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World Conference on Regenerative Medicine World Conference on Regenerative Medicine (WCRM) Leipzig, Germany, 23–25 October 2013
For the fifth time, the World Conference on Regenerative Medicine (WCRM) was held in Leipzig, Germany, an event every other year that is well worth looking foward to. Conducted entirely in English, it has quickly become a fixed date in the calendar of many applied research institutions or companies related to regenerative medicine and stem cell technologies – especially in German-speaking countries but also in Europe and beyond. The conference’s hallmarks are four parallel sessions, top-class international (keynote) speakers, a plethora of topics, a poster session with 300 contributions, approximately 1000 visitors from all over the world and innovative, new enterprises rubbing shoulders with established companies. Also noteworthy, many young scientists get a chance to showcase their projects to an appreciative audience. This year’s distinctive features were a 1-day regulatory workshop with the EMA’s Committee for Advanced Therapy and the first cooperation with a partner country, namely Canada. Thus, the ambassador from Canada was an unusual participant at the opening event. For a change, the conference started headon with the first keynote lecture – the opening ceremony was scheduled to take place in the evening. Before the first keynote speaker of the conference had even started, the visitors had to make up their minds which one of the four parallel sessions they wanted to follow. These covered an enormous variety of topics that ranged from basic research to clinical studies such as regeneration of most tissues (with an emphasis on bone and cartilage); stem cell differentiation and reprogramming, among others; tissue engineering; regulation; commercialization; technologies such as imaging, cell tracking, upscaling and
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automization; and materials such as bionanomaterials. Unable to visit them all, I describe here a personal selection of each day’s most impressive talks.
Anja C Rasch Biomedical copywriter, Falkenstr. 40, 23564 Luebeck, Germany
Day 1 Stem cell differentiation
Irvine L Weissman, Professor of the Institute for Stem Cell Biology and Regenerative Medicine, Stanford University (CA, USA), got the honor of giving the first keynote lecture of the conference  . The Weissman group has discovered that nearly every kind of cancer cell has a large amount of CD47 on the cell surface, protecting it against attack by the body’s immune system. The investigators have found that if they block the CD47 ‘don’t-eat-me’ signal by making use of antiCD47 antibodies, macrophages will consume and destroy cancer cells. Weissman said: “We can treat cancer effectively with anti-CD47 antibody.” His team recently received funding from the Medical Research Council, UK to start clinical trials addressing acute myeloid leukemia in cooperation with the University of Oxford (UK). “What happens with the cells when tissues are dissociated and put into cell culture?” Daniel H Rapoport (see also the related poster ) from the Fraunhofer Research Institution for Marine Biotechnology (Lübeck, Germany), pondered the question of plasticinduced cell plasticity  . A quick poll of the audience confirmed the prevalent hypotheses that primary cell cultures somehow keep their characteristics for days or weeks. Yet, adherence to plastic material is expected to have an impact. Rapoport cited data from his group isolating stem cell-like populations from exocrine pancreatic tissue. Using, for
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News & Views Conference Scene example, lineage-tracing technique (Cre/LoxP), cell tracking and gene-expression profiling, the results confirmed the exocrine pancreatic origin of the proliferating cells. Furthermore, microarray data were gathered from samples collected at tissue sampling and at each passage day. The results reflected rapid and profound increase of cell cycle genes and DNA remodeling genes within the immediate phase of culturing (tissue to culture transition), whereas functional pancreatic gene activity decreased. From passaging P1 onward, hardly any change in the expression pattern was detected. If cultured cells re-entered the cell cycle, they showed a stem cell-like behavior of their tissue of origin. Nanotechnology for regeneration
Björn Högberg, Assistant Professor of the Karolinska Institute (Stockholm, Sweden) captivated his audience when introducing DNA origami as a tool for drug delivery or for therapy. Högberg’s group used short single-stranded oligonucleotides as staples to twist double-stranded DNA into defined multitubular 3D shape. How to use DNA origami for drug delivery? For example, by simply using active compounds that intercalate within the DNA strands, like the anticancer drug doxorubicin (Dox). Various cell cultures were treated with the DNA-Dox structure, and a slow Dox release was shown. According to Högberg, manipulating cell signaling via receptor–ligand interaction is feasible, as well as using DNA ‘nanocalipers’ to suppress cancer-cell invasion. Professor H Kawamoto from the Riken Research Center in Yokohama (Japan) described in his keynote lecture how his group has rendered T cells antigen- specific to a molecule called PD-1. “PD-1 suppresses the immune response to attack cancer cells,” he explained. His group used induced pluripotent stem cell methods to render cytotoxic T cells into stem cells. Then, these were differentiated into T cells again, suitable for cancer immunotherapy as being able to detect cancer cells. Kawamoto was convinced that this could make a breakthrough in cancer immunotherapy, and said that efficacy and safety need to be tested in animal models in future. He speculated that the same strategy could be used for other tumor antigens like WT-1 for solid tumors or HER2 for breast and lung cancer. Kawamoto also pointed to possible side effects when normal cells also express the target antigen, then the normal tissue might be damaged. Day 2 Reprogramming & pluripotency II
Li Qian, Assistant Professor at the University of North Carolina at Chapel Hill (NC, USA), told us how her
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group directly reprogrammed cardiac fibroblasts into induced cardiomyocytes by introducing three transcription factors, namely Gata4, Mef2c and Tbx5 (GMT)  . Using an acute myocardial infarction mouse model, the introduction of GMT attenuated cardiac dysfunction (i.e., leading to reduced scar size). Currently, her group is investigating the molecular cornerstones of cardiac reprogramming in noninjury conditions, and the temporal and dosage requirements of GMT. Also, they want to improve the efficiency of converting fibroblasts into fully functional cardio myocytes by looking into potentially epigenetic hurdles or cues from the microenvironment like cytokines or shear stress. “We want to move ahead to the porcine model,” Qian finished her talk. Cell therapy for stroke & brain injury
Sean I Savitz, University of Texas Medical School (TX, USA), chose bone marrow mononuclear cells (MNCs) for cell therapy as they contain mixed cell populations, are rapidly isolated and have small cell sizes ranging from 5 to 9 μm. He started his talk with an overview of the various time windows suited for potential cell therapeutic interventions after ischemic injury, for example, for attenuating the post-ischemic proinflammatory response. Savitz presented pilot data from a trial with patients, where MNCs were isolated and injected (2 ml/kg). He also talked about a Phase II randomized trial for traumatic brain injury, stating that “In treated adult traumatic brain injury we see less injury occurring”. Tissue engineering I
Paolo Macchiarini, from the Karolinska Institute, talked about tissue-engineered replacement for patients suffering from respiratory tube damage. Specifically, decellularized – leaving only proteins and maintaining fibers – human airway constructs were repopulated with autologous epithelial cells and chondrocytes of bone marrow mesenchymal origin. Subsequently, the cellularized constructs were implanted in patients with end-stage airway disease and restored lung function. For example, in less than a month a vascularized neomucosa was noticed. In comparison with classical transplantation surgery, the recovery was more rapid and the patients do not require immunosuppressive drugs. For post-operational therapy, Macchiarini recommended pharmacological interventions, as G-CSF would boost in vivo cell seeding and engraftment, and erythropoietin would promote remodelling and suppress immune reactions. The dependence on organ donations, which also excludes children from regenerative therapy, led to consideration of artificial structures as an alternative
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approach. These should mimic the bioproperties of native tissue, provide extracellular matrix-like structure, allow for cell adhesion and be nonimmunogenic. “Take patients in early!” was the final recommendation of Macchiarini. Soft tissue regeneration
“Fecal incontinence has a similar incidence to diabetes.” Professor Andrea Frudinger, Medical University Graz (Austria), gripped the audience’s attention right from the start  . The strong impairment of quality of life was not only due to old age but also caused by damage during the vaginal birth process, when the thin muscle layer of the external anal sphincter is damaged. As the muscle cannot repair itself, noncontractable fibrotic material and fat tissue fill the gap. The Frudinger group used ultrasound to guide the injection of expanded skeletal muscle-derived cells into the external anal sphincter of 40 patients. Four weeks later, the analyses showed a response rate of 50–75%. Among the patients was an 85-year-old patient who was fully continent after 4 weeks. Frudinger announced the start of an international, multicenter Phase IIb study, consisiting of three groups with 75 patients each in Austria, Sweden, the UK, Germany and other countries. Day 3 Regeneration of sensory organs & CNS
Jane Sowden from the UCL Institute of Child Health (London, UK) presented the work of her collaboration with the Institute of Ophthalmology, also UCL, aiming at developing a photoreceptor cell-replacement therapy. Subsequent to successful transplantation of postnatal precursor cells that gave rise to new photoreceptors in adult retina in mice, the researchers had chosen induced pluripotent stem cells as a renewable source. Recently, they published data where rod precursor cells were grown in a 3D gel, labeled with GFP, and integrated effectively after subretinal transplantation and at, crucially, a certain developmental stage. To avoid genetic labeling of rod precursor cells for future clinical applications, they developed a FACS protocol using antibodies for cell surface markers to isolate cells at the relevant stages (for details see ). Sowden said that their current goal was to identify conditions for higher cone cell integration. She concluded that defining the optimal donor cell was a critical step. Regarding clinical applications the robust isolation of precursor cells as well as optimization for higher numbers were essential. Reprogramming & pluipotency III
Marius Wernig, Assistant Professor at the Stanford School of Medicine, gave a wonderful keynote lec-
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ture on direct lineage reprogramming of cell types derived from different germ layers  . He presented data – achieved in cooperation with Nobel laureate Thomas Südhoff – showing how three transcription factors, namely Ascl1, Brn2 and Myt1l, can convert mesodermal murine fibroblast to ectodermal-induced neuronal cells – skipping the pluripotent phase of induced pluripotent cells. But what were the mechanisms of induced neuronal reprogramming? Transcriptome analysis assigned significance to Ascl-1 that binds chromatin independently of Brn2 and Myt1. Further epigenetic experiments showed that Ascl-1 targets nucleosomes and acts as a pioneer factor at neurogenic loci, acting in cell-independent contexts. Ascl-1 preferentially binds in a closed ‘trivalent’ chromatin state. Chromatin immunoprecipitation sequencing analyses of Ascl-1 binding support the role of Ascl-1 as activating the induced neuron fate switch. Thus, the novel chromatin state emphasized the role of a more complex combinatorial histone code during reprogramming. The Canadian travel grant recipient Samer Hussein could not make it to Leipzig, and thus co-first author Mira C Puri presented the data of the project dubbed Grandiosa  . The unofficial name seems well deserved, as an international cooperative team that set out to investigate the transition stages from primary mouse embryonic fibroblasts to induced secondary pluripotent stem cells on the molecular level. Using a virus-free, inducible reprogramming method, samples were collected at different time points and subjected to various methods, such as whole-genome DNA methylation, chromatin immunoprecipitation, transcriptomics for both long and short RNA, and proteomics. To facilitate analyses and exchange of data, as well as to support mouse and human stem cell researchers, a website was created  . The researchers showed that CpG methylation was affected by the four ectopic reprogramming factors in the early stages of reprogramming. Promising new companies Walking the aisles of the conference along the approximately 50 new and established companies, several start-ups were especially interesting. For example, Cell.Copedia GmbH (Leipzig, Germany) gave cell sorting from whole blood a new, nonmagnetic twist. Based on affinity chromatography employing strepavidin–biotin interaction, a high amount of low-affinity antibody fragments positively selects cells to the column matrix. Nontarget cells are washed out. Then, adding biotin to the column releases the bound cells. “The low-affinity antibody fragments fall off, leaving the cells just minimally affected,” said Wilhelm Gerdes, CEO. In addition, the developers claim high
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yield and purity, envisioning the use of their system in operating theaters for autologous therapies. CytoMate Technologies BV (Zutphen, The Netherlands) came up with a neat development to remotely monitor cell culture flasks. A small camera is attached to a palm-sized platform, with its lens placed over the flask. The camera takes pictures at designated intervals, and the investigators can have them sent to their preferred mobile device (e.g., laptop, tablet or smart phone). In addition, temperature, cell numbers and confluence doubling times are logged, and the investigator is notified by e-mail if, for example, a specified confluence level is reached. Live recorded data, images and analyses are stored at the company’s server and can be accessed from any internet-compatible device. The next WCRM is scheduled for 21–23 October 2015 in Leipzig.
Information resources All abstracts from the conference are available at http:// wcrm.future-science-group.com.
Lakowski J, Han Y, Gonzalez-Cordero A. Retinal stem cell therapy: a biomarker panel for isolation of transplantationcompetentphotoreceptor precursors. http://wcrm.future-science-group.com/pdfs/pp-235.pdf
Wernig M. Direct lineage reprogramming towards the neural lineage. http://wcrm.future-science-group.com/pdfs/kl-010.pdf
Hussein S, Tonge P, Puri MC, Nagy A. Genome-wide analysis reveals multiple chromatin and transcriptional states duringreprogramming to induced pluripotent stem cells. http://wcrm.future-science-group.com/pdfs/op-103.pdf
Weissman I. Normal and neoplastic stem cells. http://wcrm.future-science-group.com/pdfs/KL-009.pdf
Frahm S, Rapoport DH. Tissue to culture transition. http://wcrm.future-science-group.com/pdfs/pp-102.pdf
Rapoport D, Frahm S. Does plastic induce plasticity? http://wcrm.future-science-group.com/pdfs/op-111.pdf
Qian L. Reprogramming fibroblasts towards cardiomyocytelike cells. http://wcrm.future-science-group.com/pdfs/kl-006.pdf
Frudinger A, Paede J, Kolovetsiou-Kreiner V, Marksteiner R. Skeletal muscle-derived cell implantation for the treatment of fecal incontinence: asingle center explorative clinical study with a 1-year follow-up. http://wcrm.future-science-group.com/pdfs/op-078.pdf
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Acknowledgements The author would like to thank S Danner (Fraunhofer EMB), for critically reviewing the article.
Financial & competing interests disclosure The author has no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties. No writing assistance was utilized in the production of this manuscript.
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