Understanding hESC-based Hematopoiesis for Therapeutic Benefit

Funding Type: 
Comprehensive Grant
Grant Number: 
Award Value: 
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 

Hematopoietic stem cell transplantation is the treatment of choice for many hematologic malignancies, and it is used to treat an expanding number of congenital blood disorders. However, only ~30% of patients who can benefit from this treatment have a matched sibling that can serve as the ideal donor. While the national marrow donor program and umbilical cord blood programs provide unrelated donor cells to many patients lacking a sibling donor, a large percentage of patients remain without a suitable donor, leaving them with suboptimal therapeutic options. This problem is more severe in certain ethnic populations, including people of Latino and Asian descent, groups that constitute a large part of California’s population. New sources of therapeutic hematopoietic stem cells are therefore needed.

Human embryonic stem cells, with their unlimited self-renewal capacity and their ability to generate all human cell types, provide a novel and exciting opportunity to obtain hematopoietic stem cells, thereby filling a critical therapeutic void. However, many hurdles remain before this vision can be realized, including the identification of more optimal human embryonic stem cell lines and better methods to direct the development of specific cell types from embryonic stem cells.

This proposal seeks to shed new insight into how we might better control and direct the development of human embryonic stem cells into therapeutically useful hematopoietic stem cells that can be used for transplantation. Our effort focuses on understanding how a specific class of small RNAs, called microRNAs, regulates the differentiation of human embryonic stem cells into specific cell types. We aim to uncover the identity of microRNAs that are important for this process, which will serve as useful biomarkers, or guides, for evaluating the therapeutic suitability of existing and newly derived human embryonic stem cell lines. In addition, we will develop techniques and reagents to modulate the expression of these critical small molecules to help direct human embryonic stem cell development for clinical therapeutic utility.

Statement of Benefit to California: 

This proposal seeks to understand how small RNA molecules help direct the differentiation of human embryonic stem cells into specific cell types suitable for therapeutic use. Our focus is the development of hematopoietic stem cells for the treatment of blood disorders. Benefits to the State of California include:

A new source of hematopoietic stem cells for patients who lack a matched donor. Some ethnic groups, such as people of Latino and Asian descent, are more likely to lack a donor, so those groups may benefit the most.
Hematopoietic stem cells have been shown to have the potential to participate in the repair of a wide range of tissues, including heart tissue, so human embryonic stem cell-generated hematopoietic cells may be useful in treating disorders outside of the blood system per se.
We will develop databases and technologies that could have wide-ranging use in the human stem cell field. To the extent that these can be commercialized, the work could be of financial benefit to California.
The kind of work proposed could attract additional economic interests to the state of California because our methods and reagents could be used by California businesses aimed at treating those afflicted with blood disease.
This work will provide new jobs for the University of California, San Francisco, and hopefully spawn additional research in the California academic and private sectors.
The proposed research is cutting-edge, and successful accomplishment of our goals would help put California at the forefront of human embryonic stem cell research, rewarding Californians for their support of proposition-71, and inspiring others to support human embryonic stem cell research.

Progress Report: 

We made good progress during the 08-09 funding period. We finished the critical work of screening 8 independent hESC lines for their ability to generate hematopoietic cells. This included 6 non-NIH-approved lines, studies that could only be performed through the innovative mechanism of CIRM funding, not through federal funding mechanisms. Interestingly, we found no meaningful differences among the lines, and are now able to focus on a single line. As a key part of achieving our goals, we generated a robust data set of miRNAs expressed in 3 different hESC lines and 4 independent human HSC samples. These data are now being scrutinized to identify possible miRNA candidates to test for their role in the development of HSCs from hESCs as well as the development/use of our more global miRNA screen. Regarding the larger miRNA screen, we now have a library that we think efficiently expresses miRNAs in hESCs, which was achieved by changing the promoter in our prior library. We are very optimistic about the new library; virus stocks are being made and will be tested in the near term. We also established and extensively characterized our mouse transplant model using human HSCs, and in the second half of the year performed many transplants of hESC-derived cells. To date we have not succeeded in identifying an engrafted HSC, but we have a number of ideas to try in year 3 to help us overcome this obstacle. We knew that this would be a huge hurdle, so while the lack of success in the transplants to date is not what we wished to have seen, it is not a major surprise. Nonetheless, we hope to overcome this problem with new ideas and studies that will be performed in funding year 3.

We continue to focus on understanding the role of microRNAs in the development of hematopoietic stem cells from human embryonic stem cells. Progress is slow, but present. We now understand that cells we expected to have hematopoietic stem cell potential appear to have a greater potential for becoming cells that line the blood vessels (endothelial cells). We are now trying to comb through our microRNA data to see if it can inform us as to how to push such cells more towards hematopoietic stem cells. We do know that these two cell types are very closely linked during mammalian development. We are also investigating new ways to develop cells from embryonic stem cells to help our chance of finding/developing the elusive hematopoietic stem cell.

Our focus in this proposal was to understand the role of miRNAs in the development of hematopoietic stem cells from human embryonic stem cells. Our work, and that of other over the past few years has demonstrated how difficult it is to generate hematopoietic stem cells from human embryonic stem cells. In fact, to date, nobody in the world has achieved that goal. However, along the way, we have come to realize that our system can help us identify and understand miRNAs involved in the critical developmental 'fork in the road' that leads to hematopoietic vs. cells that line blood vessels (endothelial cells). We worked in 2010-2011 to identify those miRNAs.

Because of the difficulty in generating hematopoietic stem cells from human embryonic stem cells, we stepped back and worked to established better reagents for understanding hematopoietic stem cell development from human embryonic stem cells. To that end, we have used homologous recombination to directly insert a reporter gene into the genome of a human embryonic stem cell. We have been characterizing this cell line, and we hope that it will help advance the field of hematopoietic stem cell development from human embryonic stem cells, both to serve our needs and those of other embryonic stem cell researchers. Using similar sophisticated techniques, we have removed (targeted) from human embryonic stem cells a critical gene required for blood development to provide yet another tool to help us understand this process. We hope that these new reagents will further our understanding of hematopoietic development.