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EJNMMI Res, 4(1), 17
2014

Visualization of angiogenesis during cancer development in the polyoma middle T breast cancer model: molecular imaging with (R)-[11C]PAQ.

Samén, Erik, Lu, Li, Mulder, Jan, Thorell, Jan-Olov, Damberg, Peter, Tegnebratt, Tetyana, Holmgren, Lars, Rundqvist, Helene, Stone-Elander, Sharon

Vascular endothelial growth factor receptor 2 (VEGFR2) is a crucial mediator of tumour angiogenesis. High expression levels of the receptor have been correlated to poor prognosis in cancer patients. Reliable imaging biomarkers for stratifying patients for anti-angiogenic therapy could therefore be valuable for increasing treatment success rates. The aim of this study was to investigate the pharmacokinetics and angiogenesis imaging abilities of the VEGFR2-targeting positron emission tomography (PET) tracer (R)-[11C]PAQ.(R)-[11C]PAQ was evaluated in the mouse mammary tumour virus-polyoma middle T (MMTV-PyMT) model of metastatic breast cancer. Mice at different stages of disease progression were imaged with (R)-[11C]PAQ PET, and results were compared to those obtained with [18 F]FDG PET and magnetic resonance imaging. (R)-[11C]PAQ uptake levels were also compared to ex vivo immunofluorescence analysis of tumour- and angiogenesis-specific biomarkers. Additional pharmacokinetic studies were performed in rat and mouse.A heterogeneous uptake of (R)-[11C]PAQ was observed in the tumorous mammary glands. Ex vivo analysis confirmed the co-localization of areas with high radioactivity uptake and areas with elevated levels of VEGFR2. In some animals, a high focal uptake was observed in the lungs. The lung uptake correlated to metastatic and angiogenic activity, but not to uptake of [18 F]FDG PET. The pharmacokinetic studies revealed a limited metabolism and excretion during the 1-h scan and a distribution of radioactivity mainly to the liver, kidneys and lungs. In rat, a high uptake was additionally observed in adrenal and parathyroid glands.The results indicate that (R)-[11C]PAQ is a promising imaging biomarker for visualization of angiogenesis, based on VEGFR2 expression, in primary tumours and during metastasis development.

Digital object identifier (DOI): 10.1186/2191-219X-4-17

J Dermatol Sci, 72(3), 304--310
December, 2013

A purified Feverfew extract protects from oxidative damage by inducing DNA repair in skin cells via a PI3-kinase-dependent Nrf2/ARE pathway.

Karien J. Rodriguez, Heng-Kuan Wong, Thierry Oddos, Michael Southall, Balz Frei, Simarna Kaur

Environmental factors such as solar ultraviolet (UV) radiation and other external aggressors provide an oxidative challenge that is detrimental to skin health. The levels of endogenous antioxidants decrease with age, thus resulting in less protection and a greater potential for skin damage. The NF-E2-related factor-2 (Nrf2) - antioxidant response element (ARE) pathway is a primary defense mechanism against oxidative stress, and induces the expression of antioxidant, detoxification and repair genes. Activation of ARE-Nrf2 can help restore oxidative homeostasis of the skin and play a role in inflammatory response and DNA repair mechanisms.To evaluate the role of a purified parthenolide-depleted Feverfew (PD-Feverfew) extract on the ARE-Nrf2 pathway and DNA repair in skin cells.These studies were undertaken in primary human keratinocytes or KB cells using Luciferase Promoter assay, siRNA transfection studies, Western blot analyses, Immunofluorescence microscopy, comet assay and quantitative real-time PCR.PD-Feverfew was found to induce Nrf2 nuclear translocation and to increase ARE activity in a dose dependent manner. Furthermore, knockdown of Nrf2 resulted in suppression of PD-Feverfew-induced ARE activity. PD-Feverfew was also found to induce phosphorylation of Akt, a kinase downstream of PI3K. Inhibition of PI3K via pre-treatment with the selective pharmacological inhibitor, LY294002, abolished PD-Feverfew-induced Nrf2/ARE activation. PD-Feverfew also reduced UV-induced DNA damage in a PI3K and Nrf2-dependent manner.Therefore, by increasing endogenous defense mechanisms and aid in DNA repair of damaged skin cells via activation of a PI3K-dependent Nrf2/ARE pathway, PD-Feverfew may help protect the skin from numerous environmental aggressors.

Digital object identifier (DOI): 10.1016/j.jdermsci.2013.08.004

Haematologica, 98(12), e166--e168
December, 2013

First description of the t(10;11)(q22;q23)/MLL-TET1 translocationin a T-cell lymphoblastic lymphoma, with subsequent lineage switchto acute myelomonocytic myeloid leukemia.

Antoine Ittel, Eric Jeandidier, Catherine Helias, Nathalie Perrusson, Catherine Humbrecht, Bruno Lioure, Isabelle Mazurier, Caroline Mayeur-Rousse, Amandine Lavaux, Sylvie Thiebault, Felix Lerintiu, Carine Gervais, Laurent Mauvieux

Reprod Biomed Online
December, 2013

Correlation between aneuploidy, apoptotic markers and DNA fragmentationin spermatozoa from normozoospermic patients.

Xavier Vendrell, Minerva Ferrer, Elena Garc{\'i}a-Mengual, Patricia Mu{\~n}oz, Juan Carlos Trivi{\~n}o, Carmen Calatayud, Vanesa Y. Rawe, Miguel Ruiz-Jorro

Genetic and biochemical sperm integrity is essential to ensure the reproductive competence. However, spermatogenesis involves physiological changes that could endanger sperm integrity. DNA protamination and apoptosis have been studied extensively. Furthermore, elevated rates of aneuploidy and DNA injury correlate with reproductive failures. Consequently, this study applied the conventional spermiogram method in combination with molecular tests to assess genetic integrity in ejaculate from normozoospermic patients with implantation failure by retrospectively analysing aneuploidy (chromosomes 18, X, Y), DNA fragmentation, externalization of phosphatidylserine and mitochondrial membrane potential status before and after magnetic activated cell sorting (MACS). Aneuploid, apoptotic and DNA-injured spermatozoa decreased significantly after MACS. A positive correlation was detected between reduction of aneuploidy and decreased DNA damage, but no correlation was determined with apoptotic markers. The interactions between apoptotic markers, DNA integrity and aneuploidy, and the effect of MACS on these parameters, remain unknown. In conclusion, use of MACS reduced aneuploidy, DNA fragmentation and apoptosis. A postulated mechanism relating aneuploidy and DNA injury is discussed; on the contrary, cell death markers could not be related. An 'apoptotic-like' route could explain this situation. Genetic and biochemical sperm integrity is essential to ensure reproductive success and support the earliest phases of embryo development. Paradoxically, spermatogenesis involves physiological changes that could endanger the DNA and cell integrity. Sperm-specific mechanisms have been studied extensively, and DNA packaging and programmed cell death (apoptosis) are potentially harmful. Also, elevated rates of chromosomal numerical abnormalities and breakage of sperm DNA have been correlated with reproductive failures. In this context, basic sperm examination methods have been combined with molecular tests to assess genetic integrity. On the other hand, magnetic activated cell sorting (MACS) can reduce the number of programmed-to-death spermatozoa. This system retains damaged spermatozoa, thereby improving the sample's quality. The relationships between apoptosis, DNA integrity and chromosomal abnormalities (aneuploidy) as a whole, and the effect of MACS on these parameters remain unknown. We analysed aneuploidy, DNA damage, and biochemical markers of cell death in ejaculate from normozoospermic patients with implantation failures before and after MACS. Aneuploid, apoptotic and DNA-injured spermatozoa decreased significantly after MACS. A positive correlation was detected between the reduction of aneuploidy and DNA damage; on the contrary, no correlation was determined with apoptotic markers. In conclusion, the use of MACS reduced aneuploidy, DNA breakages and apoptosis. A hypothesized mechanism relating aneuploidy and DNA injury is discussed; on the contrary, death cell markers could not be directly related. An 'apoptotic-like' route could explain this situation.

Neoplasia, 15(11), 1301--1313
November, 2013

Alternative Lengthening of Telomeres: Recurrent Cytogenetic Aberrations and Chromosome Stability under Extreme Telomere Dysfunction.

Despoina Sakellariou, Maria Chiourea, Christina Raftopoulou, Sarantis Gagos

Human tumors using the alternative lengthening of telomeres (ALT) exert high rates of telomere dysfunction. Numerical chromosomal aberrations are very frequent, and structural rearrangements are widely scattered among the genome. This challenging context allows the study of telomere dysfunction-driven chromosomal instability in neoplasia (CIN) in a massive scale. We used molecular cytogenetics to achieve detailed karyotyping in 10 human ALT neoplastic cell lines. We identified 518 clonal recombinant chromosomes affected by 649 structural rearrangements. While all human chromosomes were involved in random or clonal, terminal, or pericentromeric rearrangements and were capable to undergo telomere healing at broken ends, a differential recombinatorial propensity of specific genomic regions was noted. We show that ALT cells undergo epigenetic modifications rendering polycentric chromosomes functionally monocentric, and because of increased terminal recombinogenicity, they generate clonal recombinant chromosomes with interstitial telomeric repeats. Losses of chromosomes 13, X, and 22, gains of 2, 3, 5, and 20, and translocation/deletion events involving several common chromosomal fragile sites (CFSs) were recurrent. Long-term reconstitution of telomerase activity in ALT cells reduced significantly the rates of random ongoing telomeric and pericentromeric CIN. However, the contribution of CFS in overall CIN remained unaffected, suggesting that in ALT cells whole-genome replication stress is not suppressed by telomerase activation. Our results provide novel insights into ALT-driven CIN, unveiling in parallel specific genomic sites that may harbor genes critical for ALT cancerous cell growth.

Br J Haematol
October, 2013

Fusion of the additional sex combs like 1 and teashirt zinc fingerhomeobox 2 genes resulting from ider(20q) aberration in a patientwith myelodysplastic syndrome.

Jana Brezinova, Iveta Sarova, Halka Buryova, Jana Markova, Sarka Ransdorfova, Silvia Izakova, Karla Kostylkova, Jacqueline Soukupova, Zuzana Zemanova, Kyra Michalova

A variant of del(20q), an isochromosome of the long arm with the loss of an interstitial part of 20q, ider(20q), has been reported in patients with myeloid diseases (Li et al, 2004). About 40 cases with this rearrangement have been reported up to 2012 (reviewed by Mullier et al, 2012). Molecular cytogenetic and array techniques have been used for mapping of the deleted region on 20q (Douet-Guilbert et al, 2009). The proximal breakpoints are consistently located in the 20q11.21 band, and the distal breakpoints span from band 20q13.13 to band 20q13.33.

Stem Cell Res, 12(1), 1--10
September, 2013

uPAR-controlled oncolytic adenoviruses eliminate cancer stem cellsin human pancreatic tumors.

Luciano Sobrevals, Ana Mato-Berciano, Nerea Urtasun, Adela Mazo, Cristina Fillat

Pancreatic tumors contain cancer stem cells highly resistant to chemotherapy. The identification of therapies that can eliminate this population of cells might provide with more effective treatments. In the current work we evaluated the potential of oncolytic adenoviruses to act against pancreatic cancer stem cells (PCSC). PCSC from two patient-derived xenograft models were isolated from orthotopic pancreatic tumors treated with saline, or with the chemotherapeutic agent gemcitabine. An enrichment in the number of PCSC expressing the cell surface marker CD133 and a marked enhancement on tumorsphere formation was observed in gemcitabine treated tumors. No significant increase in the CD44, CD24, and epithelial-specific antigen (ESA) positive cells was observed. Neoplastic sphere-forming cells were susceptible to adenoviral infection and exposure to oncolytic adenoviruses resulted in elevated cytotoxicity with both Adwt and the tumor specific AduPARE1A adenovirus. In vivo, intravenous administration of a single dose of AduPARE1A in human-derived pancreatic xenografts led to a remarkable anti-tumor effect. In contrast to gemcitabine AduPARE1A treatment did not result in PCSC enrichment. No enrichment on tumorspheres neither on the CD133(+) population was detected. Therefore our data provide evidences of the relevance of uPAR-controlled oncolytic adenoviruses for the elimination of pancreatic cancer stem cells.

Mutat Res
June, 2013

Persisting ring chromosomes detected by mFISH in lymphocytes of acancer patient-A case report.

Sabine Schmitz, Michael Pinkawa, Michael J. Eble, Ralf Kriehuber

We report the case of an 84 years old prostate cancer patient with severe side effects after radiotherapy in 2006. He was cytogenetically analysed in 2009 and in 2012 in a comparative study for individual radiosensitivity of prostate cancer patients. No other patient had clonal aberrations, but this patient showed ring chromosomes in the range of 21-25\% of lymphocytes. He received 5 cycles of 5-fluorouracil/folic acid for chemotherapy of sigmoid colon carcinoma in 2003, three years before radiotherapy of prostate cancer. Blood samples were irradiated ex vivo with Cs-137 {\~a}-rays (0.7Gy/min) in the G0-phase of the cell cycle. 100 FISH painted metaphases were analysed for the control and the irradiated samples each. Multicolour in situ hybridisation techniques like mFISH and mBand as well as MYC locus, telomere and centromere painting probes were used to characterise ring metaphases. Metaphase search and autocapture was performed with a Zeiss Axioplan 2 imaging microscope followed by scoring and image analysis using Metafer 4/ISIS software (MetaSystems). In 2009 chromosome 8 rings were found in about 25\% of lymphocytes. Rings were stable over time and increased to about 30\% until 2012. The ring chromosome 8 always lacked telomere signals and a small amount of rings displayed up to four centromere signals. In aberrant metaphases 8pter and 8qter were either translocated or deleted. Further analyses revealed that the breakpoint at the p arm is localised at 8p21.2-22. The breakpoint at the q arm turned out to be distal from the MYC locus at 8q23-24. We hypothesise that the ring chromosome 8 has been developed during the 5 FU/folic acid treatments in 2003. The long term persistence might be due to clonal expansion of a damaged but viable hematopoietic stem cell giving rise to cycling progenitor cells that permit cell survival and proliferation.

Radiother Oncol, 107(3), 377--381
June, 2013

Early biomarkers related to secondary primary cancer risk in radiotherapytreated prostate cancer patients: IMRT versus IMAT.

Joke Werbrouck, Piet Ost, Valerie Fonteyne, Gert {De Meerleer}, Wilfried {De Neve}, Evelien Bogaert, Laurence Beels, Klaus Bacher, Anne Vral, Hubert Thierens

To investigate whether rotational techniques (Volumetric Modulated Arc Therapy - VMAT) are associated with a higher risk for secondary primary malignancies compared to step-and-shoot Intensity Modulated Radiation Therapy (ss-IMRT). To this end, radiation therapy (RT) induced DNA double-strand-breaks and the resulting chromosomal damage were assessed in peripheral blood T-lymphocytes of prostate cancer (PCa) patients applying {\~a}H2AX foci and G0 micronucleus (MN) assays.The study comprised 33PCa patients. A blood sample was taken before start of therapy and after the 1st and 3rd RT fraction to determine respectively the RT-induced {\~a}H2AX foci and MN. The equivalent total body dose (D(ETB)) was calculated based on treatment planning data.A linear dose response was obtained for {\~a}H2AX foci yields versus D(ETB) while MN showed a linear-quadratic dose response. Patients treated with large volume (LV) VMAT show a significantly higher level of induced {\~a}H2AX foci and MN compared to IMRT and small volume (SV) VMAT (p

Asian J Androl, 15(3), 421--424
May, 2013

No difference in high-magnification morphology and hyaluronic acidbinding in the selection of euploid spermatozoa with intact DNA.

Suchada Mongkolchaipak, Teraporn Vutyavanich

In this study, we compared conventional sperm selection with high-magnification morphology based on the motile sperm organellar morphology examination (MSOME) criteria, and hyaluronic acid (HA) binding for sperm chromosome aneuploidy and DNA fragmentation rates. Semen from 50 severe male factor cases was processed through density gradient centrifugation, and subjected to sperm selection by using the conventional method (control), high magnification at ?6650 or HA binding. Aneuploidy was detected by fluorescence in situ hybridization with probes for chromosomes 13, 18, 21, X and Y, and DNA fragmentation by the terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) method. Spermatozoa selected under high-magnification had a lower DNA fragmentation rate (2.6\% vs. 1.7\%; P=0.032), with no significant difference in aneuploidy rate (0.8\% vs 0.7\%; P=0.583), than those selected by the HA binding method. Spermatozoa selected by both methods had much lower aneuploidy and DNA fragmentation rate than the controls (7\% aneuploidy and 26.8\% DNA fragmentation rates, respectively). In the high-magnification group, the aneuploidy rate was lower when the best spermatozoa were selected than when only the second-best spermatozoa were available for selection, but the DNA fragmentation rate was not different. In conclusion, sperm selection under high magnification was more effective than under HA binding in selecting spermatozoa with low DNA fragmentation rate, but the small difference (0.9\%) might not be clinically meaningful. Both methods were better than the conventional method of sperm selection.

Basic and Clinical Andrology, 23(13), 1-8
2013

FISH and tips: a large scale analysis of automated versus manual scoring for sperm aneuploidy detection

Guillaume Martinez, Pierre Gillois, Marine Le Mitouard, R{\'e}my Borye, Camille Esquerr{\'e}-Lamare, V{\'e}ronique Satre, Louis Bujan, Sylviane Hennebicq

Background Approximately 1% of the spermatozoa found in ejaculate of healthy men are aneuploid and this rate increases in the population of subfertile and infertile men. Moreover, fertilization with these aneuploid sperm can lead to impaired embryo development. Fluorescent In Situ Hybridization (FISH) is the common cytogenetic tool used for aneuploidy screening on sperm. However, it is a time-consuming technique and cytogenetic or in vitro fertilization laboratories cannot routinely use it and face the increasing demand of such analyses before Assisted Reproductive Techniques (ART). As automation can be a clue for routine practice, this study compares manual and automated scoring of sperm aneuploidy rates using a Metafer Metasystems® device. The results obtained also contribute to global data about FISH on sperm cells. Methods We recruited 100 men addressed for sperm cryopreservation. They all signed an informed consent to participate in the study. 29 men were donors or consulted before vasectomy (control group) and 71 were suffering of Hodgkin’s disease or non Hodgkin lymphoma (patient group). One semen sample was collected for each patient, analyzed according to WHO criteria and prepared for a triple-color FISH using centromeric probes for chromosomes 18, X and Y. Automated scoring was performed using a Metafer Metasystems® device. Results 507,019 cells were scored. We found a strong concordance between the automated and the manual reading (d 

PLoS ONE
2013

Reduced Placental Telomere Length during Pregnancies Complicated by Intrauterine Growth Restriction

J{\'e}r{\^o}me Toutain, Martina Prochazkova-Carlotti, David Cappellen, Ana Jarne, Edith Chevret, Jacky Ferrer, Yamina Idrissi, Fanny Pelluard, Dominique Carles, Brigitte Maugey-Laulon, Didier Lacombe, Jacques Horovitz, Jean-Philippe Merlio, Robert Saura

Recent studies have shown that telomere length was significantly reduced in placentas collected at delivery from pregnancies complicated by intrauterine growth restriction secondary to placental insufficiency. Placental telomere length measurement during ongoing pregnancies complicated by intrauterine growth restriction has never been reported. This was the main objective of our study.

J Hered
October, 2012

Development and Application of Camelid Molecular Cytogenetic Tools.

Felipe Avila, Pranab J. Das, Michelle Kutzler, Elaine Owens, Polina Perelman, Jiri Rubes, Miroslav Hornak, Warren E. Johnson, Terje Raudsepp

Cytogenetic chromosome maps offer molecular tools for genome analysis and clinical cytogenetics and are of particular importance for species with difficult karyotypes, such as camelids (2n = 74). Building on the available human-camel zoo-fluorescence in situ hybridization (FISH) data, we developed the first cytogenetic map for the alpaca (Lama pacos, LPA) genome by isolating and identifying 151 alpaca bacterial artificial chromosome (BAC) clones corresponding to 44 specific genes. The genes were mapped by FISH to 31 alpaca autosomes and the sex chromosomes; 11 chromosomes had 2 markers, which were ordered by dual-color FISH. The STS gene mapped to Xpter/Ypter, demarcating the pseudoautosomal region, whereas no markers were assigned to chromosomes 14, 21, 22, 28, and 36. The chromosome-specific markers were applied in clinical cytogenetics to identify LPA20, the major histocompatibility complex (MHC)-carrying chromosome, as a part of an autosomal translocation in a sterile male llama (Lama glama, LGL; 2n = 73,XY). FISH with LPAX BACs and LPA36 paints, as well as comparative genomic hybridization, were also used to investigate the origin of the minute chromosome, an abnormally small LPA36 in infertile female alpacas. This collection of cytogenetically mapped markers represents a new tool for camelid clinical cytogenetics and has applications for the improvement of the alpaca genome map and sequence assembly.

Digital object identifier (DOI): 10.1093/jhered/ess067

Prenat Diagn, 32(8), 742--751
August, 2012

Identification of circulating fetal cell markers by microarray analysis.

Marie Brinch, Lotte Hatt, Ripudaman Singh, Kristine M{\o}ller, Steffen Sommer, Niels Uldbjerg, Britta Christensen, Steen K{\o}lvraa

Different fetal cell types have been found in the maternal blood during pregnancy in the past, but fetal cells are scarce, and the proportions of the different cell types are unclear. The objective of the present study was to identify specific fetal cell markers from fetal cells found in the maternal blood circulation at the end of the first trimester.Twenty-three fetal cells were isolated from maternal blood by removing the red blood cells by lysis or combining this with removal of large proportions of maternal white blood cells by magnetic-activated cell sorting. Fetal cells identified by XY fluorescence in situ hybridization and confirmed by reverse-color fluorescence in situ hybridization were shot off microscope slides by laser capture microdissection. The expression pattern of a subset of expressed genes was compared between fetal cells and maternal blood cells using stem cell microarray analysis.Twenty-eight genes were identified as fetal cell marker candidates.Of the 28 fetal marker candidate genes, five coded for proteins, which are located on the outer surface of the cell membrane and not expressed in blood. The protein product of these five genes, MMP14, MCAM, KCNQ4, CLDN6, and F3, may be used as markers for fetal cell enrichment.

Am J Med Genet B Neuropsychiatr Genet, 159B(5), 598--604
July, 2012

Mild cognitive impairment identified in older individuals with Downsyndrome by reduced telomere signal numbers and shorter telomeresmeasured in microns.

Edmund C. Jenkins, Lingling Ye, Milen Velinov, Sharon J. Krinsky-McHale, Warren B. Zigman, Nicole Schupf, Wayne P. Silverman

Previously, we established that short-term T lymphocyte cultures from people with Down syndrome (DS) and dementia (Alzheimer's disease) had shorter telomeres than did those from age- and sex-matched people with DS only, quantified as significantly reduced numbers of signals of peptide nucleic acid (PNA) telomere probes in whole metaphases [Jenkins et al. (2008); Neurosci Lett 440:340-343] as well as reduced telomere probe light intensity values in interphases [Jenkins et al. (2010); Neurobiol Aging 31:765-771]. We now describe shorter telomere length in adults with DS and mild cognitive impairment (MCI) compared to age- and sex-matched individuals with DS without MCI. Telomere length is quantified by reduced telomere signal numbers and shorter chromosome 1 telomeres measured in micrometers (microns). These findings were in agreement with quantitative light intensity measurements of chromosome 1 and chromosome 21 PNA telomere probes with and without the use of a #normalizing##ratio# involving the fluorescence exhibited by a PNA probe for centromere 2, and with the use of light intensity measurements of interphase preparations. Most importantly, the distributions of chromosome 1 telomere lengths (in microns) were completely non-overlapping for adults with and without MCI, indicating that this measure has great promise as a biomarker for MCI as well as dementia in this population.

Leukemia, 26(7), 1695--1697
July, 2012

Molecular characterization of deletions of the long arm of chromosome5 (del(5q)) in 94 MDS/AML patients.

N. Douet-Guilbert, E. {De Braekeleer}, A. Basinko, A. Herry, N. Gueganic, C. Bovo, K. Trillet, A. {Dos Santos}, M. J. {Le Bris}, F. Morel, J. R. Eveillard, C. Berthou, M. {De Braekeleer}

Deletion of the long arm of chromosome 5 (del(5q)) is a common finding in myelodysplastic syndrome (MDS) and in acute myeloid leukemia (AML). First described in 1974 by Van den Berghe et al.,1 the 5q- syndrome, more frequently found in old-aged females, is characterized by erythroid hypoplasia, macrocytic anemia, normal to elevated platelets count, preponderance of monolobulated megakaryocytes, isolated 5q deletion and low rate of progression to AML.

J Clin Invest, 122(2), 569--574
February, 2012

Recurrent genomic instability of chromosome 1q in neural derivativesof human embryonic stem cells.

Christine Varela, J{\'e}r{\^o}me Alexandre Denis, J{\'e}r{\^o}me Polentes, Maxime Feyeux, Sophie Aubert, Benoite Champon, Genevi{\`e}ve Pi{\'e}tu, Marc Peschanski, Nathalie Lefort

Human pluripotent stem cells offer a limitless source of cells for regenerative medicine. Neural derivatives of human embryonic stem cells (hESCs) are currently being used for cell therapy in 3 clinical trials. However, hESCs are prone to genomic instability, which could limit their clinical utility. Here, we report that neural differentiation of hESCs systematically produced a neural stem cell population that could be propagated for more than 50 passages without entering senescence; this was true for all 6 hESC lines tested. The apparent spontaneous loss of evolution toward normal senescence of somatic cells was associated with a jumping translocation of chromosome 1q. This chromosomal defect has previously been associated with hematologic malignancies and pediatric brain tumors with poor clinical outcome. Neural stem cells carrying the 1q defect implanted into the brains of rats failed to integrate and expand, whereas normal cells engrafted. Our results call for additional quality controls to be implemented to ensure genomic integrity not only of undifferentiated pluripotent stem cells, but also of hESC derivatives that form cell therapy end products, particularly neural lines.

PLoS One, 7(10), e47185
2012

Evaluation of different biomarkers to predict individual radiosensitivityin an inter-laboratory comparison--lessons for future studies.

Burkhard Greve, Tobias B{\"o}lling, Susanne Amler, Ute R{\"o}ssler, Maria Gomolka, Claudia Mayer, Odilia Popanda, Kristin Dreffke, Astrid Rickinger, Eberhard Fritz, Friederike Eckardt-Schupp, Christina Sauerland, Herbert Braselmann, Wiebke Sauter, Thomas Illig, Dorothea Riesenbeck, Stefan K{\"o}nemann, Normann Willich, Simone M{\"o}rtl, Hans Theodor Eich, Peter Schmezer

Radiotherapy is a powerful cure for several types of solid tumours, but its application is often limited because of severe side effects in individual patients. With the aim to find biomarkers capable of predicting normal tissue side reactions we analysed the radiation responses of cells from individual head and neck tumour and breast cancer patients of different clinical radiosensitivity in a multicentric study. Multiple parameters of cellular radiosensitivity were analysed in coded samples of peripheral blood lymphocytes (PBLs) and derived lymphoblastoid cell lines (LCLs) from 15 clinical radio-hypersensitive tumour patients and compared to age- and sex-matched non-radiosensitive patient controls and 15 lymphoblastoid cell lines from age- and sex- matched healthy controls of the KORA study. Experimental parameters included ionizing radiation (IR)-induced cell death (AnnexinV), induction and repair of DNA strand breaks (Comet assay), induction of yH2AX foci (as a result of DNA double strand breaks), and whole genome expression analyses. Considerable inter-individual differences in IR-induced DNA strand breaks and their repair and/or cell death could be detected in primary and immortalised cells with the applied assays. The group of clinically radiosensitive patients was not unequivocally distinguishable from normal responding patients nor were individual overreacting patients in the test system unambiguously identified by two different laboratories. Thus, the in vitro test systems investigated here seem not to be appropriate for a general prediction of clinical reactions during or after radiotherapy due to the experimental variability compared to the small effect of radiation sensitivity. Genome-wide expression analysis however revealed a set of 67 marker genes which were differentially induced 6 h after in vitro-irradiation in lymphocytes from radio-hypersensitive and non-radiosensitive patients. These results warrant future validation in larger cohorts in order to determine parameters potentially predictive for clinical radiosensitivity.

Blood, 118(26), 6760--6768
December, 2011

Impact of additional cytogenetic aberrations at diagnosis on prognosisof CML: long-term observation of 1151 patients from the randomizedCML Study IV.

Alice Fabarius, Armin Leitner, Andreas Hochhaus, Martin C M{\"u}ller, Benjamin Hanfstein, Claudia Haferlach, Gudrun G{\"o}hring, Brigitte Schlegelberger, Martine Jotterand, Andreas Reiter, Susanne Jung-Munkwitz, Ulrike Proetel, Juliana Schwaab, Wolf-Karsten Hofmann, J{\"o}rg Schubert, Hermann Einsele, Anthony D Ho, Christiane Falge, Lothar Kanz, Andreas Neubauer, Michael Kneba, Frank Stegelmann, Michael Pfreundschuh, Cornelius F Waller, Karsten Spiekermann, Gabriela M Baerlocher, Michael Lauseker, Markus Pfirrmann, Joerg Hasford, Susanne Saussele, R{\"u}diger Hehlmann, f{\"u}r Klinische Krebsforschung (SAKK), Schweizerische Arbeitsg, the German CML Study Group,

The prognostic relevance of additional cytogenetic findings at diagnosis of chronic myeloid leukemia (CML) is unclear. The impact of additional cytogenetic findings at diagnosis on time to complete cytogenetic (CCR) and major molecular remission (MMR) and progression-free (PFS) and overall survival (OS) was analyzed using data from 1151 Philadelphia chromosome-positive (Ph(+)) CML patients randomized to the German CML Study IV. At diagnosis, 1003 of 1151 patients (87\%) had standard t(9;22)(q34;q11) only, 69 patients (6.0\%) had variant t(v;22), and 79 (6.9\%) additional cytogenetic aberrations (ACAs). Of these, 38 patients (3.3\%) lacked the Y chromosome (-Y) and 41 patients (3.6\%) had ACAs except -Y; 16 of these (1.4\%) were major route (second Philadelphia [Ph] chromosome, trisomy 8, isochromosome 17q, or trisomy 19) and 25 minor route (all other) ACAs. After a median observation time of 5.3 years for patients with t(9;22), t(v;22), -Y, minor- and major-route ACAs, the 5-year PFS was 90\%, 81\%, 88\%, 96\%, and 50\%, and the 5-year OS was 92\%, 87\%, 91\%, 96\%, and 53\%, respectively. In patients with major-route ACAs, the times to CCR and MMR were longer and PFS and OS were shorter (P