Day 2 :
1Institute of Molecular Genetics, Italy
Keynote: Tumorspheres from in vitro transformed cell lines show molecular signatures related to stemness and transformation
Time : 09:30-10:10
Chiara Mondello is Senior Researcher and Group Leader at the Institute of Molecular Genetics of the National Research Council in Pavia (Italy). Her main interests concern the study of genome instability and cellular transformation in mammalian cells. She has published more than 80 papers in peer-reviewed journals and several chapters in books. She has been the Editor of the book “Multiple Pathways in Cancer Development” (Transwell Research Network).
Evidence indicates that a subset of cells endowed with high tumorigenic potential and stemness features (cancer stem cells: CSCs) is responsible for tumor initiation and maintenance in several cancers. In this study, we used a tumor cellular model developed in our laboratory from telomerase immortalized human fibroblasts (named cen3tel) and the tumorsphere assay to possibly isolate and characterize CSCs from in vitro transformed cells. We found that cen3tel cells were able to form spheres (frequency ~ 2-10%) and sphere cells showed self renewal capacity and Sox2 overexpression, suggesting that in these populations there is a subset of cells with CSC-like features. The characterization of sphere cells revealed that they displayed the activation of pro-survival mechanisms, through the expression modulation of genes as c-MYC, GNL3 and Notch, and the tumor suppressor miR-34a, which could favor the growth of cells in suspension upon detachment from a solid support. Moreover, genome wide gene expression profiles of sphere cells relatively to adherent cells revealed an extensive transcriptional reprogramming involving several stemness and cancer-related genes, indicating that transformed cells are highly plastic entities adopting specific gene expression programs depending on different environmental conditions. However, sphere cells were only slightly more tumorigenic in vivo than adherent cells suggesting that different subpopulations can support tumorigenicity in transformed cells, highlighting a further level of complexity in tumor heterogeneity.
Gillies McIndoe Research Institute, New Zealand
Time : 10:10-10:50
Dr Tinte Itinteang serves as the current Chief Scientific Officer and the Evans Family Research Fellow of the Gillies McIndoe Research Institute (GMRI) in Wellington, New Zealand. Tinte completed medical training at the Melbourne University in 2001, and then completed his basic medical residency in New Zealand, from 2008- 2010 he completed his PhD from Victoria University of Wellington, NZ on the role of stem cells and the renin-angiotensin system (RAS) in infantile haemangioma. From 2012-2014, he was appointed as a research fellow at the Gillies McIndoe Research institute, during which time he spent six weeks at the Friedlander laboratory at The Scripps Research Institute in San Diego investigating the role of iPSCs for disease modelling. He was then appointed as the Chief Scientist of the GMRI from 2015. His work on the role of stem cells and the RAS in infantile haemangioma has been acknowledged with the International Society for the Study of Vascular Anomalies John Mulliken award as well as several national and international honours. He is the author of over 50 peer reviewed articles and over 100 presentations at international conferences.
Infantile haemangioma (IH) is the most common tumour of infancy, characterised by an initial proliferation with aggressive vasculogenesis, followed by spontaneous slow involution leaving a fibro-fatty residuum. IH affects about 10% of infants and has a predilection for female, Caucasian and premature infants. We have demonstrated the presence of embryonic-like stem cells in the endothelium of proliferating IH that express markers associated with mesenchymal and haematopoietic plasticity. These primitive cells are also the putative source of the fibro-fatty tissue that naturally occurs during involution of this tumour. This presentation focuses on the role of the primitive endothelium in the pathoetiogensis of IH and their ability to form downstream definitive mesenchymal and haematopoietic cells. It will also cover our insights into the role of the renin-angiotensin system (RAS) in the regulation of this primitive population, underscoring the novel use of RAS modulators in the treatment of problematic IH. The potential to exploit IH as a human model for directed regenerative medicine will be discussed.
Minerva Biotechnologies, USA
Keynote: Are cancer stem cells really healthy cells being reprogrammed back to a naïve stem cell state?
Time : 11:10-11:50
Cynthia Bamdad holds a BS in Physics from Northeastern University and a PhD in Biophysics from Harvard University. As a PhD candidate and without the benefit of an advisor, she invented the first electronic DNA chip that she commercialized at a California startup, which was then sold to Motorola for $300M; the chip is now the core of an FDA-approved diagnostic device marketed by Genmark. She is sole or lead inventor on over a hundred patent applications, including for novel technologies that enabled Minerva Biotechnologies’ groundbreaking discoveries in basic cancer biology.
The intersecting space between stem cells and cancer stem cells is rapidly expanding. Many of the markers, previously thought to be stem cell markers, are now being shown to be markers of metastasis as well. Similarly, metastatic markers such as CXCR4, have now been shown to be markers of the earliest stem cells, called ‘naïve’ stem cells. Here, we report the discovery of a novel stem cell growth factor, NME7-AB, that induces human stem cells to revert to the earliest naïve state. NME7-AB is naturally expressed in every cell of a human Day 3 morula, but by Day 5 is only expressed in the cells of the inner cell mass, which are naïve by definition. Although this new growth factor should be turned off for adult life, we have found it expressed in nearly every metastatic tissue we have examined. In vitro and in vivo, NME7-AB induces cancer cells to become more metastatic; they upregulate metastatic markers and form tumors in mice from as few as 50 cells. Daily injections of NME7-AB into the tumor-bearing mice caused the solid tumor cancers to metastasize. These data are consistent with the idea that cancer cells are being reprogrammed back to a stem cell-like state, wherein the metastatic cancer cells, aka cancer stem cells, are reverted all the way back to the naïve stem cell state. These data suggest that agents that inhibit naïve stem cell pluripotency or growth will also be inhibitors of cancer cells and, more particularly, cancer stem cells.
Tolero Pharmaceuticals, Inc., USA
Keynote: Defining and exploiting the clinical mechanism of activity for alvocidib in acute myeloid leukemia patients
Time : 11:50-12:30
Steven L Warner specializes in small molecule drug discovery, new screening platforms in drug discovery, and translational research focusing on cancer therapeutics. He is an expert in the discovery of novel cancer agents and has played integral roles in moving multiple compounds into clinical trials. He earned his graduate degree in Pharmaceutical Sciences at the University of Arizona. He completed a postdoctoral fellowship under the mentoring of Dan Von Hoff at the Translational Genomics Research Institute (TGen). He is currently the Vice President of Drug Discovery and Development at Tolero Pharmaceuticals.
Alvocidib drug development has largely focused on the inhibition of cell cycle regulating CDKs. However, recent clinical activity in leukemia is not fully accounted for by cell cycle inhibition. We set out to more fully understand the mechanism of alvocidib particularly in leukemia patients that were benefiting significantly from the treatment. In addition to the cell cycle regulating CDKs, alvocidib has very potent activity against CDK9 (IC50=1.5 nM). CDK9 is responsible for controlling the transcriptional pause release that is important for the expression of certain genes. One of the most universally regulated transcripts by CDK9 is the anti-apoptotic family member, MCL-1, a member of the BCL-2 family involved suppressing cell death. Alvocidib treatment results in a concentration and time-dependent inhibition of MCL-1 expression in cancer cells. To further validate the CDK9/MCL-1 axis as the therapeutic target of alvocidib in, we utilized an approach called mitochondrial profiling to interrogate the dependencies that leukemia cells have on BCL-2 family members and retrospectively screened archived patient samples from a previously completed phase II clinical trial. We discovered that the patients that had a strong dependency on MCL-1, identified by a high NOXA priming score, where those that showed profound benefits from alvocidib. These findings led us to conduct an on-going phase II prospective biomarker trial where we are prescreening patients for NOXA priming and enrolling patients with a score ³40%. Taken together, these results highly support the conclusion that alvocidib works through a mechanism that targets CDK9 activity and MCL-1 expression.
- Cancer stem cell | Molecular Medicines for Cancers| Stem cell markers | Breast Cancer Stem Cells | Cancer Stem Cells in Brain Gliomas
Location: Waterfront 1
1Institute of Molecular Genetics, Italy
StemTek Therapeutics, Derio, Spain
Time : 12:30-13:00
Ines Prieto Remon is currently Senior Researcher at StemTek Therapeutics, a biopharmaceutical company located in the Basque Country, Spain. She earned her Bachelor of Biochemistry at the University of the Basque Country, Spain, in 2006, and her PhD in Molecular Biology and Biomedicine from the University of Cantabria, Spain, in 2013, working with Dr Carlos Pipaon Gonzalez and Dr Marian Ros. In her thesis work she studied signaling pathways in Fanconi anemia patients samples, regarding their aberrant acute sensitivity to chemotherapeutic agents. She also studied microphthalmia with Fancd2-/- mouse model. After her PhD, Ines accepted a postdoctoral position at the Laboratory for Experimental Hematology & DNA Repair, at Herman B Wells Center in Indianapolis, IN, US. There, she worked with Dr. Helmut Hanenberg in a project which aimed to study the protective effect of compounds in order to reduce/remove side effects of chemotherapy.
The cancer stem cell (CSC) concept has important implications not only for our understanding of carcinogenesis, but also for the development of cancer therapeutics. There is a growing body of preclinical evidence showing that cancer stem cells contribute to chemotherapy and radiation resistance in breast cancer. The use of drugs that interfere with stem cell self-renewal represents the strategy of choice for novel effective anti-cancer treatments, but also a great challenge because cancer stem cells and their normal counterparts share many pathways. The biology of cancer stem cells has proven complex and difficult to translate into effective therapeutic strategies. The question arises as: how do we test compounds for anti-cancer stem cell activity? The answer is: phenotypic screening. There are indeed several functional assays well validated in the scientific literature that have been used for years associated to the ability of cancer cells to demonstrate stem cell behavior. The most relevant is the 3D tumor spheroid assay. This assay has been used to uncover and culture stem cells from many tissues as well as from tumors. There are multiple reports now, that show that spheroid derived cells are enriched in tumor initiating or cancer stem cells, derived from cell lines and from natural fresh tumors as well. Here, we describe the use of 3D spheroid models to profile compound activity against cancer stem cells. Furthermore, a case of compounds preventing hypoxia-inducible transcription factor (HIFs) activity is presented. Recently, HIF transcription factor biology has been linked to pathways that regulate stem cell self-renewal and pluripotency, suggesting a new mechanism whereby HIF proteins may drive tumor growth, through the generation of tumour-initiating cells or cancer stem cells. Therefore, targeting the HIF pathway may provide a novel therapeutic avenue to target cancer stem cells. We demonstrate that interfering with HIF pathway activation prevents mammosphere formation, validated through independent confirmation through Sox2 promoter activation, Aldefluor® assay and in vivo proof-of concept experiments targeting
Mardin Artuklu University, Turkey
Title: Hypericum triquetrifolium Turra against cyclophosphamide-induced hemorrhagic cystitis in rats
Time : 14:00-14:30
Songul Cetik is an Assistant Professor at Vocational Higher of Health Services, Mardin Artuklu University in Turkey. She has completed her graduation from Eskisehir Osmangazi University, Faculty of Arts and Sciences, Department of Biology, 2009. She has done her Post-graduation from Eskisehir Osmangazi University, Faculty of Science, 2014.
A cyclophosphamide (CYP) usage is limited by side effects of it, are commonly used as antineoplastic drug. Hemorrhagic cystitis is one of the most important side effects of CYP chemotherapy. Antioxidants such as Hypericum triquetrifolium Turra (HT) show an important antioxidant and anti-carcinogenic properties with its rich contents. This study investigated the possible cytoprotection effect of HT (25, 50, 100 mg/kg, i.p., for 6 days) in CYP (150 mg/kg, single dose, i.p.) treated rats, and attempted to obtain a suitable new agents. Creatinin (CK), malondialdehyde (MDA), total oxidant capacity (TOC), total antioxidant capacity (TAC) and oxidative stress index (OSI) levels were measured in blood serum. Furthermore, the bladder tissue samples were investigated histopathological. In the only CYP group CK, MDA, TOC and OSI levels found increased while TAC level decreased. According our results high dose CYP caused the edema, necrosis, bleeding and tissue erosions, hemorrhage and separation of the muscle fibers supported the our biochemical results. After pretreatment with HT doses observed an important decrease in the CYP toxicity, decreased the cell damage and oxidative stress parameters while increased TAC. Based on the present experimental study’s findings, we may say that HT pretreatment has the potential to be a therapeutic option for the management of CYP-induced HC.
Gillies McIndoe Research Institute, Wellington, New Zealand
Title: Cancer stem cells in isocitrate dehydrogenase wild type glioblastoma express components of the renin-angiotensin system and cathepsins B, D and G
Time : 14:30-15:00
Tinte Itinteang serves as the current Chief Scientific Officer and the Evans Family Research Fellow of the Gillies McIndoe Research Institute (GMRI) in Wellington, New Zealand. He completed his Medical Training at the Melbourne University in 2001, and then completed his Basic Medical Residency in New Zealand, from 2008-2010. He completed his PhD from Victoria University of Wellington, NZ on the role of stem cells and the renin-angiotensin system (RAS) in infantile haemangioma. From 2012-2014, he was appointed as a Research Fellow at the Gillies McIndoe Research Institute, during which he spent six weeks at the Friedlander laboratory at The Scripps Research Institute in San Diego investigating the role of iPSCs for disease modelling. He was then appointed as the Chief Scientist of the GMRI from 2015. His work on the role of stem cells and the RAS in infantile haemangioma has been acknowledged with the International Society for the Study of Vascular Anomalies John Mulliken award as well as several national and international honours. He is the author of over 50 peer reviewed articles and has given over 100 presentations at international conferences.
Introduction: Isocitrate dehydrogenase wildtype glioblastoma (GB), the most aggressive form of brain glioma, is associated with a median survival of 25 months. Cancer stem cells (CSCs) have been proposed to be the origin of many cancers, including GB. Renin-angiotensin system (RAS) has been associated in CSCs in different types of cancers. This study aimed at identifying and characterising the CSC population within GB tissues for the CSC markers, components of the RAS, and the protease cathepsins B (CathB), D (CathD) and G (CathG), which provide potential bypass loops for the RAS. As well neuro-spheres derived from fresh GB samples were investigated their expression of stem cell markers, TRA 1-60, OCT4, SOX2 and SSEA-1, and the aforementioned markers.
Methodology: DAB and immunoflourenscent immunohistochemical (IHC) staining was performed on 7 GB samples for the expression of CSC markers SALL4, OCT4, SOX2, pSTAT3 and NANOG; components of the RAS, namely pro-renin receptor (PRR), angiotensin converting enzyme (ACE), angiotensin II receptor 1 (ATIIR1) and angiotensin II receptor 2 (ATIIR2); and CathB, CathD and CathG. NanoString mRNA analysis was performed on 5 of the original 7 GB samples, for the transcriptional expression of the same markers. 6 fresh samples of the original cohort of GB were grown in culture and stained for TRA 1-60, OCT4, SSEA-1 and SOX2, markers associated with CSCs. These cells were subjected to transcriptional analysis for CSC makers, components of RAS and CathB, CathD and CathG.
Results: IHC staining demonstrated a significant number of GB cells expressing SOX2 and pSTAT3. A subset of these expressed OCT4, SALL4 and NANOG. NanoString mRNA analysis demonstrated the expression of mRNA transcripts for the markers examined. Cultures of GB tissues yielded tumour-spheres which expressed TRA 1-60, OCT4, SSEA-1 and SOX2. These tumour-spheres also expressed mRNA transcripts for the CSC and RAS markers, CathB and CathD demonstrated by IHC staining on GB tissues.
Conclusion: The finding of this study confirms the putative presence of 2 CSCs within GB. The ability for primary cells derived from GB to form tumour-spheres in vitro, that express CSCs markers underscores the critical role of CSCs in the biology of GB. The expression of the components of the RAS, CathB and CathD, but not CathG by CSCs in GB and the GB-derived tumour-spheres, suggests CSCs as a novel therapeutic target by modulation of the RAS using existing medications.
Newcastle University, UK
Title: The expression of theThe expression of the classical stem cell markers in pancreatic adenocarcinoma cell line classical stem cell markers in pancreatic adenocarcinoma cell line
Time : 15:00-15:30
Hussain R Al-Turaifi obtained his PhD from North East England Stem Cell Institute, Faculty of Medical Sciences, Newcastle University UK and is focusing on Translation Medical Research through enrolling in Translational Medicine Program at The University of Edinburgh, College of Medicine and Veterinary Medicine, School of Biomedical Sciences Edinburgh. As Head of Referral Laboratory, Head of Blood Bank Donation Testing Center and Consultant of Molecular Pathology and Clinical Biochemistry he concentrates on diagnostic clinical laboratory at King Fahad Hofuf Hospital, KSA. He worked in academic field as a faculty of Biomedical School, Newcastle University and in the Department of Medical Biochemistry, College of Medicine, Dammam University, KSA.
Pancreatic cancer has been the third leading cause of cancer-related death in USA. Most of the cancer patients get diagnosed in late stage and this minimizes the effectiveness of surgical intervention to less than 20 percentage. Moreover, chemo-radio therapy is not curative thus the survival rate of patients with pancreatic cancer after 5-years was 7%. In USA, 53,070 new cases were estimated diagnosed with pancreatic cancer in 2016, while 41,780 patients was the estimated death from pancreatic cancers. Similar percentage was reported globally, estimated by World Health Organization in 2012 . Presence of cancer stem cells (CSCs) within pancreatic tumor was reported by several groups using unspecific biomarkers. Pluripotent transcription factors such as OCT4, SOX2 and NANOG, that upregulated in embryonic stem cells in contrast to somatic cells, were detected in various types of cancer tumors from adult patients. The aims of this study was to investigate the expression of the classical stem cell markers in pancreatic adenocarcinoma cell line (PANC1). PANC1 cells were characterized by RT-PCR/immuno-staining. Transient over-expression of stem cell promoter-driven reporter plasmid Oct4-eGFP was undertaken using Lipofectamine 2000 transfection reagent. Several embryonic stem cell markers and other cancer related markers were detected which illustrate the nature of pancreatic cancer.
National Cancer Centre Singapore, Singapore
Title: The activation of RAF/MEK/ERK kinase cascade by variable β3-αC loop deletions triggers oncogenesis
Time : 15:30-16:00
Hu Jiancheng received his PhD from University of Colorado Denver in 2007 and then Post-doctoral training at Washington University in St. Louis and Howard Hughes Medical Institute. Since 2014, he has joined National Cancer Centre Singapore where he has served as the Principal Investigator at the Laboratory of Cancer Signaling. He has published more than 15 papers in international renowned journals. His research interests include: (1) the regulatory mechanism of RAF kinase and other oncogenic protein kinases under normal/pathological conditions; (2) molecular basis that underlie intrinsic and acquired resistance of kinase inhibitors in clinic treatment of cancers; (3) the development of novel kinase inhibitors.
RAF/MEK/ERK kinase cascade has well-defined role in cancer development. Aberrant activation of this kinase by genetic alterations exists in >40% human cancers, which functions as a driver to trigger cancer pathogenesis. In current study, we identified a new catalogue of mutations in RAF and MEK with variable deletions of β3-αC loop. These mutants are constitutively active and highly oncogenic in vitro and in vivo. To develop strategies for targeting these mutants-driven cancers, we tested whether they were sensitive to RAF/MEK inhibitors that used for clinic treatment of BRAF (V600E)-harboring cancers or undergoing clinic trials, and found that all of them exhibit a strong drug resistance at cellular level and in tumor-xenograft mouse model. To explore molecular mechanism that underlies this phenomenia, we next carried out a serial of biochemistry and structral analysis and demonstrated that β3-αC loop deletions stimulate the homo-oligomerization of both RAF and MEK, which not only triggers their kinase activity but also dramatically decreases their drug affinity. Together, our study provides a solid evidence that RAF and MEK mutants with β3-αC loop deletions function as a cancer driver and a clear molecular basis that β3-αC loop deletions activate RAF and MEK and lead to strong inhbitor resistance, and appeals a development of new inhibitors.
King Faisal Specialist Hospital and Research Centre, Saudi Arabia
Time : 16:10-16:40
Monther Al-Alwan has completed his PhD in immunology from Dalhousie University and postdoctoral studies from University of Manitoba, Canada. He is a Scientist at the stem cell and tissue re-engineering program (SCTRP) at King Faisal Specialist Hospital and Research Centre and Adjunct Associate Professor at AlFaisal University, Riyadh, Saudi Arabia. Currently, he is actively involved in dissecting the molecular pathways that regulate the function of cancer stem cells and how this is related to chemoresistance and metastasis. He has more than 22 peer-reviewed publications in reputed journals and has been serving as an Editorial Board Member of various international journals.
There have been significant advances in breast cancer treatment, which have been attributed to the use of targeted therapy in combination with surgery and chemotherapy. However, the tumor-related mortality remained high mainly due to chemoresistance resulting in relapse and metastasis. Chemoresistance is widely believed to be regulated by a small subpopulation of the tumor bulk that possess stem cell-like features and thus are called cancer stem cells (CSCs). We have shown significant association between worse clinical outcome in breast cancer patients, including metastasis and shorter survival, and expression of fascin, an actin-bundling protein. Moreover, we have also reported that fascin is a critical mediator of breast CSCs and chemoresistance, via the activation of focal adhesion kinase (FAK), which is known to directly bind members of the integrin adhesion molecules. Here we have used fascin loss and gain of function approaches to examine if fascin influences integrin expression to regulate breast CSC function. Our results have demonstrated that fascin expression in breast cancer cells is directly associated with increased expression of integrins including: CD49a, CD49C, CD49f, CD29 and CD61. Fascin-mediated integrin expression on breast cancer cells enhances their adhesion, chemoresistance and tumorsphere formation ability. This study supports a role for fascin in the maintenance of breast CSCs via the regulation of integrin expression. The outcome of this study is expected to provide another evidence that fascin targeting may present a new approach for optimal treatment of breast cancer from the root.
Institut de Biologie Valrose- Ibv, France
Time : 16:40-17:10
Thierry Virolle is a Research Director (permanent position) at Institut National de la Santé et de la Recherche Médicale (INSERM), Head of the Team Cancer Stem Cell Plasticity and Functional intra-tumor Heterogeneity at the Institute of Biologie Valrose (iBV). He is Co-Founder of the French National Sud Cancer Stem Cell Network, SUNRiSE dedicated to the study of cancer stem cell. He is Doctor of Science (PhD) at Nice Sophia Antipolis University (2000), his researches focus on the regulation of the plasticity of glioblastoma cancer stem cells and its contribution in the genesis of functionally divergent tumor territories.
Glioblastomas are incurable primary brain tumors that affect patients of all ages. The aggressiveness of this cancer has been attributed in part to the persistence of treatment-resistant glioblastoma stem-like cells (GSC). We have demonstrated that microRNA cluster miR-302-367 has the potential to force GSC exit from stemness, promoting loss of stemness properties and tumorigenicity and revealing a great therapeutic interest. In our study we attempt to develop a cell-based therapy for miR-302-367 continuous delivery by taking advantage of the capability of glioma cells to secrete exosomes that enclose small RNA molecules. We engineered primary glioma cells to stably express the miR-302-367. Remarkably, these cells altered, in a paracrine manner, the expression of stemness markers, the proliferation and the tumorigenicity of neighboring glioblastoma cells. Further characterization of the secretome derived from miR-302-367 expressing cells showed that a large amount of miR-302-367 was enclosed in exosomes, which were internalized by the neighboring glioblastoma cells. This miR-302-367 cell-to-cell transfer resulted in the inhibition of its targets such as CXCR4/SDF1, SHH, cyclin D, cyclin A and E2F1. Orthotopic xenograft of miR-302-367-expressing cells together with glioblastoma stem-like cells efficiently altered initiation and tumor development in mice brain.