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Browsing by Subject "Cancer Stem Cells"

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    Development and characterization of CD133 positive cancer stem cell targeted toxins for use in carcinoma therapy
    (2013-05) Waldron, Nate N.
    Cancer Stem cells (CSC) have been shown to play an important role in a number of carcinomas. Although representing a subpopulation of many cancers, CSC are extremely important because they are more drug resistant than the more differentiated cancer cells which make up the bulk of most solid tumors. High numbers of CSC is an indicator of poor clinical outcome and have been shown to be a cause in drug refractory relapse, which is the single most urgent problem in carcinoma therapy. CD133 is a cell surface receptor that has been identified as a CSC marker in breast, brain, colon, pancreatic, and recently in Head and Neck (HN) carcinoma. Our laboratory bioengineered a targeted toxin called dCD133KDEL consisting of the scFv portion of a novel anti-CD133 monoclonal antibody on the same molecule as truncated pseudomonas exotoxin. Binding of dCD133KDEL was demonstrated on a variety of carcinoma lines and we verified the ability of the anti-CD133 scFv to sort tumor initiating cells. Since enhanced tumor initiation is a hallmark of CSC, we demonstrated that dCD133KDEL was able to prevent tumor initiation. Importantly, even though CD133 was expressed only on a subpopulation of cells, dCD133KDELprevented cell proliferation in vitro and had powerful anti-cancer effects in vivo in xenograft mouse models of head and neck cancer. The therapeutic potential of dCD133KDEL was further investigated in xenograft models of human breast and ovarian cancer where it was effective when administered systemically as well. To further study therapeutic potential, we assessed the reactivity of this drug on normal human progenitor cells since CD133 is a known progenitor cell marker. dCD133KDEL did not kill normal human CD133+ stem cells at the same concentrations as it did for carcinoma cells, indicating a therapeutic window exists. Drug safety studies were performed in mice and the maximum tolerated dose of dCD133KDEL was established. Liver damage was shown to be the dose limiting toxicity. Since CD133 positive cells can develop from CD133 negative cell populations, a phenomenon known as stem cell plasticity, we developed a bispecific targeted toxin (dEpCAMCD133KDEL) capable of targeting both CD133 and the epithelial cell adhesion molecule, EpCAM, an overexpressed marker on most carcinomas. dEpCAMCD133KDEL potently inhibited cell proliferation of a number of carcinoma lines in vitro and was also effective at eliminating tumor spheroids, which have been shown to be enriched for CSCs. This bispecific agent was also effective at causing tumor regression in a model of HN cancer in vivo. Because of the preliminary effectiveness of dCD133KDEL and dEpCAMCD133KDEL in preclinical evaluation, these drugs warrant further development for possible use in carcinoma therapy. These two CSC targeted toxins could be extremely useful in situations where our current drugs are failing because of the progression of a critical drug resistant CSC population. Since, targeted toxins work synergistically with current chemotherapy, these two targeted toxins could be used as an adjunct to current therapy to target the CSC population specifically, while traditional chemotherapy and radiation can still be used to target the rapidly dividing bulk of the tumor. We believe that future success in cancer treatment must include approaches to target CSC as well as the majority of less differentiated cancer cells that comprise a tumor.
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    Molecular Mechanisms of Stemness and Invasion in Pancreatic Cancer
    (2016-05) Nomura, Alice
    Pancreatic cancer remains a cancer with the lowest survival rate and shortest median survival. It is predicted to accelerate to the 2nd cause of cancer related death in the next 15 years. Poor survival rates stem from late disease detection, aggressive tumor biology, and unsuccessful treatments. These dismal statistics highlight the urgency for increased knowledge of the biology of pancreatic cancer and the need for effective therapies. Within the past decade, cancer stem cells, a population within the tumor, have been exposed for their role in tumor initiation, progression, chemoresistance, and metastasis. These studies focus to examine the role of CD133 positive cancer stem cells in stemness and metastasis within pancreatic cancer. We demonstrated that CD133 positive pancreatic cancer cells, from human pancreatic cancer cell lines and a spontaneous murine model of pancreatic cancer, are capable of tumor initiation at very low cell numbers in both immunocompromised and immunocompetent mice, respectively. These cells exhibited chemoresistance properties by the upregulation of drug transporters, active drug efflux capabilities, and resistance to cell death upon treatment with conventional chemotherapies, such as gemcitabine and 5-fluorouracil. CD133 positive cancer stem cell population was, however, sensitive to a novel therapy- Minnelide™. In both in vitro and in vivo studies, CD133 positive cells responded to Minnelide™ treatment and underwent apoptosis in addition to the CD133 negative population. These investigations led to questions regarding the function of this surface marker. CD133 played no functional role in conferring the cancer stem cell phenotype to this population. Through overexpression of CD133 in a pancreatic cancer cell line with very low endogenous CD133 expression, we determined that CD133 expression influences both stemness and invasiveness. Cells overexpressing CD133 were capable of initiating tumors at low cell numbers, as compared to control cells, and exhibited an upregulation in pluripotency and developmental signaling gene expression. Additionally, tumors derived from CD133 overexpressing cells demonstrated a marked increase in metastasis to several distant sites. This was shown to occur through the activation of NF-kB signaling and induction of the epithelial-mesenchymal transition; resulting in increased cellular invasiveness. Further, CD133 expressing cells displayed increased expression and secretion of the cytokine, interleukin-1 beta. Inhibition of IL-1 signaling through various methods established a significant role for IL-1 in the induction of the epithelial-mesenchymal transition and cellular invasiveness. Significantly, interleukin-1 beta positively correlates with CD133 gene expression in pancreatic cancer cell lines of varying aggressiveness. In cell lines with high levels of CD133 positive populations, inhibition of IL-1 signaling demonstrated its critical role in epithelial-mesenchymal transition induction and invasiveness. This exhibited that IL-1 signaling functions within CD133 positive populations during the metastasis process. Finally, these studies demonstrated the pivotal role of NF-kB activation in the induction of the epithelial-mesenchymal transition, cellular invasion, and metastasis. Using triptolide and NF-kB signaling inhibitor treatment, as well as, NF-kB signaling pathway modulation via constitutively active or inactive plasmid expression, this signaling pathway was decisively confirmed to mediate invasion and metastasis in pancreatic cancer. Taken together, this work establishes the functional role of the cancer stem cell marker, CD133; IL-1 signaling; and NF-kB activation in pancreatic cancer stemness and metastasis.