Tools and Technologies I
Statement of Benefit to California:
This application describes the identification of small molecule ligands for promoting self-renewal, differentiation, and maturation of stem cells. To achieve these ends, the applicant proposes to use combinatorial chemistry to create peptide libraries that are coupled to beads. This library will be used to screen various pluripotent cell lines that have been lentivirally engineered to express stage and/or cell type specific markers, in order to identify peptides that bind to cell surface receptors on these cells. Finally, the molecules of interest will be comprehensively assessed for their ability to promote stem cell behaviors such as growth, differentiation, and maturation, with a particular emphasis on cardiomyocyte generation. Investigators will use imaging, electrophysiology, proteomic, and microarray techniques to assess cellular responses. The reviewers were enthusiastic about the proposed technology and its potential advance the field of stem cell biology. The identification of small molecule ligands to manipulate self-renewal, pluripotency and directed differentiation of human embryonic stem cells (hESC) and induced pluripotent stem cells (iPSCs) would be an important advance. Equally, identification of ligands that promote cardiomyocyte differentiation might help in the effort to produce mature cardiac cells from hESCs for cell therapy. Other molecules identified in this screen could serve as markers for stem cells, to be used as imaging probes. Finally, an ability to mimic extracellular matrices might offer new scaffolding possibilities for improved tissue engineering. The potential impact of these studies was considered very high, both for basic research purposes and to advance the field of stem cell biology into the clinic. In terms of feasibility, reviewers agreed that this proposal would have a high likelihood of success due to the eminent qualifications of the investigators and a “first rate” experimental design. The principal investigator (PI)’s laboratory has created combinatorial libraries that were used successfully in other screens. In preliminary work presented in the application, the applicant identified ligands that promote hESC growth by screening one of these peptide libraries against mouse embryonic stem cells. In addition, the applicant has access to a wide diversity of embryonic cell lines available for this project. Reviewers complained that the research plan lacked experimental details, which impaired their ability to assess the project extensively – for instance, it was not clear how many lines would be tested, whether they would take into account gender differences in the lines, and what the size of the screening library was. One reviewer worried that the use of two peptides on a single bead might result in aggregation, a potential pitfall that was not addressed in the application. Despite these issues, reviewers commented on the proposal’s overall strengths in both concept and methodology, and expressed confidence that the project was feasible given the applicant’s success with previous screens. The strongest aspect of this application is the research team involved. The applicants were described as highly qualified to conduct the described experiments: the PI is a leader in the field of combinatorial chemistry, with extensive experience in the proposed methodologies, whereas the co-investigator provides valuable and complementary expertise with stem cells, cardiac differentiation and lentiviral delivery. Overall, this is an ambitious and feasible proposal from a talented team of investigators that could potentially advance the field of stem cell science.