Day 2 :
National Institute of Standards and Technology, USA
Time : 10:00 :11:00
Miral Dizdaroglu has obtained his PhD in physical chemistry at the Karlsruhe Technical University, Germany, and subsequently worked for seven years at the Max-Planck-Institute for Radiation Chemistry, Germany, before moving to USA. He has been at the National Institute of Standards and Technology (NIST) for more 30 years. In 2006, Dr. Dizdaroglu was conferred upon the rank of NIST Fellow. To date, he published highly cited 247 papers and gave numerous presentations on his work around the world. Dr. Dizdaroglu received numerous scientific awards including the Hillebrand Prize of the American Chemical Society, and the Silver and Gold Medal Awards of the US Department of Commerce. He was also awarded two Honorary Doctorates.
Statement of the Problem: Most chemotherapeutic agents kill cancer cells by damaging DNA. Cancer cells overexpress DNA repair proteins and thus increase DNA repair capacity that can cause resistance to therapy by removing DNA lesions before they become toxic. DNA repair proteins constitute targets for inhibitors to overcome the therapy resistance. Inhibition of DNA repair proteins is a promising approach to enhance the efficacy of cancer therapy. Despite the successes with other proteins, the development of inhibitors has been lagging for DNA glycosylases involved in the base excision repair mechanism. The purpose of this study was to discover small molecule inhibitors of the major human DNA glycosylases NEIL1, NTH1 and OGG1. First, we developed a fluorescence-based assay using double-stranded oligodeoxynucleotides containing one substrate lesion to detect both glycosylase and apyrimidinic/apurinic lyase activities of DNA glycosylases. From a screen of ∼400,000 compunds, a number of inhibitors were identified. Subsequently, we applied gas chromatography/isotope-dilution tandem mass spectrometry to measure the glycosylase activities of NEIL1, NTH1 and OGG1 using damaged DNA containing multiple lesions. Findings: Four purine analogs were found to be potent inhibitors of excision of the main substrates of NEIL1. Three of NEIL1 inhibitors also inhibited the excision of NTH1 substrates, but did not affect OGG1 activity. From a screen of ∼50,000 molecules, five hydrazydes were identified as potent inhibitors of OGG1 (Figure 1). These compounds exhibited no inhibition of NEIL1 and NTH1 activities for all their substrates analyzed by two different methodologies used in this work. Conclusion & Significance: Overall, this work forms the foundation for future drug discovery for the entire family of DNA glycosylases. The inhibitors identified serve as a proof-of-concept for the initial phase of the drug discovery process. Future work will involve the screening of additional compound libraries for different types of inhibitors of DNA glycosylases.
- Track 7: Chemical Biology
Track 8: DNA Damage and Repair
Track 9: RNA and DNA Nanotechnology
Track 10: Sequencing
Track 11: Molecular Therapeutics
Track 12: Molecular Biologics
Track 13: Computational Molecular Biology
National University of Singapore , Singapore
Yvonne began her research career in Bing Lim’s lab at the Genome Institute of Singapore, supported by an A*STAR Graduate Scholarship, and her PhD work led to key insights into the scope and mechanisms of microRNA activity (Miranda et al, Cell 2006; Tay et al, Nature 2008). Subsequently, she pursued her postdoctoral training in Pier Paolo Pandolfi’s lab at Harvard Medical School and Beth Israel Deaconess Medical Center under a Special Fellow award from the Leukemia & Lymphoma Society, and this research led to the discovery that protein-coding transcripts can co-regulate the tumor suppressor PTEN by competing for shared microRNAs (Tay et al, Cell 2011; Karreth et al, Cell 2011). Yvonne has recently returned to Singapore to begin her independent research career, and has been awarded the Singapore National Research Foundation Fellowship and NUS President’s Assistant Professorship to continue her research into this previously uncharacterized layer of gene regulation.
Pseudogenes, non-coding homologs of protein-coding genes, were once considered non-functional evolutional relics. Recent studies have shown that pseudogene transcripts can regulate their parental transcripts by sequestering shared microRNAs, thus acting as competing endogenous RNAs (ceRNAs). In this study, we utilize an unbiased screen to identify the ferritin heavy chain 1 (FTH1) transcript and multiple FTH1 pseudogenes as targets of several oncogenic miRNAs in prostate cancer. We characterize the critical role of this FTH1 gene:pseudogene:microRNA network in regulating tumorigenesis in prostate cancer, and show that impairing microRNA binding and subsequent ceRNA crosstalk results in complete phenotype rescue. Our results also demonstrate that pseudogenes are able to regulate intracellular iron levels, which are crucial for multiple physiological and pathophysiological processes. In summary, we describe a novel and extensive gene:pseudogene ceRNA network comprising multiple microRNAs and multiple pseudogenes derived from a single parental gene, which regulates iron storage and tumorigenesis in prostate cancer.
Jagiellonian University, Poland
Jolanta Jura (maiden name: Kwiatkowska by 2000) is professor at Faculty of Biochemistry, Biophysics and Biotechnology of Jagiellonian University in Krakow. After completing her Ph.D. in 2004 in Human Genetic Institute in Poznan, Jolanta Jura joined the group at Brigham and Women's Hospital of Harvard Medical School in Boston for two years, focusing on identification and characteristic of gene involved in tuberous sclerosis. Currently, the laboratory of Jolanta Jura studies the molecular mechanism of regulation of inflammation and inflammation related disorders focusing on gene expression and transcript stability, role of RNases in regulation of RNA profiles, signaling pathways.
The response of cell to external and internal stimuli results in the global changes in gene expression profile and accumulation of molecules that are essential for cell to adjust to a new conditions.One of recently discovered endonuclease capable of mRNA cleavage is Monocyte Chemoattractant Protein-induced protein 1 (MCPIP1), known also as Regnase-1 (Reg1). The protein is essential for degradation of short-living transcripts coding for inflammation-related proteins, including IL-1β, IL-6, IL-2, IL-8, IL-12b, IER-3, c-Rel and many other transcripts coding for proteins engaged in many other cellular processes such as: cell differentiation, metabolism and apoptosis. Besides mRNA, MCPIP1 regulates also miRNA processing. Ribonucleolytic activity of MCPIP11 has been attributed to a PIN (PilT N terminus) like domain, where four Asp residues (D141, D225, D226, D244) in the catalytic center determine RNase activity. It has been proven that MCPIP1 binding of an mRNA depends on a conserved stem-loop structures present in the 3’UTR of mRNA templates. Level of MCPIP1 changes during inflammatory processes generated by pathogen infections and also during sterile inflammation induced for example by cancer development. We found that the MCPIP1 levels are significantly lower in ccRCC samples than in surrounding tissues. Furthermore, lower level of MCPIP1 influences very important signaling pathways what corresponds to higher level of proangiogenic factors and enhanced metabolism and proliferation rate. Overall, these results suggest that MCPIP1 is an important player in ccRCC development.
Alexandria University, Egypt
Reham Abdel Haleem Aboelwafa, MD, a Lecturer of Clinical Pathology, Faculty of Medicine, Alexandria University, Egypt. She has expertise in research, teaching and administration both in hospital and education institution. She is expert in Molecular Techniques: PCR (conventional, and Real time PCR), microarray, Pyrosequencing and NGS in addition to FISH techniques including Prenatal Genetic Diagnosis (PGD) and Flow cytometric mmunophenotyping of different types of hematologic malignancies. She has several international publications in the field of molecular biology, genetics and epigenetics.
Overexpression of the Wilms tumor 1 gene (WT1) is implicated in the prognosis of acute myeloid leukemia (AML) with high expression predicting disease progression, as well as being intensively studied as a potential molecular marker for minimal residual disease (MRD) and treatment response. Many different isoforms for WT1 are generated by alternative transcription initiation, mRNA splicing and alternative translation initiation. Recently, an alternative promoter incorporating a unique first exon, alternative WT1 transcript (AWT1), has been described. The AWT1 expression and the underlying epigenetic alterations associated with its expression in AML are still unknown.
Saint Petersburg State University, Russia
Oleg Tikhodeyev is the author of the original approach for resolving multiple ambiguities and contradictions in current genetic concepts. He has shown that the key source of such ambiguities and contradictions is the erroneous belief that the same genetic term (for example, mutation) is able to comprise both specific phenomenology and the underlying mechanisms (Tikhodeyev, 2015). This belief became widely accepted after 1952, when the hereditary role of DNA had been demonstrated. In modern genetic concepts, the terms describing molecular mechanisms should be clearly distinguished from those describing phenomenology because there is no strict correlation between phenomenology and molecular mechanisms
Statement of the Problem: It has long been accepted that any hereditary factor in any organism was represented by DNA sequences. This idea became fundamental in molecular genetics, and was implicitly transformed into the DNA theory of inheritance. All basic genetic terms (genotype, gene, allele, mutation, recombination, etc.) were considered as specific DNA sequences or their alterations. However, multiple examples of stable epigenetic inheritance lacking any distinctions in DNA sequences were recently discovered, and the most exciting among them is protein inheritance. In particular, amyloid hereditary prions in fungi were considered as “protein-only” hereditary factors, which features were determined entirely by protein conformation. As a result, the principal question arises whether the DNA theory of inheritance is wrong or not. Methodology & Theoretical Orientation: Considering different variants of the same hereditary prion as prion alleles, we examined the molecular nature of such variety. Findings: To perpetuate stably in cell generations a certain prion allele requires two entities: (i) specific state of the prion protein, and (ii) the corresponding DNA sequence to provide reproduction of the prion particles. We name these entities as the DNA determinant and the epigenetic determinant, respectively. Thus, a certain prion allele is a bimodular hereditary system depending on both the DNA determinant and the epigenetic determinant. Alteration of any of these two determinants may result in establishment of a novel prion allele. Moreover, similar regularities are characteristic to all other cases of epigenetic inheritance, irrespective to the underlying mechanisms. Conclusion & Significance: The hereditary role of DNA is fundamental for any known mechanisms of inheritance, including epigenetic. However, it becomes an element of a more complicated concept: in addition to “DNA-only” hereditary factors, various bimodular hereditary factors also exist.
University of the Witwatersrand, South Africa
Patrick Arbuthnot is currently personal professor and director of the Wits/SAMRC Antiviral Gene Therapy Research Unit at the University of the Witwatersrand in South Africa. After graduating with a medical degree he completed a PhD in 1992 then carried out postdoctoral work at Necker Hospital in Paris, France. It was during this time that Dr Arbuthnot developed an interest in advancing gene therapy for treatment of liver cancer and hepatitis B virus (HBV) infection. On returning to South Africa, he established the Antiviral Gene Therapy Research Unit, which has now published widely on HBV infection, liver cancer, HIV-1 infection and developing new methods of treating these diseases. Patrick Arbuthnot’s main research interest is in advancing use of biological and synthetic nanoparticles to carry potentially therapeutic nucleic acids (DNA or RNA) that are capable of permanently disabling HBV.
Chronic infection with hepatitis B virus (HBV) remains and important global health problem. Carriers of the virus are at high risk for cirrhosis and liver cancer. Available treatment only has modest curative efficacy and improved therapy is a priority to prevent the life-threatening complications that accompany the infection. The viral replication intermediate comprising covalently closed circular DNA (cccDNA) exists as a stable minichromosome in infected hepatocytes. Licensed treatment has no effect on cccDNA and devising methods based on gene therapy to disable this replication intermediate has considerable potential. Previous work from our laboratory demonstrated effective inhibition of HBV replication and targeted disruption of cccDNA by Transcription Activator-Like Effector Nucleases (TALENs). Although this approach is promising, unintended mutagenesis may occur in chronic carriers as a result of TALEN activity at HBV sequences that are integrated into the host genome. To circumvent this problem, we have produced repressor TALEs (rTALEs) that were designed to induce transcriptional repression at essential HBV transcriptional regulatory elements: the basic core promoter/enhancer II and preS2 promoter sequences. KRAB-encoding sequences were fused to the N-terminal regions of TALEs contain sequence-specific DNA binding domains derived from the AvrBs4 N1 Xanthomas TALE. Each rTALE was expressed from the CMV promoter and engineered to interact with an HBV-specific 18 bp target. The repressors were incorporated into recombinant adeno-associated viral vectors which were used to deliver the antiviral
Shantou University, China
Imran Rashid has his expertise in cell culture research model development and working at Shantou University China. His expertise to develop animal cell culture model development bring him to the Shantou university to introduce new strategies to diagnose the stranding reasons and to find the possible reason to solve the problems. He is young dynamic and energetic rising scientist, his four years research experience in Zhejiang University China and later in Shantou University till now, proves that he is emerging scientist in the field of animal cell culture model development.
Statement of the Problem: Rapid declining population of cetacean is moving towards their extinctions, and protection department and researchers could not find the biological cause despite of physical injury and plastic materials in cetacean stomach. The sampling of cetaceans is restricted and field trails are likely to be impossible in the sea or oceans to diagnose cetaceans. Our presentation describes the state of art to know the unknow reason of cetacean death. Skin biopsy is only possible to some extent for sample collection, and we suggest that fibroblast cells culturing is possible to grow in the laboratory. These fibroblast cells can be genetically modified and converted into immortalized cell line. In the meantime, primary cells can also be converted into neuron and hepatocyte like cells by inducing some specific factors to conduct neurological and hepatological effects and response studies. These can be used to diagnose and estimate the molecular changes in the cetacean body. Several studies in the human has proven that fibroblast cells express about 100 genes related to neurodegenerative change and hepatic cells response related genes also found in fibroblast cells. This presentation demonstrates the new strategy to cetacean in the laboratory.