Tools and Technologies I
Human embryonic stem cells, also known as pluripotent stem cells, are recognized as a valuable tool for advancing our knowledge of human development and biology, and also for their potential in regenerative medicine. Recently, new embryonic stem cell derivation technology has allowed scientists to derive the equivalent of human embryonic stem cells from adult cells, rather than from human embryos. This important advance allows for the possibility of eliminating the problem of immune system rejection of transplanted cells. The use of embryonic stem cells, because they have thus far been derived from human embryos, is controversial. Recent technical innovations, however, have shown that there are now new opportunities for scientific progress with human pluripotent stem cells obtained from sources other than human embryos. Studies using human skin cells have shown that these skin cells can be directly reprogrammed back to their embryonic stem cell state without going through an embryonic state; thus, no embryos are created or destroyed using this new technology. These human pluripotent stem cells obtained from skin have been termed induced pluripotent stem cells (iPSCs) as they have been induced by the forced expression of four human protein factors introduced into these skin cells using viruses. iPSC technology holds the promise that iPSCs can be derived from an individual's skin cells and from this state be triggered to form heart, brain and other cell types required for regenerative repair. As these cells may be derived from an individuals own skin and transplanted back into the same individual, the derived repair cells are likely recognized as self and thus unlikely subject to immune rejection. Currently, the iPSC technology is unsuitable for use in humans as the viruses used to express the four factors required to induce the formation of the iPSC become part of DNA of the cell and are closely associated with causing cancer or other life-threatening diseases. Here, we propose a study to side step this difficulty by using a novel reagent to actively express the same four factors expressed in the virus-derived iPSCs but without the use of virus. Introducing active proteins into the cell in a non-integrative fashion helps eliminate the risk of cancer. When successful, the iPSCs derived using this technology will be safe for use in humans and this will allow a faster movement of successful patient-derived stem cell therapies into the clinic.
Statement of Benefit to California:
Human induced pluripotent stem cells (iPSCs) derived from differentiated human adult tissue cells following viral-driven expression of a small number of “reprogramming factors” are a monumental discovery. The possibility of translating iPSC technology to patient-derived stem cell therapy is nothing short of revolutionary and is of immediate benefit to every Californian and, indeed, all of humankind. This proposal will apply pre-existing, California-invented technology, in the form of a novel reagent to deliver the same active “reprogramming factors” directly to the nucleus of fibroblast cells. The resulting iPSCs will be suitable for clinical use as they are directly derived from the active proteins in the absence of cancer-associated viruses. This project is highly relevant to the goals of CIRM, and will result in the removal of a major block in the translation of iPSC technology to the clinic, namely the presence of random integrative cancer-causing viral sequences in the genome. The safe induction of patient-specific iPSCs will allow safe, effective and immune-tolerated stem cell therapies for the people of California and the world. This successful technology will have an immense economic benefit to California and illustrate that the people of California were justified in their support of stem cell research and have helped propel this research into the clinic for Californians and the world.
The goal of the research described in this proposal is to develop an improved method for generating induced pluripotent stem cells (iPSC). Established approaches for iPSC derivation require viral vectors that may cause insertional mutations, and thus dysregulation of endogenous genes, or other genetic damage to reprogrammed cells. The proposed method involves delivery of reprogramming proteins directly into cells and into cell nuclei in the absence of viral transduction. The proposed research has a high potential impact in that it could yield iPSC without the use of viral vectors. This would represent an important advance in the generation of pluripotent cells. The approach would facilitate the generation of patient-specific or disease-specific stem cell lines for basic or translational research and eventual clinical application. The rationale for the approach is clear, and the outlined experiments are relatively straightforward and an obvious extension of pioneering work carried out by others. The applicant presented some preliminary evidence that a protein can be delivered into cells. However, reviewers had strong reservations about the likelihood for success, because the applicant did not present a systematic strategy for optimizing components and procedures, assessing the quality of recombinant proteins, evaluating the activity of individual factors, and standardizing the dose, order, and frequency of delivery of reprogramming factors. Reviewers were concerned that experiments to evaluate the role of epigenetic factors were poorly developed and utilized reagents that could damage cells. Moreover, reviewer enthusiasm was further diminished by the lack of novelty or creativity in the approach. The applicant is a junior investigator, has published only one research paper as a senior or first author and has no apparent experience in leading a research program.