Day 3

Wednesday, August 16

STEM CELLS: CULTURE AND CREATING 3D MODELS

7:30 Breakfast Workshops (sponsorships available)

8:15 Chair's Remarks

Plenary Keynote Presentations
	8:25 Spatial-Mechanical Regulation of Morphogenesis, Malignancy and Therapeutic Responsiveness Valerie M. Weaver, Ph.D., Assistant Professor of Pathology & Lab Medicine, University of Pennsylvania Using a quantitative molecular approach, animal models, 3D mammary epithelial cultures (MECs) and mechanically-defined extracellular matrices (ECMs) we have been studying how physical force could modulate the normal and malignant tissue phenotype. We found that malignant transformation of the breast is associated with a sustained and progressive increase in ECM stiffness. ECM stiffness elicits these cellular effects by driving integrin aggregation to facilitate focal adhesion maturation through enhancement of Rho-dependent cell contractility. A growth factor transformed mammary epithelium that exerts abnormally high integrin-generated tension could be phenotypically-reverted if Rho-dependent force was normalized. Intriguingly, we recently found that matrix stiffness also alters the responsiveness of an epithelium to exogenous death cues including gamma radiation, chemotherapy and immune receptor activators. Thus, tissue homeostasis and apoptotic responsiveness appear to be functionally-linked to tensional-homeostasis through integrin-growth factor receptor-GTPase crosstalk. The relevance of this paradigm to tissue morphogenesis, malignant transformation and tumor therapy will be discussed.
	9:05 Functional Tissue Engineering and Disease Models David Kaplan, Ph.D., Professor and Chair, Department of Biomedical Engineering, Science and Technology Center, Tufts University School of Engineering  Tissue engineering, through the in vitro cultivation of human stem cells, bioengineered 3D scaffolds and bioreactors, provides new options for the study of tissue structure and function. These human tissues provide a new venue to the study of disease in vitro under controlled conditions that can fill an important gap between cell-based assays and human clinical trials.
	9:50 The Convergence of Science and Commercial Considerations in Tissue Engineering James W. Burns, Ph.D., Senior Vice President and Head of Drug and Biomaterial R& D, Genzyme Tissue engineering can be considered to be the use of therapy for influencing biology locally to repair tissue, regenerate organs, or restore health. We have used biomaterials, cells, genes, and drugs in combination and on their own, for tissue engineering applications in orthopaedics, burn repair and cardiovascular medicine. My talk will focus on the technical development of our tissue engineering approaches and lessons learned on how science and commercial considerations converge in developing successful tissue engineering therapies.

10:30 Coffee Break, Poster and Exhibit Viewing

SOURCES, ORIGINS, AND FATE

11:30 A Good Ending Makes a Good Beginning - Telomere Function in Stem Cells
David Keefe, M.D., James M. Ingram Professor and Chair, OB/GYN, University of South Florida
Telomeres have long been known to mediate senescence in somatic cells. Virtually all stem cells express telomerase, the enzyme responsible for maintaining telomere length, but mammalian oocytes and early embryos do not. So how do reconstituted embryos reset telomere length? Recent work from our laboratory has shown that early embryos can reset telomere length during early embryonic development even in the absence of telomerase.

12:00 Challenges of Primate Embryonic Stem Cell Research
Carol Brenner, Ph.D., Associate Professor, Department of Biology, University of New Orleans
Embryonic stem (ES) cell research is controversial to some because producing ES cells requires destroying embryos. However, some of the surplus human embryos available from in vitro fertilization (IVF) clinics may have a high rate of genetic errors and therefore would be unsuitable for ES cell research. An insurmountable problem is that there are no human ES cells derived from in vivo-produced embryos to provide normal comparative data. In contrast, some monkey ES cell lines have been produced using in vivo-generated, normal embryos obtained from fertile animals; these can represent a "gold standard" for primate ES cells. In this review, we argue a need for strong research programs using rhesus monkey ES cells, conducted in parallel with studies on human ES and adult stem cells, to derive the maximum information about the biology of normal stem cells and to produce technical protocols for their directed differentiation into safe and functional replacement cells, tissues, and organs. In contrast, ES cell research using only human cell lines is likely to be incomplete, which could hinder research progress, and delay or diminish the effective application of ES cell technology to the treatment of human diseases.

12:30 Reconstructing and Deconstructing the Progression of Human Squamous Cell Carcinoma in 3D Tissue Models
Jonathan Garlick, DDS, Ph.D., Professor, Division of Cancer Biology and Tissue Engineering, Tufts University

1:00 Lunch on Your Own

EXPLORING SCREENING TOOLS

2:00 Chair's Remarks

2:05 Applications of ES Cell-Derived Neurons in a High-Throughput Screen to Identify Potentiators of the AMPA Subtype of Glutamate Receptors
Marsha Roach, MS, Scientist, Pfizer Global Research and Development
Schizophrenia is a chronic, highly debilitating mental disorder afflicting over 3 million individuals in the US. Current antipsychotic therapy effectively treats the symptoms, but not the cognitive deficits. Since hypofunction of glutamatergic neurotransmission has been implicated in the pathophysiology underlying schizophrenia, the identification of compounds that potentiate the AMPA subtype of glutamate receptors may offer therapeutic benefit. We have developed a novel high throughput screen using murine embryonic stem cell-derived neuronal cell types to identify compounds that potentiate AMPA receptors. This technology was utilized in a functional high-throughput screen for novel allosteric potentiators of the AMPA subtype of glutamate receptor. Initial validation studies confirmed the expected range of potencies for reference compounds. Key benefits of this technology include the capability for a full file screen using a single batch of cells and the ability to assay endogenously expressed neuronal receptors in physiologically relevant cell types.

2:35 High-Content Drug Screening with 3-D Bioengineered Muscle
Herman Vandenburgh, Ph.D., CSO, Myomics, Inc.
Muscle (cardiac, smooth and skeletal) is a complex tissue with numerous, interdependent biochemical pathways which regulate muscle contractility. We have developed a high-content physiological screening method for quantifying muscle force generation using 3-D bioengineered muscle in a 96 well format. Using an in vitro physiological assay will help identify muscle-active compounds and eliminate those exhibiting adverse side effects prior to testing in vivo. This technology is currently used to screen for drugs capable of attenuating skeletal muscle wasting disorders using primary adult rodent and human skeletal muscle stem cells.

3:05 3D Culture of Ovarian Follicles: From Immature Eggs to Live Births
Lonnie Shea, Ph.D., Associate Professor, Chemical and Biological Engineering, Northwestern University
In vitro fertilization techniques are not applicable to all women with compromised fertility. We have designed microenvironments for the culture of immature mouse follicles that enables coordinated growth of germ and somatic cells leading to follicle growth and maturation of the oocyte, which can be fertilized and subsequently re-implanted to obtain births of healthy pups. Successful maturation results from soluble factors in the culture media, insoluble factors in the 3D environment, and the mechanical properties of the environment. This 3-D culture model can serve as the basis for high-throughput screening to identify drugs that promote or limit follicle maturation. Additionally, this technology may enable egg banking as a means to preserve fertility for cancer patients or other fertility disorders.

3:35 Refreshment Break, Last Chance to View Poster and Exhibits

4:15 Human Organoids for Immunogenicity and Immunotoxicity Testing
Christoph Giese, Ph.D., Department Head, Tissue Engineering-Cell Assays, ProBioGen AG
Preclinical evaluation of biological drug candidates, e.g. antibodies, vaccines and growth factors, demands novel predictive cell-based assays. Species specificity of those tests is crucial for valid efficacy and side effect data. Three-dimensional tissue culture of the respective species, e.g. human, seems to be mandatory. In contrast to chemical drugs, any injection of biologics induces a light to severe immune response. In cases of vaccines this particular response is the most wanted. For therapeutical antibodies for example, immunogenicity is a no go criteria. We will present concepts of organoid culture, to address vaccine efficacy and immunogenicity of biologics. Complex co-cultures of human dendritic cells and lymphocytes were performed in miniaturized perfusion cell culture devices. Cytokine patterns, histological analyses and cell imaging were used to evaluate functionality and responsiveness of these tissue cultures. First attempts were made to multiplex, miniaturize and automatize the systems for later high content. Process performance was evaluated for robustness, consistency and standardization.

4:45 High-Throughput Approaches to Drug Delivery and Tissue Engineering
Daniel G. Anderson, Ph.D, Research Associate, Chemical Engineering, Massachusetts Institute of Technology
The combinatorial, automated high-throughput synthesis and evaluation of small molecules has revolutionized modern drug discovery. We have developed platforms that enable the rapid synthesis and testing of large libraries of synthetic biomaterials for delivery of genes, RNA, and drugs, and for characterization of cell interactions. Using the methods, we have identified degradable polymers with excellent in vivo delivery properties, and identified materials that offer new levels of control over stem cell behavior.

5:15 Panel Discussion

5:45 Close of Day