Human Marrow Stromal Mesenchymal Progenitor Cell Lines from Patients with Distinct Hematopoietic Diseases
Tools and Technologies I
Leukemia represents a variety of blood cancers that originate in bone marrow where blood cells are produced. Bone marrow from leukemia patients is widely populated with leukemia cells. A pathologist makes a diagnosis of leukemia by looking at blood and marrow films. Current successful treatments focus on eliminating the leukemic blood cells, and in some cases are followed by bone marrow transplantation. Bone marrow contains many different types of stem cells, which are regenerative cells. Bone marrow continually produces new blood cells—red cells, white cells, and platelets—throughout a person’s life. Supporting cells in the bone marrow have been known for over a century; these cells are called stromal cells (as the word “stroma”, in Latin, means mattress). Stromal cells have recently proven to be connective tissue or mesenchymal stem cells. Like all stem cells, the numbers of these cells are limited in bone marrow, but still they have the potential to populate the colonies of cells found throughout the body. Stromal cells thus constitute the principal components of various microenvironments—not only of marrow but also of breast, ovary, lymph nodes, etc. Scientists have studied and developed well-established cell growth systems for these easily reproduced cells to determine the function of both healthy and diseased cells and ultimately to be able to modify or destroy diseased cells. By studying not just cancer cells (i.e., the seeds) but also their microenvironments (i.e., the soil in which cancer “seeds” germinate, live and thrive) and by establishing new well-characterized stromal cell lines, scientists expect to identify the switches that turn on/off various kinds of diseases of the blood and other organs. In this application, the investigator proposes to establish unique stromal cell lines from a spectrum of blood disorders. The research team proposes to grow, expand, and isolate marrow stromal cells from patients having various types of leukemia, multiple myeloma, myelodysplastic syndromes, and aplastic anemia, and then immortalize them using state-of-the-art technologies. Once established and characterized, these immortalized cell lines will be made widely available to the scientific community. This research will enable scientists to conduct their investigations using standardized cells, thus advancing the search for causes and treatments of devastating diseases. Because of the potential of stromal cells to develop into a variety of connective tissues, like muscle, bone, fat and even nerve cells, the generation of these cell lines offers virtually unlimited practical research opportunities to investigators in many different disciplines in California and throughout the world.
Statement of Benefit to California:
Bone marrow stromal mesenchymal stem cells are the chief component of the marrow microenvironment, which controls the production of normal blood cells as well as promotes the development of leukemia, a deadly disease that represents a variety of blood cancers originating in the marrow. Like all other cancers, leukemia is a purposeless, progressive proliferation of blood cells, growing out of control and ultimately killing the patient if untreated, and in some unfortunate cases even if treated. According to the Leukemia & Lymphoma Society’s “Facts 2007-2008”, new cases of leukemia, Hodgkin and non-Hodgkin lymphoma, and myeloma will account for nearly 9.4 percent of the 1,444,920 new cancer cases diagnosed in the US this year. Every five minutes someone in the US is diagnosed with a blood cancer, and every 10 minutes someone dies from this terrible disease. Furthermore, according to Cancer Statistics 2007, there were an estimated 4,610 new cases of leukemia and an estimated 2,150 deaths from leukemia in California, as compared to 44,240 new cases of leukemia and 21,790 deaths from leukemia in the US in 2007. The broad objectives of our proposed studies thus are to understand the role of the marrow microenvironment in leukemia development and conversely the deleterious impact of leukemia cells on stromal cells and their ability to support normal blood cell production, which is almost always compromised in leukemia patients. Because marrow stromal cells by nature represent adult mesenchymal stem or progenitor cells (MPCs) with multi-lineage differentiation capacity, they offer promising potential for regenerative medicine. One of the largest barriers to advancing stem cell biology, including studies of MPCs and ultimately their application to therapy, is the establishment of well-characterized, immortalized cell lines to promote investigational standardization. Moreover, MPCs are not merely multi-potential but they are also multi-functional in the physiological sense, a fact not adequately appreciated. Functional properties of stromal cells include the following. a) MPCs are crucial to normal blood cell production. b) MPCs likely influence the genesis and progression of chronic blood diseases. c) MPCs’ role in leukemia is viewed as part of an emerging paradigm by which neighboring non-malignant cells regulate growth and development of malignant cells. d) MPCs have anti-proliferative and immunomodulatory properties. Still, little is known about the molecular mechanisms governing the critical biological roles of MPCs. MPC lines should, therefore, be valuable experimental tools. In addition to being useful for regenerative medicine purposes, they are critical for investigating various functional properties of stromal cells, including their roles in regulating leukemia cell growth and progression. This research will result in the development of novel diagnostic tools and treatment modalities for cancer patients in California and throughout the world.
The goal of this proposal is to generate multiple lines of stromal cells, particularly mesenchymal progenitor cells (MPC) from normal donors as well as from patients with several types of hematopoietic diseases. The Principal Investigator (PI) hypothesizes that certain aspects of these disorders could be attributed to underlying changes or defects in the microenvironment (stromal compartment) that would correspond to aberrant phenotypes in the resulting cell lines. Thus, comparison of normal and diseased cell lines might offer new insights into the mechanisms of pathology as well as provide new information about stromal cell biology. The research team proposes to grow, expand and isolate marrow stromal cells from patients having various types of leukemia, multiple myeloma, myelodysplastic syndromes and aplastic anemia, and then immortalized these cells using current molecular techniques. The reviewers felt that while the premise of this research has merit, the overall goals were too ambitious and risky. The success of the project relies on an unproven hypothesis. This uncertainty, combined with a lack of preliminary data, significantly diminished the reviewers’ enthusiasm. The scope was thought to be excessive, and the applicants gave little consideration to how the work would be focused or prioritized. Finally, given the ambitious nature of this proposal, it was difficult to evaluate the sufficiency of the research staff, two of which have yet to be hired. The impact of this proposal is uncertain, largely due to the unproven premise on which it is based. If it is true that some or all of the proposed diseases are due to stromal perturbations, and that these changes correspond to stable phenotypes in the immortalized cell lines, then the reviewers felt that the impact would be high. Renewable sources of the proposed cells would provide a valuable resource for studying the mechanisms of hematologic disease, the biological functions of MPC, and might eventually facilitate the development of new therapies. The reviewers questioned the feasibility of the project. They felt that the scope and magnitude of the proposal were excessive, especially given the lack of preliminary data to support the initial premise. The sheer quantity of cell lines and the number of assays that were proposed seemed unlikely to be accomplished in a two year time frame, although the reviewers praised the PI’s’ choice of criteria for characterizing MPC. The PI showed good judgment in his/her decision to isolate multiple lines from individual patients, but one reviewer indicated that there were insufficient numbers of patients for at least one of the diseases to draw statistically significant conclusions. Additional concerns were raised about the lack of focus and prioritization in the overall strategy. As an example, one reviewer noted that before immortalizing the cells, it would make sense to evaluate the three immortalization technologies proposed to determine which is most effective. As written in the proposal, it appears that all three would be used simultaneously. Another reviewer suggested that the PI should have initially focused on two diseases rather than four. Most reviewers agreed that the proposal would have been substantially improved if a pilot experiment was designed to provide proof of principle, and the ensuing strategies were prioritized and scaled down appropriately. Inclusion of studies in preclinical models could also facilitate the correlation of disease features with in vitro cellular phenotypes. The PI has a proven track record in the area of proteomics and single cell genomics and holds patents relevant to the proposed work. While the reviewers felt that the PI is qualified to conduct the proposed research, they were unclear as to how much of a team was actually in place. Overall, while the tools described in this proposal would be valuable, the research plan was deemed too risky and ambitious to warrant its pursuit.