Theses and Dissertations
This collection includes both ASU Theses and Dissertations, submitted by graduate students, and the Barrett, Honors College theses submitted by undergraduate students.
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Description
Human Immunodeficiency Virus type 1 (HIV-1) causes millions of deaths every year, but a protective vaccine remains elusive. A promising vaccine strategy is to use virus-like particles (VLPs) for HIV-1. To this end, HIV-1 VLPs were produced in Nicotiana benthamiana plants that were stably expressing the HIV-1 Gag protein and transiently expressing a truncated form of gp41. These VLPs were tested to determine their inherent adjuvant effects due to their production in plants in order to dissect the previously observed stimulating activity of these VLPs in a prime-boost vaccine approach. THP1 human monocytes were differentiated using PMA or IL-4 and GM-CSF to form macrophages and dendritic cells, respectively. These cells were treated with purified VLPs or control samples to determine the individual adjuvant effects of the plant, bacterial, and VLP components in the purified VLP samples. It was postulated that the PMA-differentiated THP1 cells were not induced to become macrophages due to the lack of CD11b+ cells in the sample and the lack of increased TNFα expression in response to LPS treatment. It was also determined that the VLPs have inherent adjuvant properties to dendritic cells due to bacterial and VLP components, but not due to plant components.
ContributorsDickey, Rebekah Marie (Author) / Mor, Tsafrir (Thesis director) / Blattman, Joseph (Committee member) / Meador, Lydia (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Vaccinia virus is a cytoplasmic, double-stranded DNA orthopoxvirus. Unlike mammalian cells, vaccinia virus produces double-stranded RNA (dsRNA) during its viral life cycle. The protein kinase R, PKR, is one of the principal host defense mechanisms against orthopoxvirus infection. PKR can bind double-stranded RNA and phosphorylate eukaryotic translation initiation factor, eIF2α, shutting down protein synthesis and halting the viral life cycle. To combat host defenses, vaccinia virus encodes E3, a potent inhibitor of the cellular anti-viral eIF2α kinase, PKR. The E3 protein contains a C-terminal dsRNA-binding motif that sequesters dsRNA and inhibits PKR activation. We demonstrate that E3 also interacts with PKR by co-immunoprecipitation. This interaction is independent of the presence of dsRNA and dsRNA-binding by E3, indicating that the interaction is not due to dsRNA-bridging.
PKR interaction mapped to a region within the dsRNA-binding domain of E3 and overlapped with sequences in the C-terminus of this domain that are necessary for binding to dsRNA. Point mutants of E3 were generated and screened for PKR inhibition and direct interaction. Analysis of these mutants demonstrates that dsRNA-binding but not PKR interaction plays a critical role in the broad host range of VACV. Nonetheless, full inhibition of PKR in cells in culture requires both dsRNA-binding and PKR interaction. Because E3 is highly conserved among orthopoxviruses, understanding the mechanisms that E3 uses to inhibit PKR can give insight into host range pathogenesis of dsRNA producing viruses.
PKR interaction mapped to a region within the dsRNA-binding domain of E3 and overlapped with sequences in the C-terminus of this domain that are necessary for binding to dsRNA. Point mutants of E3 were generated and screened for PKR inhibition and direct interaction. Analysis of these mutants demonstrates that dsRNA-binding but not PKR interaction plays a critical role in the broad host range of VACV. Nonetheless, full inhibition of PKR in cells in culture requires both dsRNA-binding and PKR interaction. Because E3 is highly conserved among orthopoxviruses, understanding the mechanisms that E3 uses to inhibit PKR can give insight into host range pathogenesis of dsRNA producing viruses.
ContributorsFoster, Clayton (Co-author) / Alattar, Hamed (Co-author) / Jacobs, Bertram (Thesis director) / Blattman, Joseph (Committee member) / McFadden, Grant (Committee member) / School of Life Sciences (Contributor) / W. P. Carey School of Business (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2017-05
Description
Since the Acquired Immune Deficiency Syndrome (AIDS) crisis began in the early 1980s, there has been a significant amount of stigma attached to the disease and the virus that causes it, Human Immunodeficiency Virus (HIV). At the time, HIV/AIDS was viewed as a death sentence. A large part of the stigma came from the fact that in the early days of the crisis, AIDS patients were predominantly part of the LGBTQ+ community. With the discovery of effective antiretroviral therapies, today HIV can be thought of as a preventable, yet manageable, chronic illness, although it remains a huge public health concern (About HIV/AIDS, 2018). While the virus is now rarely viewed as a death sentence, there is still considerable stigma that surrounds people living with HIV/AIDS (PLWHA). Research shows that the shows and movies people watch can affect their attitudes on a variety of issues, and HIV is no exception. Because HIV is such a big threat to public health, and because people often adopt views they see in media, analyzing the ways shows and movies portray PLWHA is an important aspect in understanding where stigma surrounding HIV/AIDS comes from. The writers behind today's HIV+ characters on television and in movies all seemingly made an effort to decrease stigma, but they went about it in different ways, and with varying amounts of success. A common method to dispel stigma was to use the entertainment-education method (Singhal & Rogers, 1999), which in these cases means characters had discussions about topics like safe sex, Pre-Exposure Prophylaxis (PrEP), and the importance of getting tested. A few shows showed serodiscordant couples, which was also effective at fighting stigma. In contrast, by trying to be representative of PLWHA, some shows actually contributed to the stereotypes behind the stigma, or had characters be openly stigmatizing towards PLWHA. After analyzing what I found the shows and movies did well and what they did poorly, I'll analyze why it is important that shows maintained historical accuracy, and how doing so appeared to fight the stigma associated with HIV/AIDS. I will also evaluate what's missing \u2014 such as which high-risk groups are not represented. Ultimately, this thesis will argue that shows and movies made in the last 12 years all aimed to decrease stigma, through a variety of techniques.
ContributorsEvans, Celia Grace (Author) / Hurlbut, Ben (Thesis director) / Berkel, Cady (Committee member) / Blattman, Joseph (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2018-12
Description
The p53 gene functions as a tumor suppressor that inhibits proliferation, regulates apoptosis, DNA repair, and normal cell cycle arrest. Mutation of the p53 gene is linked to be prevalent in 50% of all human cancers. In this paper, we are exploring triple negative breast cancer and the effects of simvastatin on tumor growth and survival. Simvastatin is a drug that is primarily used to treat high cholesterol and heart disease. Simvastatin is unique because it is able to inhibit protein prenylation through regulation of the mevalonate pathway. This makes it a potential targeted drug for therapy against p53 mutant cancer. The mechanism behind this is hypothesized to be correlated to aberrant activation of the Ras pathway. The Ras subfamily functions to transcriptionally regulate cell growth and survival, and will therefore allow for a tumor to thrive if the pathway is continually and abnormally activated. The Ras protein has to be prenylated in order for activation of this pathway to occur, making statin drug treatment a viable option as a cancer treatment. This is because it acts as a regulator of the mevalonate pathway which is upstream of protein prenylation. It is thus vital to understand these pathways at both the gene and protein level in different p53 mutants to further understand if simvastatin is indeed a drug with anti-cancer properties and can be used to target cancers with p53 mutation. The goal of this project is to study the biochemistry behind the mutation of p53's sensitivity to statin. With this information we can create a possible signature for those who could benefit from Simvastatin drug treatment as a possible targeted treatment for p53 mutant cancers.
ContributorsGrewal, Harneet (Co-author) / Loo, Yi Jia Valerie (Co-author) / Anderson, Karen (Thesis director) / Blattman, Joseph (Committee member) / Ferdosi, Shayesteh (Committee member) / Department of Psychology (Contributor) / School of Life Sciences (Contributor) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2016-12
Description
Adaptive therapy utilizes competitive interactions between resistant and sensitive cells by keeping some sensitive cells to control tumor burden with the aim of increasing overall survival and time to progression. The use of adaptive therapy to treat breast cancer, ovarian cancer, and pancreatic cancer in preclinical models has shown significant results in controlling tumor growth. The purpose of this thesis is to draft a protocol to study adaptive therapy in a preclinical model of breast cancer on MCF7, estrogen receptor-positive, cells that have evolved resistance to fulvestrant and palbociclib (MCF7 R). In this study, we used two protocols: drug dose adjustment and intermittent therapy. The MCF7 R cell lines were injected into the mammary fat pads of 11-month-old NOD/SCID gamma (NSG) mice (18 mice) which were then treated with gemcitabine.<br/>The results of this experiment did not provide complete information because of the short-term treatments. In addition, we saw an increase in the tumor size of a few of the treated mice, which could be due to the metabolism of the drug at that age, or because of the difference in injection times. Therefore, these adaptive therapy protocols on hormone-refractory breast cancer cell lines will be repeated on young, 6-week old mice by injecting the cell lines at the same time for all mice, which helps the results to be more consistent and accurate.
ContributorsConti, Aviona (Author) / Maley, Carlo (Thesis director) / Blattman, Joseph (Committee member) / Seyedi, Sareh (Committee member) / School of Life Sciences (Contributor, Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
T cells, a component of the adaptive immune system, play an instrumental role in directing immune responses and direct cell killing in response to pathogens and cancers. T cells recognize and signal through the T cell receptor, a protein heterodimer on the surface of T cells. The T cell receptor is a highly variable structure formed via somatic recombination; the structure recognizes peptides presented on the surface of nucleated cells by major histocompatibility complex proteins in a specific receptor-restricted, peptide-restricted manner. This balance between T cell diversity and T cell specificity stands as a barrier to efficacious development of articificial T cell receptors capable of clearing disease. T cell receptors may be tailored to produce pathogen- or cancer-specific immune responses from autologous T cell populations. This necessitates a pipeline for amplification, cloning, and expression of antigen-specific T cell receptors. This study aims to utilize influenza-specific T cell receptor chains from healthy donor T cells to test a model for T cell receptor cloning and expression. This study utilizes Gateway recombination for high-throughput cloning into mammalian expression vectors. This study has successfully amplified and cloned T cell receptor chains from a population of influenza-specific T cells from donor cell transcripts into mammalian cell expression vectors. Additionally, CD8, a coreceptor for the T cell receptor complex, was successfully cloned and inserted into a vector for expression in mammalian cells. Sanger sequencing has confirmed sequences for influenza-specific T cell receptor chains and the CD8 chain. Future application of this project includes expression in mammalian non-T cells to test for efficacy of expression and, ultimately, expression in cytotoxic cells to create lymphocytes capable of antigen-specific recognition and cytolytic killing of cells of interest.
ContributorsVale, Nolan Richard (Author) / Anderson, Karen (Thesis director) / Blattman, Joseph (Committee member) / Department of Psychology (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
Novel biological strategies for cancer therapy have recently been able to generate antitumor effects in the clinic. Of these new advancements, oncolytic virotherapy seems to be a promising strategy through a dual mechanism of oncolysis and immunogenicity of the host to the target cells. Myxoma virus (MYXV) is an oncolytic poxvirus that has a natural tropism for European rabbits, being nonpathogenic in humans and all other known vertebrates. MYXV is able to infect cancer cells which, due to mutations, have defects in many signaling pathways, notably pathways involved in antiviral responses. While MYXV alone elicits lysis of cancer cells, recombinant techniques allow for the implementation of transgenes, which have the potential of ‘arming’ the virus to enhance its potential as an oncolytic virus. The implementation of certain transgenes allow for the promotion of robust anti-tumor immune responses. To investigate the potential of immune-inducing transgenes in MYXV, in vitro experiments were performed with several armed recombinant MYXVs as well as unarmed wild-type and rabbit-attenuated MYXV. As recent studies have shown the ability of MYXV to uniquely target malignant human hematopoietic stem cells, the potential of oncolytic MYXV armed with immune-inducing transgenes was investigated through in vitro killing analysis using human acute myeloid leukemia (AML) and multiple myeloma (MM) cell lines. Furthermore, in vitro experiments were also performed using primary bone marrow (BM) cells obtained from human patients diagnosed with MM. In this study, armed MYXV-infected human AML and MM cells resulted in increased cell death relative to unarmed MYXV-infected cells, suggesting enhanced killing via induced mechanisms of cell death from the immune-inducing transgenes. Furthermore, increased killing of primary BM cells with multiple myeloma was seen in armed MYXV-infected primary cells relative to unarmed MYXV-infected primary cells.
ContributorsMamola, Joseph (Author) / McFadden, Grant (Thesis director) / Jacobs, Bertram (Committee member) / Blattman, Joseph (Committee member) / School of Life Sciences (Contributor, Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
ContributorsLuca, Michael (Author) / Yan, Hao (Thesis director) / Stephanopoulos, Nicholas (Committee member) / Blattman, Joseph (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor) / School of International Letters and Cultures (Contributor) / School of Molecular Sciences (Contributor)
Created2024-05
Description
Cell immunotherapies have revolutionized clinical oncology. While CAR T cell therapy has been very effective in clinical studies, off-target immune toxicity limits eligible patients. Thus, NK cells have been approached with the same therapy design since NK cells have a more favorable safety profile. Therefore, the purpose of this research project is to explore DNA nanotech-based NK cell engagers (NKCEs) that force an immunological synapse between the NK cell and the cancer cell, leading to cancer death. DNA tetrabody (TB) and DNA tetrahedron (TDN) are fabricated and armed with HER2 affibody for tight adhesion to HER2+ cancer cell lines like SKBR3. Overall, relationship between TB-NK treatment and cancer cell apoptosis is still unclear. TB-NK treatment induces an apoptotic profile similar to PMA/IO stimulation. Pilot cell assay needs to be replicated with additional controls and a shortened treatment window. For DNA TDN fabrication, HER2 affibody polishing with Ni-NTA affinity chromatography achieves high purity with 20% to 100% high-imidazole elution gradient. ssDNA-HER2 affibody conjugation is optimal when ssDNA is treated with 40-fold excess sulfo-SMCC for 4 hours. In conclusion, the manufacturing of DNA-based NKCEs is rapid and streamlined, which gives these NKCEs the potential to become a ready to use immunotherapy.
ContributorsLuca, Michael (Author) / Yan, Hao (Thesis director) / Stephanopoulos, Nicholas (Committee member) / Blattman, Joseph (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor) / School of International Letters and Cultures (Contributor) / School of Molecular Sciences (Contributor)
Created2024-05
ContributorsLuca, Michael (Author) / Yan, Hao (Thesis director) / Stephanopoulos, Nicholas (Committee member) / Blattman, Joseph (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor) / School of International Letters and Cultures (Contributor) / School of Molecular Sciences (Contributor)
Created2024-05