Day 2 :
University of Bradford, United Kingdom
Keynote: General Overview of the Comet Assay
Time : 09:35-10:00
Diana Anderson has completed her PhD at the University of Manchester, UK in the Faculty of Medicine. She is the Established Chair in Biomedical Sciences at the University of Bradford. She has published more than 450 papers, 8 books and successfully supervised 30 PhD students, she has Hirsch factor of 51. She is the Editor-in-Chief of a Book Series for the Royal Society of Chemistry and Consultant to many International Organizations such as the World Health Organisation/International Programme of Chemical Safety. She is/has been Member of the Editorial Board of ten international journals.
The comet assay is an important toxicological method for measuring DNA strand breaks in single cells. Cells embedded in agarose on a microscope slide are lysed with detergent and high salt. Electrophoresis at high pH results in structures resembling comets, observed by fluorescence microscopy. The comet tail is formed by DNA fragments moving towards the anode. The assay can be used for genotoxicity testing of novel compounds and exotoxins, human bio-monitoring, molecular epidemiology, basic research into DNA damage and repair and effects of nanoparticles. DNA repair can be measured after treatment of cells with a DNA damaging (challenging) agent and measuring damage remaining after different time intervals or with repair enzymes. This overview will be illustrated with examples from the work of the author’s group and collaborators. These will include studies for genotoxicity testing of various agents for exotoxins the work with DBP halogenated acetic acids, for human biomonitoring and molecular epidemiology the work with mother and babies, diabetes and lead-exposed children and for fundamental research that with drug resistant non-Hodgkin’s lymphoma patients over-expressing p53 mutant protein and lacking DNA repair. Most of this work has been carried out in somatic human lymphocytes but it can also be carried out in germ cells as human sperm. Using sperm, the oestrogens can cause DNA damage which can be diminished with anti-oxidants suggesting an oxygen radical involvement. Also as age increases in men so there is an increase in DNA damage. The comet assay can be modified and used as an innovative blood test to predict cancer. From a regulatory viewpoint, the assay is regarded as an indicator test but is incorporated into guidelines in some countries.
University of Cincinnati Academic Health Center, USA
Time : 09:30-10:00
Georg F. Weber attended medical school in Würzburg, Germany. He worked at the Dana-Farber Cancer Institute, Harvard Medical School from 1990 through 1999 and is currently on the faculty at the University of Cincinnati. Georg F. Weber has published about 90 scientific reports, including many in the most respected professional journals, and various monographs, including textbooks on molecular oncology and anti-cancer drugs. He holds several U.S. and international patents. As a component of his mission to combat cancer dissemination, Georg F. Weber is the founder and chief executive officer of MetaMol Theranostics, a company specialized in diagnosis and treatment of cancer metastasis.
Metastasis formation is an essential aspect of cancer. While the organ preference for dissemination is largely governed by tumor–host interactions on the epigenetic level, there is a genetic basis underlying the ability of cancer cells to disseminate. Metastasis genes are comprised of developmentally non-essential stress response genes, which encode homing receptors, their ligands, and extracellular matrix-degrading proteinases. They jointly cause invasion and anchorage-independence. Metastatic potential is conferred to cancer cells by aberrant expression or splicing of these genes, which include variant CD44 and osteopontin. The CD44-dependent spread of tumor cells is mediated by its cytokine ligand osteopontin, which induces directed migration. A C-terminal osteopontin domain ligates the variant exon 6 of CD44 through a protein-protein interaction and is likely to bind to variant exon 3 through a heparin bridge.rnOsteopontin is expressed at high levels by various cancers and contributes importantly to their invasive potential. In contrast, osteopontin derived from host cells induces cellular immunity and could bolster anti-tumor protection by cytotoxic T-lymphocytes. Underlying the functional differences between tumor-derived and host-derived osteopontin are structural characteristics. The osteopontin gene product is subject to alternative splicing, which yields three RNA messages, osteopontin-a (full length), osteopontin-b (lacking exon 5), and osteopontin-c (lacking exon 4). The shorter forms -b and -c are differentially expressed in cancers, but are absent from healthy tissues.rnThe major limiting factor in the process of metastasis formation is the death of the tumor cells before their implantation in target organs. Hence, anchorage-independent survival is essential for metastasis. While untransformed non-hematopoietic cells undergo anoikis consecutive to losing contact with their substratum, cancer cells can survive in the circulation for extended periods of time. The detachment of mammary epithelial cells prompts a loss of glucose transport and ATP deficiency, thus compromising the energy metabolism. Invasive breast tumor cells abundantly express two splice variants of the metastasis gene osteopontin. Osteopontin-a and osteopontin-c synergize in supporting tumor progression via up-regulating the energy production, which leads to deadherent survival. Osteopontin splice variants hold promise as potential drug targets.rn