Human ES (hES) cells offer the opportunity to be converted into replacement tissues for diseased organs and provide cures for diseases like Parkinson’s, diabetes, and a host of neurological disorders. Unfortunately due to political considerations, scientific space containing equipment and other resources provided by the federal government are off limits for work on unapproved hES cell lines. Space where unimpeded hES cell work can be carried out is a major limitation for many scientists at our institute to initiate scientific inquiry in the very exciting field of regenerative medicine. We are applying for funds for a Shared Research Laboratory Grant to work on non-federal government-approved hES cells, which are superior to the approved hES cell lines. Eighteen faculty here have no laboratory space in which to do research in hES cells. We are requesting funds to renovate and equip ~2,000 sq ft of independently-operated, stand-alone space which will include laboratory space for generating lentiviral vectors (LV) capable of delivering genes and/or small interfering RNA (siRNA) to manipulate hES cells, and tissue culture facilities to grow and propagate non-NIH approved hES cell lines. To study and to induce differentiation of hES cells to different lineages we will require the availability of tools that activate or suppress gene expression. Furthermore to correct a genetic defect, for instance a defective insulin gene that leads to type I diabetes, one needs to introduce a functional insulin gene in the regenerated pancreas from the ES cells of a diabetic patient. If the introduced gene is not integrated in the genome, it will be lost in the subsequent progeny. Thus the need for a delivery vehicle that will become part and parcel of the chromosome in both the progeny cells and the self-renewing cells. We have developed delivery vehicles based on retro- and lentiviruses that can safely and efficiently deliver and transcribe genes in both the embryonic and adult stem cells. We believe that we can provide such vectors to all the stem cell researchers in the neighboring institutions working on CIRM related hES cell projects, which will not only be cost effective, but accelerate the pace of science in the exciting field of hES cells. We plan to hire a full-time core director and 3 research assistants who will be responsible for setting up and maintaining the stem cell facility and managing the core facility for generating viral vector. We will offer a hands-on laboratory course to all participants in CIRM related research to generate retro- and LV, safety associated with use of these vectors and state-of-the-art methods to utilize hES cells. We hope to train the next generation of human stem cell biologists who will play an essential role in bringing the fruits of regenerative medicine to the people of California and the world.
Statement of Benefit to California:
Human embryonic stem (hES) cells offer the opportunity to be converted into replacement tissues for diseased organs and provide cures for diseases like Parkinson’s, diabetes, and a host of neurological disorders. To realize the potential of this revolutionary concept, scientists must understand the basic mechanisms of how ES cells can become a liver, pancreatic cell, muscle, or neuron. What are the signals that trigger differentiation to a specific cell lineage and what molecular events must transpire to allow self-renewal of the ES cells? Eighteen faculty here wish to pursue answers to these fundamental questions. Several scientists want to understand the maintenance and self-renewal capacity of hES cells. Other investigators are proposing ways to induce hES cells to spinal chord or dopaminergic neurons. Converting hES cells to a variety of cell types will require genetic manipulation; therefore we are proposing the generation of delivery vehicles (vectors) that can be used to introduce or inactivate genes into hES cells. Because most of the scientists here do not have a facility to work on non-federally funded hES cell lines, we are requesting funds to build such a facilty. We believe that scientific work carried on hES cells will help to cure diseases like Parkinson’s, Lou Gehrig’s disease, and diabetes. The work proposed here will lead to establishing biotech companies in California to expand basic research into products thereby not only improving the health of Californians, but also become a significant economic engine. The facility we are proposing will provide gene delivery vehicles to scientists from neighboring institutions working with CIRM funds that will be cost effective and establish collaborations. The facility will also train the next generation of stem cell biologists to continue this very important scientific work and keep California in the forefront of biomedical research.
SHARED LABORATORY SYNOPSIS OF PROPOSAL: This is an application for a facility to develop retroviral and lentivirus vectors for use in the genetic manipulation of human embryonic stem cells (hESCs). Included are plans for a tissue culture space to manipulate non-presidential hESC lines. The ~2000 square foot facility will be renovated and equipped with the solicited grant money and will be an addition to the already extant 1000 square foot facility currently serving the needs of two faculty. The facility will be housed at the Salk, will serve as a core for investigators from The Salk Institute, the Burnham Institute for Medical Research, the University of California, San Diego and the Scripps Research Institute, and will be led by Dr. Inder Verma. A total of 18 faculty members are listed as potential users. A course is also proposed that will focus on the generation of viral vectors to manipulate human stem cells. QUALITY AND IMPACT OF THE SCIENCE: A large portion of the work in this core focuses on production of viral vectors. While the Program Director (Dr. Verma) is a leader in this area, this work is not likely to be focused exclusively on hESC manipulation. However, this submission has been coordinated with the other institutions noted above, with each institution submitting a core proposal that focuses on a unique aspect of hESC technology. The proposed users of the core facility outside the vector core have outstanding track records and propose a wide range of research varying from determinants of ESC totipotency and proliferation, to lineage-specific differentiation factors. Noted cell systems under study include the central nervous system, hepatocytes and hematopoietic cells. Thus, while the proposed research is not unique, it is clearly important to the field. Two researchers, Drs. Gage and Belmonte, already have active hESC programs that utilize space developed with philanthropic funding. The proposed faculty members have outstanding track records with widely varied expertise encompassing structural biology, cell signaling pathways, transcription factors, and cell biology with publications in the highest ranking journals. There are multiple members of the National Academy of Sciences among the proposed core users as well as researchers approved for CIRM awards. APPROPRIATENESS OF SPACE AND EQUIPMENT TO SCOPE OF PLAN: The facility proposed includes a viral vector core contiguous with a tissue culture facility. The Salk Institute already has a non-NIH hESC space that is apparently utilized primarily by Drs. Gage and Belmonte. The proposed hESC facility will house 4 laminar flow hoods, necessary tissue culture equipment, and work stations dedicated to biochemical and molecular work (6 stations encompassing 1.5 benches). The proposal calls for the purchase of a high-speed sorter (since genetic manipulation of hESCs will likely not be 100% efficient). There is contiguous office space. The proposed space is modest if all proposed users are truly active in using the core facility. The proposed core also includes a high-end live imaging spinning disk confocal capable of monitoring proteins in 3D over time (Salk to pay 50% of the costs). The proposal calls for the overall facility to be under the direction of Inder Verma (5% effort) with day-to-day operations to be the responsibility of a Core Director (to-be-named; proposed as a Ph.D.-level individual, 75% effort), three full-time research assistants (only 1 listed in key personnel on 100% effort, the second will be funded by Salk, the 3rd by the course budget-see below) and a part-time administrative coordinator (25% effort). QUALITY OF MANAGEMENT PLAN: The management plan includes an oversight committee made up of John Young (Chair), Steve Kay, Jerry Olefsky and Stuart Lipton. All members have requisite experience in management of programs and cores to serve in this capacity. Dr. Verma is particularly suited to the unique focus of this proposed core in the area of virus vectors. The Core Director will be responsible for scheduling the tissue culture spaces and use of equipment. The Core Director will also be responsible for maintaining hESC lines, ensuring ‘safe practices’, and ensuring access to faculty from other institutions to the facility. Much of the management plan is not detailed but apparently will rely largely on the Core Director who is to be named. There is a letter of support from individuals at the four institutions explaining the strategy of each institution submitting separate grant applications, although there appears to be no formal support letter from the leadership of these institutions. DISCUSSION: Reviewers agreed that the quality of the institution and investigators is strong, and the viral tools may have many different applications, but the proposal is not well put-together with a sound management plan. This is an outstanding Program Director with a real track record in the field, it seems that the space would be well-utilized, and the scientific impact would likely be important. A reviewer commented that the space requested might be a little small and it would be advantageous if Dr. Gage were more involved in the management and oversight, as the management plan leaves much to be desired. The application is focused on retroviruses and lentiviruses, which are interesting and will compliment the other San Diego consortium members. In fact, this application is written largely as a vector core. Some reviewers questioned the applicant’s underlying assumption that viruses are needed for hESC research. A discussant pointed out that a strong argument can be made that viral vectors will have many different applications. In addition, reviewers described the consortium approach as thoughtful; while all 4 institutions in the Mesa group will be doing hESC work, they will each have their specialization and their own strengths. A reviewer points out that strategically this is a good idea, although the details are not worked out fully. Another considers it important to have multiple sites to do this work given the large number of people in the area who are doing stem cell research. PROGRAMMATIC REVIEW: A motion was made to recommend this Shared Research Laboratory application for funding. One reviewer believes that this would be a weak shared research laboratory and that it would not be likely to benefit the entire region. However, from a portfolio point of view, another points out that there are advantages to this application, and it would be hard to find a better place than this to do the work. The question was raised as to how important it is for the state to have a shared research laboratory with this emphasis? Several Working Group members echo the point that the real benefit to be gained is the vector technology and that this program director is the best person to lead this effort. The motion to recommend this Shared Research Laboratory application for funding passed. TECHNIQUES COURSE QUALITY OF THE PROPOSED TECHNIQUES COURSE: The course will be focused on the generation of viral vectors to manipulate human stem cells. A similar course is not currently in existence. The proposed course includes 12 sessions, with 3 out of 12 of these sessions focused on hESC manipulation. However, the proposal also states that the course will include a focus on research strategies and state-of-the-art methods to maintain, characterize, and utilize hESCs. The course will be ‘continuous’ and will include all levels of trainees. The faculty for the course will be identified at the beginning of each academic year and the identified faculty will present a seminar that fits into the course every month. Thus, overall, the major focus is on viral vectors and the proposed course planning is still somewhat preliminary. The course will be run by Dr. Inder Verma and the Core Director (to be named). The primary organizers of the course will be Drs. Verma, Gage and Belmonte. The participants will be appointed after review of their applications, and are expected to be “highly recommended by scientists familiar with their performance” and from laboratories with projects approved for CIRM funding. The course budget includes Dr. Verma (10% effort), the Core Director (to be named, 25% effort), a technician (100% effort) and an administrative coordinator (25% effort). QUALIFICATIONS OF THE INSTITUTION: The diversity of the potential faculty and the quality of their science would predict that this course would be highly meritorious. However, the details for the course seem focused on virus vectors with few details provided on the hESC component. DISCUSSION: A reviewer points out that the course topics all have to do with viral vectors and that this institution is capable of mounting a dynamic course, but that this application is a poor effort. The description of the course is considered too vague and resembles a seminar series as there was not an explicit discussion of a laboratory component to the course. Another discussant agrees and points out that with time and further development, the course would provide distinctive training for those using sophisticated vectors. PROGRAMMATIC REVIEW: A motion was made to recommend this Techniques Course application not be funded and this motion passed.