The intent of the proposed shared research facility is to provide a state-wide resource for qualified scientists in California to study human embryonic stem cells (hESC) without federal restrictions. The shared facility will encourage a spirit of collaboration and include laboratories for investigators to culture, collect, store, and analyze hESC, provide necessary services that will be cost-effective and assist with research productivity, and ensure an environment that will facilitate the essential interactions among scientists. This approach will advance the use of hESC for regenerative medicine purposes and aid in developing new technologies and therapies for the treatment of human disease. Using established methods that have proven successful for other collaborative and service-based structures, this facility will encourage scientists to work together and provide the necessary resources to ensure their success. Investigators new to hESC research will benefit greatly by having this facility available because it will have a centralized supportive structure where experienced personnel will provide the necessary assistance and guidance. For those investigators with hESC research experience, new opportunities will be available to work with cell lines that can be obtained but not be used in laboratories that are supported by federal funding. This will greatly expand research programs that are focused, for example, on studying ways to differentiate hESC towards blood cells and vessels for the treatment of disorders such as sickle cell disease and vascular abnormalities associated with heart disease. In addition, regeneration of damaged organs such as the heart, lung, liver, or kidney may require methods to reconstruct these tissues using scaffolds on which to grow the cells. These approaches require the ideas of cell biologists, engineers, biomedical researchers, and clinicians working together, and testing these ideas to ensure the procedures are safe before considering treatments of human patients. Techniques such as those that focus on ways to monitor cells once they are injected into the body will provide a powerful tool to study the outcome of these therapies.
A techniques course will be offered to scientists, students, and staff which will result in more laboratories in California working with hESC. The cells are difficult to grow and specialized training is required by personnel that are highly skilled and can provide the necessary information and direction to ensure success. The course is designed to provide qualified applicants the training experiences that will reinforce the basics, and ensure they are able to establish these techniques in their laboratories. The training course will be offered 4 times during the calendar year and include presentations and ‘hands-on’ experiences. Continuing education through the facility will ensure trainees have the necessary support when they return to their respective laboratories.
The Translational Human Embryonic Stem Cell Shared Research Facility will serve the state and its citizens by providing unparalleled opportunities to investigators, and establish a model for the manner in which researchers throughout California can work together to advance the use of cellular therapies for the treatment of human disease. This facility will remove barriers preventing the transfer of promising stem cell therapies to human patients by connecting people with expertise and new ideas with the resources necessary to develop and to evaluate new technologies and therapies under the necessary conditions before they are assessed in humans. The California community will benefit from the results of this collaborative environment because it will facilitate and advance research findings, promote a culture of sharing, and educate and train a new generation of scientists in human embryonic stem cell research. This approach, the infrastructure and goals of the proposed facility, and the plans for the techniques course will all provide opportunities for scientists, students, fellows, and staff, and increase the number of qualified scientific and medical personnel that will be able to make new discoveries and ultimately improve health care for patients.
SYNOPSIS OF PROPOSAL: The applicants propose the establishment and maintenance of a non-federally funded shared research facility at UC Davis for the culture of hESCs, flow cytometry and cell sorting, quantitative PCR, immunohistochemistry, controlled rate cryopreservation, and essential services for the state-wide hESC research community. There is currently no such space on the UC Davis campus. This space will include three fully-equipped cell culture labs. The applicants are proposing to use hESC lines from five sources in these studies. Three are from the registry, one is from the Harvard collection, and one is new lines from the University of Minnesota and other institutions “to be determined”. Half of their work could have been funded by the NIH; however, establishment of new lines and derivation is part of the endeavor of the center which is not covered by federal funds.
QUALITY AND IMPACT OF THE SCIENCE: This application from Dr. Alice Tarantal is for 3 years of funding to establish a shared research laboratory off campus near UC Davis. It is predicted that the UC Davis facility will provide services for 32 investigators - 18 from UC Davis, 5 from UC Merced, 5 from Stanford, and 4 from Children’s Hospital of Los Angeles (CHLA). A broad range of scientific questions are being addressed by the 32 investigators, and this proposal aims to form a laboratory that is not focused on a few areas but rather covers the entire spectrum of differential lineages providing support for this large number of investigators. The basic biological studies performed in this facility have four aims: 1) the description of molecular pathways underlying epigenetic silencing in hESCs; 2) studies on changes in chromatin structure that occurs during differentiation of hESCs; 3) use of chemical biology to guide hESCs toward differentiation into specific pathways (e.g. hepatocyte differentiation); 4) the “potential” for establishing new lines. The two first aims address epigenetic control of hESCs, which one reviewer feels is interesting but certainly not of the highest priority in the hESC field or for CIRM. The third is sound, but it will be difficult for Davis to compete with other UCs and private universities all converging toward this goal with larger numbers of investigators and bigger collections of chemical compounds. Regarding the fourth aim, the meaning of “potential” is unclear in this context. Dr. Ericson is currently on sabbatical in the UK in Dr. Andrews’ laboratory, and upon his return he apparently will bring this technology to Davis. Nevertheless, many of the studies proposed will require the use of hESC lines that are not approved for federal funding. It is not exactly clear what this facility will do for the four CHLA investigators. The likely draw for these PIs may be their collaborations with Dr. Tarantal, who is a highly productive and helpful colleague. Finally, the applicants suggest the use of nonhuman primates to facilitate the development of new cellular therapies for human disease. An important potential strength is the presence of this primate center, thus positioning this proposal closer to clinical applications. One reviewer notes that the recommendation of the National Academy of Sciences currently prohibits mixing experiments between hESCs and non-human primate embryos. Therefore, any planned experiments in monkeys will have to be done in adults.
APPROPRIATENESS OF SPACE AND EQUIPMENT TO SCOPE OF PLAN: The shared research facility, which will be located about 1/2 mile off the main UC Davis campus near the primate center, will be 2,257 square feet of laboratory area which will house cell culture laboratories along with facilities for FACS analysis and cell sorting, a molecular core laboratory, cryopreservation and cell storage facilities. The facility that has been designed is excellent, a nice floor plan is provided, and the work proposed with hESC is appropriate for the proposed facilities. There will be three cell culture rooms. Two will have one biosafety cabinet, and the third will have two biosaftey cabinets. The third and largest cell culture room would also be used for the stem cell techniques course proposed. There will be a total of 12 incubators. Each lab will have one Zeiss Axiovert microscope with camera, a refrigerated centrifuge, and a microfuge. In addition, there will be two separate labs where investigators can work – one for histology with a cryostat and one for molecular work. The FACS sorting facility is directly adjacent to the cell culture rooms. Equipment proposed for this facility is adequate, which in addition to the equipment mentioned above includes CO2 incubators, RT-PCR systems, -80 and -20 freezers, cryotanks, and an irradiator for mouse embryonic fibroblast feeders. Space might be limiting for the amount of equipment and number of rooms suggested, especially if the first year follows as predicted by the university regarding its popularity, but the applicants have not made an effort to justify the size based on the number of proposed users or the types of hESC research that will be performed.
QUALITY OF MANAGEMENT PLAN: One of the major strengths of this proposal is how well the laboratory will be managed. This facility will be overseen by Dr. Tarantal in the capacity of Program Director (PD), who has demonstrated expertise in multidisciplinary collaborations, and in the direction of core facilities focused on stem cells. Dr. Tarantal has a lot of experience running similarly complex cores that require multiple employees, quality control, federal and state oversight, and security. Well-trained and experienced technicians and administrators will assist Dr. Tarantal in the management of this facility. Dr. Tarantal currently serves as the chair of the UC Davis Stem Cell Research Oversight Committee (SCRO) and the UC Davis IACUC. The issue of conflict of interest or at least the appearance of conflict of interest will need to be dealt with accordingly, as the head of this facility cannot be both head of the scientific facility and head of IACUC and SCRO. Reviewers are confident that this would be resolved. The PD has amassed an impressive number of supporting letters, yet one reviewer feels that a rather bureaucratic system has been developed for California investigators wishing to gain access to this facility. This process will require the preparation of an “Investigator Request for Access” form documenting signed MTAs, necessary approvals from campus committees (IRB, SCRO, IACUC, and biological use authorization), evidence of campus training in biological and chemical safety, bloodborne pathogens, and other required training and/or screening on an annual basis; evidence for prior laboratory experiences relevant to hESC research; and the completion of facility training requirements for all users. Once approved for access, investigators will be given key card access and user identification for online booking of the facility’s equipment and requests for reagents and supplies. This website will also include a cost structure for use of this facility (with hourly rates posted where applicable). This plan seems to place a great deal of faith in the administrative process of this shared laboratory. While important, these procedures should not take precedence over scientific inquiry. An overly ambitious set of criteria on the administrative level will act as a deterrent to serious scientists interested in this field. On the other hand, until the federal government loosens its restrictions on studies with hESC, this level of assurance of compliance is likely very important.
DISCUSSION: This institution has been approved for a good number of SEED grants, thus there is clearly a great deal of interest on the part of this institution in stem cell research. The goals of the center are straightforward, and the faculty possesses the requisite expertise for this kind of work. Reviewers generally believe they will do a good job. However, the science as presented is incredibly vague. The third aim, which is to look for compounds that change the epigenetic differentiation properties, is less well-developed than others in terms of the basic number of compound to screen and how to follow up on hits. One reviewer questioned the competitiveness of this institution in chemical biology given that this competency was not as well developed compared to other institutions. The description of the derivation of new lines in aim four is also problematic. One of the shared lab investigators is being trained in line derivation technology while on sabbatical, but relevant issues for line derivation such as the origin/availability of embryos, patient consent requirements, and any connection to an IVF clinic are all missing from the application. Progress in the research will depend on the availability of embryos, and not addressing these points lowered enthusiasm for this application.
There is a strong emphasis in the proposal on the fact that a non-human primate center will be located adjacent to the hESC lab, but the relationship of the primate center and the shared lab is unclear. One would assume the use of non-human primates in facilitating cell therapy development, but the downstream translational uses were not described in the application. Reviewers generally were not enthused by this prospect because they believe that the field is not quite ready to use non-human primates. Several discussants agreed that the applicants’ argument that it is valuable for the hESC lab to be located near the primate center does not make sense. Four investigators from CHLA were included based on their intention to use the non-human primate facility, but one reviewer had trouble judging the science because it was described too vaguely. Dr. Tarantal is a very good collaborator but the rationale for their inclusion as users is unclear. Another reviewer mentioned that the imaging proposed would be critical because one can’t sacrifice a large number of non-human primates, but the description of the imaging is very vague. This reviewer noted that the imaging instrument mentioned (microPET) may only suitable for rodents or other small non-human primates. Another discussant countered, stating that s/he believed this technology is being used for rhesus and marmoset studies. Finally, one discussant expressed mild concern over the location of the facility being adjacent to the primate center near the main campus. Another discussant pointed out that while this is not near the medical center it is easy to reach by a 15 minute drive, where traffic is not an issue.
PROGRAMMATIC REVIEW: A motion was made to recommend this Shared Research Laboratory application for funding. Panel members cited two main problems: 1) there is a lack of clarity around the relationship with and utility of the primate center and 2) the style of this plan (including the location off the main campus and the location of the collaborators) is not adequate to justify funding a core lab. One panel member noted even though this application has a number of flaws, this institution is trying to ramp up work in this area and so it should be funded. Another panel member stated that this represents our only grant with anything to do with non-human primates, and asked whether this is an important resource to build now. A discussant responded that this was in fact one reason not to fund because the applicant does not articulate the role of the primate center and what it contributes in the context of the shared lab. There are human reproduction techniques in use that weren’t ever tested in non-human primates. A panel member cited the availability of imaging resources once again, but there was no discussion of how they would be used in the context of their contribution to a unique shared lab resource. The motion to recommend this Shared Research Laboratory application for funding failed.
QUALITY OF THE PROPOSED TECHNIQUES COURSE: This application proposes a technique course to be conducted four times a year, each made of a two-day “intensive” period. The proposed course is too short, and clearly overambitious in proposing that students will learn about the growth of hESC in this short amount of time. In fact, the proposal reflects a real lack of experience with the derivation, culture, characterization and maintenance of hESCs. The cell cycle of a human embryonic stem cell is 24 hours; thus, at best, the students will be exposed to one cell cycle of analysis of hESCs. Moreover, the generation of embryoid bodies without analysis requires several days. During these two days, students are to be trained on feeders and hESC culture, morphological analysis and characterization, thawing cells (both feeders and hESCs), cell staining and flow cytometry, PCR and immunohistochemistry (of both differentiated and undifferentiated cells). This is only the syllabus for day one. On day two, the students are meant to learn passaging and cryopreservation, establishing cell banks, formation of embryoid bodies, and preparation of cells for karyotyping. In addition to the experiments of these two days, students would have a total of 4.5 hours on day one and 4 hours on day two of theoretical teaching, which includes issues from general techniques to discussions of MTAs, ESCRO and IRBs. This unrealistic scheduling, in addition to a lack of expertise among the resident faculty as compared to other California institutions, significantly dampened the enthusiasm of reviewers.
QUALIFICATIONS OF THE INSTITUTION: The institution is excellent, among the best of the UC system. The qualifications of specific trainers are also excellent as the applicants propose to bring in Dr. Meri Firpo from the University of Minnesota, who has extensive experience in teaching hESC culture courses, and Dr. Carolyn Lutzko from CHLA who has worked on differentiating hESC down pulmonary epithelial lineages. Unfortunately, the resident faculty are clearly behind in their own expertise of hESCs, and the type of program they are proposing will not help this situation.
DISCUSSION: There was no further discussion following reviewers’ comments.