The ability of human pluripotent stem cells (hPS) to both self-renew and differentiate into virtually any cell type make them a promising source of cells for drug research and development, toxicity screening, developmental research and cell based therapies. Indeed, the derivation of patient matched cells from induced pluripotent stem cells derived from patient skin samples has the potential to treat conditions where cells are lost to disease or injury. However, stem cell identity, purity and scalability remain formidable challenges for translating pluripotent stem cell research to clinical applications. There is a lack of highly efficient methods for directed differentiation from hPS cell lines to mature cell types o interest and scale-up from hPS cells is costly. The derivation of scalable (self-renewing) embryonic progenitor stem cell lines offers a solution because they are well defined, clonally pure, simpler and less costly to produce on an industrial scale. Previous studies have demonstrated that a highly diverse collection of hundreds of distinct scalable embryonic progenitor cell lines can be isolated from human embryonic stem (hES) cells. Recent studies have demonstrated the utility of one of these lines for repairing femoral condyle cartilage and bone in an animal model. The production of scalable patient- matched progenitor stem cells and genetically diverse banks of progenitor stem cell lines for drug screening or disease modeling will require repeated consistent progenitor isolation from donor derived induced pluripotent stem (iPS) cell lines. We propose here to develop peptide based reagents to enable simple cost effective isolation of scalable progenitor cells of known scalability and differentiation capacity from pluripotent cell lines. In phase I, we will demonstrate feasibility of using cell targeting peptides selected for their ability to bind the scalable osteogenic progenitor cell line, SM30, to derive the cell line from hES cells. The re-derived SM30 cell line, SM30R, and similarly derived cell line isolates from iPS cell lines will be compared to the original SM30 cell line.
In Aim 1, w will isolate the SM30 cells from hES cells using peptide targeted quantum dots and compare them to the original SM30 cell line by gene expression analysis and functional differentiation assays.
In Aim 2 we will characterize the scalability and stability of the cell lines and in Aim 3 e will isolate SM30R progenitors from a variety of donor derived iPS cell lines. In phase 2, we will identify peptides for additional progenitor cell lines, formulate commercial peptide conjugates for cell labeling, and test applicability on a broad range of iPS cell lines. The long term goal is to develop processes, reagents and kits for progenitor cell isolation that will be marketed to both the research community and companies developing human cell lines for drug development and cell therapy.
Stem cell technologies offer great promise for revolutionizing human health care by providing cells for improved disease modeling, drug testing, and for cell transplantation to treat degenerative disease and injury. Manufacturing the desired stem cell types in sufficient quantity and purity remains a significant challenge for the field. Consistent isolation of the same stem cell type from multiple human donors is particularly challenging. We propose to develop reagents and methods to enable streamlined reproducible isolation and scale up of pure human stem cell types from reprogrammed donor cells.
