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- All Subjects: Glioblastoma
- Creators: Anderson, Karen
- Creators: Blattman, Joseph
Description
Glioblastoma multiforme (GBM) is an aggressive malignant brain tumor with a median prognosis of 14 months. Human hairless protein (HR) is a 130 kDa nuclear transcription factor that plays a critical role in skin and hair function but was found to be highly expressed in neural tissue as well. The expression of HR in GBM tumor cells is significantly decreased compared to the normal brain tissue and low levels of HR expression is associated with shortened patient survival. We have recently reported that HR is a DNA binding phosphoprotein, which binds to p53 protein and p53 responsive element (p53RE) in vitro and in intact cells. We hypothesized that HR can regulate p53 downstream target genes, and consequently affects cellular function and activity. To test the hypothesis, we overexpressed HR in normal human embryonic kidney HEK293 and GBM U87MG cell lines and characterized these cells by analyzing p53 target gene expression, viability, cell-cycle arrest, and apoptosis. The results revealed that the overexpressed HR not only regulates p53-mediated target gene expression, but also significantly inhibit cell viability, induced early apoptosis, and G2/M cell cycle arrest in U87MG cells, compared to mock groups. Translating the knowledge gained from this research on the connections between HR and GBM could aid in identifying novel therapies to circumvent GBM progression or improve clinical outcome.
ContributorsBrook, Lemlem Addis (Author) / Blattman, Joseph (Thesis director) / Hsieh, Jui-Cheng (Committee member) / Goldstein, Elliott (Committee member) / Harrington Bioengineering Program (Contributor) / School of Social Transformation (Contributor) / Barrett, The Honors College (Contributor)
Created2018-05
Description
The long-term survival of patients with glioblastoma multiforme is compromised by the tumor's proclivity for local invasion into the surrounding normal brain. These invasive cells escape surgery and display resistance to chemotherapeutic- and radiation-induced apoptosis. We have previously shown that tumor necrosis factor-like weak inducer of apoptosis (TWEAK), a member of the tumor necrosis factor superfamily, can stimulate glioma cell invasion and survival via binding to the fibroblast growth factor-inducible 14 (Fn14) receptor and subsequent activation of the Rac1/NF-kappaB pathway. In addition, we have reported previously that Fn14 is expressed at high levels in migrating glioma cells in vitro and invading glioma cells in vivo. Here we demonstrate that TWEAK can act as a chemotactic factor for glioma cells, a potential process to drive cell invasion into the surrounding brain tissue. Specifically, we detected a chemotactic migration of glioma cells to the concentration gradient of TWEAK. Since Src family kinases (SFK) have been implicated in chemotaxis, we next determined whether TWEAK:Fn14 engagement activated these cytoplasmic tyrosine kinases. Our data shows that TWEAK stimulation of glioma cells results in a rapid phosphorylation of the SFK member Lyn as determined by multiplex Luminex assay and verified by immunoprecipitation. Immunodepletion of Lyn by siRNA oligonucleotides suppressed the chemoattractive effect of TWEAK on glioma cells. We hypothesize that TWEAK secretion by cells present in the glioma microenvironment induce invasion of glioma cells into the brain parenchyma. Understanding the function and signaling of the TWEAK-Fn14 ligand-receptor system may lead to development of novel therapies to therapeutically target invasive glioma cells.
ContributorsJameson, Nathan Meade (Author) / Anderson, Karen (Thesis director) / Lake, Douglas (Committee member) / Tran, Nhan (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor)
Created2013-05
Description
The growing field of immunotherapy has generated numerous promising diseasetreatment platforms in recent years. By utilizing the innate capabilities of the immune system, these treatments have provided a unique, simplistic approach to targeting and eliminating cancer. Among these, the bispecific T cell engager (BiTEÒ) model has demonstrated potential as a treatment capable of bringing immune cells into contact with cancer cells of interest and initiating perforin/granzyme-mediated cell death of the tumor. While standard BiTE platforms rely on targeting a tumor-specific receptor via its complementary antibody, no such universal receptor has been reported for glioblastoma (GBM), the most common and aggressive primary brain tumor which boasts a median survival of only 15 months. In addition to its dismal prognosis, GBM deploys several immune-evasion tactics that further complicate treatment and make targeted therapy difficult. However, it has been reported that chlorotoxin, a 36-amino acid peptide found in the venom of Leiurus quinquestriatus, binds specifically to glioma cells while not binding healthy tissue in humans. This specificity positions chlorotoxin as a prime candidate to act as a GBM-targeting moiety as one half of an immunotherapeutic treatment platform resembling the BiTE design which I describe here. Named ACDClx∆15, this fusion protein tethers a truncated chlorotoxin molecule to the variable region of a monoclonal antibody targeted to CD3ε on both CD8+ and CD4+ T cells and is theorized to bring T cells into contact with GBM in order to stimulate an artificial immune response against the tumor. Here I describe the design and production of ACDClx∆15 and test its ability to bind and activate T lymphocytes against murine GBM in vitro. ACDClx∆15 was shown to bind both GBM and T cells without binding healthy cells in vitro but did not demonstrate the ability to activate T cells in the presence of GBM.
ContributorsSchaefer, Braeden Scott (Author) / Mor, Tsafrir (Thesis advisor) / Mason, Hugh (Committee member) / Blattman, Joseph (Committee member) / Arizona State University (Publisher)
Created2021
Description
Glioblastoma (GBM) is a highly invasive and deadly late stage tumor that develops from abnormal astrocytes in the brain. With few improvements in treatment over many decades, median patient survival is only 15 months and the 5-year survival rate hovers at 6%. Numerous challenges are encountered in the development of treatments for GBM. The blood-brain barrier (BBB) serves as a primary obstacle due to its innate ability to prevent unwanted molecules, such as most chemotherapeutics, from entering the brain tissue and reaching malignant cells. The GBM cells themselves serve as a second obstacle, having a high level of genetic and phenotypic heterogeneity. This characteristic improves the probability of a population of cells to have resistance to treatment, which ensures the survival of the tumor. Here, the development and testing of two different modes of therapy for treating GBM is described. These therapeutics were enhanced by pathogenic peptides known to improve entry into brain tissue or to bind GBM cells to overcome the BBB and/or tumor cell heterogeneity. The first therapeutic utilizes a small peptide, RVG-29, derived from the rabies virus glycoprotein to improve brain-specific delivery of nanoparticles encapsulated with a small molecule payload. RVG-29-targeted nanoparticles were observed to reach the brain of healthy mice in higher concentrations 2 hours following intravenous injection compared to control particles. However, targeted camptothecin-loaded nanoparticles were not capable of producing significant treatment benefits compared to non-targeted particles in an orthotopic mouse model of GBM. Peptide degradation following injection was shown to be a likely cause for reduced treatment benefit. The second therapeutic utilizes chlorotoxin, a non-toxic 36-amino acid peptide found in the venom of the deathstalker scorpion, expressed as a fusion to antibody fragments to enhance T cell recognition and killing of GBM. This candidate biologic, known as anti-CD3/chlorotoxin (ACDClx) is expressed as an insoluble protein in Nicotiana benthamiana and Escherichia coli and must be purified in denaturing and reducing conditions prior to being refolded. ACDClx was shown to selectively activate T cells only in the presence of GBM cells, providing evidence that further preclinical development of ACDClx as a GBM immunotherapy is warranted.
ContributorsCook, Rebecca Leanne (Author) / Blattman, Joseph N (Thesis advisor) / Sirianni, Rachael W. (Thesis advisor) / Mor, Tsafrir (Committee member) / Anderson, Karen (Committee member) / Arizona State University (Publisher)
Created2019