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Characterization of Cancer Stem Cells in Multiple Myeloma

Funding Type: 
Early Translational I
Grant Number: 
Funds requested: 
$740 860
Funding Recommendations: 
Not recommended
Grant approved: 
Public Abstract: 
Cancer stem cells were first reported in acute myelogenous leukemia. Following this initial discovery, similar types of cells were identified in breast cancer and brain tumors. Recently, Matsui and his colleagues have reported the identification of clonogenic multiple myeloma (MM) cells and identified their progenitors as MM stem cells from severe combined immunodeficient (SCID) mice using a MM cell line. These cancer stem cells were shown to be a type of stem cell capable of forming tumors while showing properties of stem cells such as self-renewal and the ability to differentiate into multiple cell types. Until recently, most MM patients were treated with chemotherapy and glucocorticosteroids as initial therapy. Novel anti-myeloma agents such as thalidomide, lenalidomide, the proteasome inhibitor bortezomib, the doxorubicin derivative Doxil, and arsenic trioxide have proven to be potent inhibitors of myeloma cell growth in laboratory studies. They have been widely used alone and in combination therapies for myeloma patients, primarily for the treatment of relapsed or refractory disease but their efficacy have moved these newer agents to the frontline setting in many patients with promising results. Despite these improvements in anti-myeloma therapy, the disease remains incurable. Thus, development of new therapies that especially target the clone that maintains the tumor and gives rise to all of its subclones will be critical to eliminating the tumor population altogether. In order to characterize whether cancer stem cells can be identified in these patients, we have implanted fresh bone marrow biopsies from MM patients were into the superficial gluteal muscle of C.B-17 SCID mice. The tumors were excised from donor mice two months following implantation, and digested with protnase-E to produce a single cell suspension. These cells were analyzed using flow cytometry to identify specific cellular phenotypes within the tumor population. For accurate targeting MM cancer stem cell, we further isolate the tumor cells into several subpopulations by using anti-CD138, CD20, and CD133 antibodies with magnetic beads and then inject the each population into MM mouse model. Gene expression and phenotypes of each subpopulation cells will be analyzed by different biotechnologies. Through characterizing this population, we should be able to identify potential specific treatments that are capable of eliminating this critical tumor subclone. We will also determine the sensitivity of this stem cell population in vivo to treatment with a variety of agents. We will optimize combination therapies that can effectively eliminate this stem cell clone in MM. Importantly, development of specific therapies targeted at cancer stem cells in MM patients should produce not only a better survival but the ultimate goal of cure for these patients.
Statement of Benefit to California: 
Multiple myeloma (MM) is an incurable hematological malignancy that accounts for 10% death caused by blood cancer. MM is a malignancy of plasma cells with low proliferative activity in the bone marrow. The incurable disease has a median survival of approximately 5 years. Until recently, most patients were treated with chemotherapy and glucocorticosteroids as initial therapy. Novel anti-myeloma agents such as thalidomide, lenalidomide, the proteasome inhibitor bortezomib, and arsenic trioxide have proven to be potent inhibitors of myeloma cell growth in laboratory studies. Initially they were used alone but their efficacy was shown to be much improved when combined with each other or with chemotherapy. Despite these improvements in anti MM therapy, the disease remains incurable and nearly all patients eventually develop resistance to these therapies. New cases of MM are expected to be approximately 19,920 in 2008 in the United States. Most of patients are older than 60y. However, California is a big state with more population than other state and the MM patients number is continue increasing. Most of our clinical patients are from California although some patients from all of the United States. Scientists in California were the first to discover and isolate human leukemia and human breast cancer stem cells. Their efforts are now close to isolating stem cells for brain cancer, ovarian cancer, melanoma and bladder cancer. Scientists in our research laboratory has stared MM cancer research several year ago (see preliminary data). We hypothesize that the multiple myeloma (MM) cancer stem cells are a distinct population within the tumor clone. These myeloma “stem cells” are more likely to be drug resistant, and lead to more aggressive disease that does not respond to conventional therapies. The development of specific therapies targeted at MM cancer stem cells gives hope for not only improving survival but ultimately curing these diseases. In addition, the major clinical manifestations of MM are related to bone disease resulting from enhanced bone loss. When we treat the MM patient we have to treat both cancer and bone damage. Although MM accounts for only a small percentage of all cancer types, the costs associated with treating and managing it are among the highest. Recent developments in diagnosing, treating, and managing myeloma have led to novel treatment strategies. Immunomodulators, proteasome inhibitors, and bisphosphonates are improving response rates and preserving quality of life. However, these agents are not replacing older treatment modalities, but being used in addition to them. State will spend lots of money to do so. If we can cure MM by targeting cancer stem cell it’s not only MM patient gets quality life but also California will be benefit to reduce huge medical expense in the future.
Review Summary: 
Cancer stem cells are believed to exist in some tumors as a distinct subpopulation of cells causing relapse and metastasis by giving rise to new tumors. Multiple myeloma (MM), which is characterized by an aberrant accumulation of malignant plasma cells in the bone marrow (BM), is the target of this proposal. The principal investigator (PI) proposes to overcome a bottleneck for cell-based therapies by isolating and characterizing myeloma initiating cells (MICs) from the tumors of patients with MM. MICs have been detected in MM tumor cell lines established to grow in vitro, but they have not been identified in the tumors of patients. The ultimate goal of the proposal is to determine whether existing chemotherapies used to treat MM patients target MM MIC, and if not, to identify combinations of these chemotherapeutic agents that target these cells. The PI also suggests that his/her studies may lead to discovery of new molecular targets in MM MIC-enriched cell populations. The review panel was united in its agreement that MM poses a significant health problem and is currently incurable; therefore, this proposal addresses a critical area in need for research and development of a therapy. These myeloma “stem cells” are more likely to be drug resistant, and lead to a more aggressive disease state that does not respond to conventional therapies. Despite its focus on isolation and characterization of MICs, one reviewer believed most of the proposal was clearly focused on fundamental cancer stem cell biology, not early translation. Though the proposal itself was very straightforward, reviewers found it to be limited in scope and suffering from a number of substantial shortcomings. First, the applicant assumes that the three antibodies named would prove useful in separating tumorigenic from non-tumorigenic MM tumor cells; however, this assumption is not supported by data or previous studies. Furthermore, the applicant does not indicate whether she/he would use limiting dilution cell transplantation experiments to calculate the frequency of MICs in any of the subpopulations. Unless the latter analyses are carried out, it will not be possible to know which of the cellular fractions comprise the MIC subpopulation. Second, the applicant seemingly assumes that one of the six defined cell subpopulations would comprise a substantial MIC fraction, thus making it possible to characterize the presumed abundant MIC in one of the subpopulations. Compelling evidence that this would be the case is lacking, and hence, characterization of the cells in the MIC-enriched fraction(s) will be uninformative. Third, the characterization of the MIC in any of the MIC-enriched populations would yield minimal new information. Fourth, reviewers found the third specific aim to be confusing, as it was not clear whether the applicant would use existing MM xenograft models and also derive new models using fresh tumor samples. Reviewers were also confused by the rationale in which the PI argues that MM MIC may be resistant to existing therapies yet plans to test these very same drugs for their capacity to target MM MIC. In addition, reviewers were not convinced by preliminary data suggesting that complete selectivity for MIC could be achieved with the proposed studies. Technically, highly efficient and selective methods, required for the isolation of rare cell populations, are not proposed. Reviewers also felt that the proposal may be misdirected, as the identification of new compounds is not proposed and the PI already has tools in hand with existing chemo-resistant tumor lines to analyze and characterize MICs. Preliminary studies and data with these cell lines would have been extremely useful for the evaluation of this application and would help focus the proposal in pertinent directions. The applicant has appropriate qualifications to perform the research program described in the proposal, and the collaborations between team members are well documented in publications. There does not seem to be a great deal of collaborations with individuals outside of the immediate team, however. The research environment is adequate for the proposed research. In summary, the applicant presents a broad approach to search for cells from multiple myeloma patients with genetic or surface markers consistent with cancer stem cells. Despite attempting to address a critical bottleneck, the application’s proposed solution is narrow in scope and possesses critical flaws that undermine its feasibility.

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