS &lt; 7 and those with GS &gt; 7, using a Bonferroni adjusted T-Test with a p-value threshold of 0.05. The classifier was subsequently validated in two independent patient cohorts (Memorial Sloan Kettering [MSKCC] and German Cancer Research Center [DKFZ]) and compared to a state of the art biomarker signature (Penney et al. 2011). Results: In the MSKCC dataset (GSE21034) our model segregated GS &lt; 7 from GS &gt; 7 patients (n = 56) with an area under the receiver operating characteristic curve (AUC) of 0.97, comparable to the AUC of 0.94 obtained by Penney et al. in their independent validation set (n = 45). Strong performance was observed by our model when discriminating between primary Gleason grade (pGG) 3 and pGG 4 &amp; 5 patients in the MSKCC (n = 130) and DKFZ prostate cancer (GSE29079, n = 47) datasets, achieving AUCs of 0.75 and 0.87, respectively. In the challenging GS 7 subset, our model segregated GS 3 + 4 and 4 + 3 patients in the DKFZ dataset (n = 64) with an AUC of 0.81 outperforming the Penney et al. signature (AUC = 0.60). Conclusions: A biomarker signature was developed which discriminates between low and high GS patients, outperforming a previously reported signature. Further validation of this biomarker signature in additional post RP patients, as well as, pretreatment biopsy specimens is warranted.

Deriving New Treatment Strategies in Multiple Myeloma

Klaus Podar, MD PhD, is an Associate Professor of Experimental Oncology and Hematology and a hematologist/oncologist at the National Center for Tumor Diseases, University of Heidelberg, and the German Cancer Research Center (DKFZ) in Heidelberg, Germany. He has published more than 100 peer-reviewed manuscripts and keynote reviews, is a member of several scientific societies including the American Association of Cancer Research (AACR), American Society of Hematology (ASH) and European Hematology Association (EHA), and is the recipient of several prestigious awards.

Precision radiotherapy for brain tumors: A 10-year bibliometric analysis.

OBJECTIVE:Precision radiotherapy plays an important role in the management of brain tumors. This study aimed to identify global research trends in precision radiotherapy for brain tumors using a bibliometric analysis of the Web of Science.DATA RETRIEVAL:We performed a bibliometric analysis of data retrievals for precision radiotherapy for brain tumors containing the key words cerebral tumor, brain tumor, intensity-modulated radiotherapy, stereotactic body radiation therapy, stereotactic ablative radiotherapy, imaging-guided radiotherapy, dose-guided radiotherapy, stereotactic brachytherapy, and stereotactic radiotherapy using the Web of Science.SELECTION CRITERIA:Inclusion criteria: (a) peer-reviewed articles on precision radiotherapy for brain tumors which were published and indexed in the Web of Science; (b) type of articles: original research articles and reviews; (c) year of publication: 2002-2011. Exclusion criteria: (a) articles that required manual searching or telephone access; (b) Corrected papers or book chapters.MAIN OUTCOME MEASURES:(1) Annual publication output; (2) distribution according to country; (3) distribution according to institution; (4) top cited publications; (5) distribution according to journals; and (6) comparison of study results on precision radiotherapy for brain tumors.RESULTS:The stereotactic radiotherapy, intensity-modulated radiotherapy, and imaging-guided radiotherapy are three major methods of precision radiotherapy for brain tumors. There were 260 research articles addressing precision radiotherapy for brain tumors found within the Web of Science. The USA published the most papers on precision radiotherapy for brain tumors, followed by Germany and France. European Synchrotron Radiation Facility, German Cancer Research Center and Heidelberg University were the most prolific research institutes for publications on precision radiotherapy for brain tumors. Among the top 13 research institutes publishing in this field, seven are in the USA, three are in Germany, two are in France, and there is one institute in India. Research interests including urology and nephrology, clinical neurology, as well as rehabilitation are involved in precision radiotherapy for brain tumors studies.CONCLUSION:Precision radiotherapy for brain tumors remains a highly active area of research and development.

Exact solution of the 2-dimensional grid arrangement problem

Given an undirected graph G=(V,E), we consider injective mappings of its vertices to the k-dimensional Cartesian integer grid. Among such embeddings we are interested in those that minimize the sum of the resulting edge lengths, where the length of an edge is defined by the L1-metric. The case k=1 is the well-known Minimum Linear Arrangement Problem. We prove that the general problem is NP-hard for any fixed grid dimension. Our computational study focuses on the case k=2. We present as a first exact optimization algorithm a branch-and-cut scheme for sparse graphs. Several classes of valid inequalities are introduced and analyzed for facet defining properties of two corresponding polyhedra. Finally, computational results from a successful real-world application of the problem at the German Cancer Research Center (DKFZ) are presented.

Impact of polymorphic variation at 7p15.3, 3p22.1 and 2p23.3 loci on risk of multiple myeloma

Impact of polymorphic variation at 7p15.3, 3p22.1 and 2p23.3 loci on risk of multiple myeloma

A bias in genotyping the miR-27a rs895819 and rs11671784 variants

Due to the oncogenetic function of miR-27a, SNPs affecting this gene are frequently investigated for an influence on cancer risk. Yang et al. [1] has reported the association between the minor [G] allele of rs895819 on miR-27a and a reduced familial breast cancer risk in German population by direct sequencing. One study in Chinese Han population also observed a tumor suppressive effect of the rs895819 [G] allele in gastric cancer using MALDITOF MassARRAY [2]. One very recent publication has analysed the association of rs895819 with familial breast cancer risk in Italian population using TaqMan allelic discrimination assay [3]. However, this genotyping method appears to be inappropriate for rs895819 (chr19:13808292) due to the adjacent variant rs11671784 (chr19:13808296, which is located only 4 nucleotides distance and represents a rare variant, with the rare [T] allele frequency of 2.4 % and 1.9 % in German familial breast cancer cases and healthy controls, respectively [1]). Unfortunately, commercially available pre-designed assays of Applied Biosystems do not consider SNPs (especially SNPs with a low allele frequency) in their probe design nor provide access to the probe sequences [4]. Any TaqMan probe overlapping with the respective other SNP will give false calls in case of certain SNP constellations. To be sure about the primer and probe design, we ordered a self-designed TaqMan allelic discrimination probes for rs895819 and rs11671784 genotyping using the Applied Biosystems Assay-by-Design service. The two TaqMan probes comprise only the target SNP sites and avoid any overlap with the adjacent SNP (please see the primers and probes in Table 1). The observed genotyping bias introduced by the TaqMan allelic discrimination assay was as follows: First, we genotyped rs895819 in 1,217 German familial breast cancer patients and 1,422 unrelated healthy German controls as the samples described by Yang et al. [1] by both sequencing and TaqMan allelic discrimination method. All the uncertain or inconsistent genotypes determined by both assays were investigated for the second time for confirmation. As a result, the rs895819 genotyping using and TaqMan allelic discrimination probes were not concordant with our sequencing results (Table 2). The genotype constellation rs895819-AA/rs11671784-TT all failed when genotyping; the genotype constellation rs895819-AG/ rs11671784-CT resulted in false rs895819-GG instead of rs895819-AG calls (Table 2). Remarkably, researchers will not notify the bias via genotyping duplicates as the bias will be the same in both duplicates. As the genotype constellation frequencies of rs895819-AA/rs11671784-TT and rs895819-AG/rs11671784-CT are rather rare in Caucasian population (about 1.9 % in familial breast cancer cases and 1.4 % healthy controls [1]), it is also possible that the genotyping results are in Hardy–Weinberg equilibrium (HWE) although using biased genotyping assay. Nevertheless, this bias might have important implications on the final result. We further genotyped rs11671784 in a subset of 413 samples containing all the observed genotypes by TaqMan R. Yang (&) B. Burwinkel Helmholtz-University Group Molecular Epidemiology, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 581, 69120 Heidelberg, Germany e-mail: r.yang@dkfz.de

Epidemiological Follow-up 15 Years after the Breast Cancer Scandal in Essen.

Purpose: In the years 1993-1996 a number of presumably false-positive breast cancer diagnoses were made by a pathologist in Essen. A follow-up, undertaken 15 years later, investigated how many women had tumour recurrence and/or metastasis or had died from breast cancer. Material and Methods: A total of 151 (68 %) out of 222 women could be traced. One hundred and forty-seven (66.2 %) of the 222 women were alive. The observed survival rate, number of recurrences and/or metastases, and number of deaths from breast cancer were compared with data from the Munich Tumour Registry. The number of breast cancer cases among daughters of the affected women was ascertained. Results: The total observed survival rate at follow-up after 15 years was 93 %, a much higher figure than the survival rate of 45 % given by the Munich Tumour Registry. Recurrence and/or metastasis or death from breast cancer occurred in 9/222 cases (4.1 %). The incidence for these events calculated according to data from the Munich Tumour Registry is 13 %. Two daughters (2.2 %) out of a total of 90 were diagnosed with breast cancer whereas, according to the German Cancer Research Centre, the expected rate would have been between 5 and 10 %. Conclusions: The results of our follow-up after 15 years show that more women survived than expected and that the number of recurrences and/or metastases and deaths due to breast cancer was lower than expected. Fewer daughters of affected women were diagnosed with breast cancer than expected. These results support our suspicion that not all women diagnosed with breast cancer by a pathologist in Essen actually had breast cancer.

Mechanism of Evenness Interrupted (Evi)-Exosome Release at Synaptic Boutons

Wnt signaling plays critical roles during synaptic development and plasticity. However, the mechanisms by which Wnts are released and travel to target cells are unresolved. During synaptic development, the secretion of Drosophila Wnt1, Wingless, requires the function of Evenness Interrupted (Evi)/Wls, a Wingless-binding protein that is secreted along with Wingless at the neuromuscular junction. Given that Evi is a transmembrane protein, these studies suggested the presence of a novel vesicular mechanism of trans-synaptic communication, potentially in the form of exosomes. To establish the mechanisms for the release of Evi vesicles, we used a dsRNA assay in cultured cells to screen for genes that when down-regulated prevent the release of Evi vesicles. We identified two proteins, Rab11 and Syntaxin 1A (Syx1A), that were required for Evi vesicle release. To determine whether the same mechanisms were used in vivo at the neuromuscular junction, we altered the activity of Rab11 and Syx1A in motoneurons and determined the impact on Evi release. We found that Syx1A, Rab11, and its effector Myosin5 were required for proper Evi vesicle release. Furthermore, ultrastructural analysis of synaptic boutons demonstrated the presence of multivesicular bodies, organelles involved in the production and release of exosomes, and these multivesicular bodies contained Evi. We also used mass spectrometry, electron microscopy, and biochemical techniques to characterize the exosome fraction from cultured cells. Our studies revealed that secreted Evi vesicles show remarkable conservation with exosomes in other systems. In summary, our observations unravel some of the in vivo mechanisms required for Evi vesicle release.

Obituary

Obituary