Browsing by Subject "APOBEC3B"

Now showing 1 - 4 of 4
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    APOBEC Mutagenesis and DNA (Mis)Repair
    (2019-08) Serebrenik, Artur
    Cancer imposes a burden on the entire world. As of 2019, cancer accounts for 1 in every 6 deaths worldwide. This is a difficult disease to treat because of the immense heterogeneity created by mutations in tumors. These mutations allow tumors to evolve mechanism to avoid cell death, evade immune detection, develop drug resistance and metastasize. Considerable research has undergone to better understand the underlying molecular basis for the mutations observed in tumors. A better understanding of these processes is critical for development of effective therapies that kill cancer cells with minimal adverse effects. The advent of deep sequencing technologies has given researchers vast insights into the many cellular perturbations that give rise to the mutations observed in cancer. One of these molecular mechanisms is the cytosine deaminase activity attributed to the APOBEC3B (A3B) enzyme. Previous studies have implicated A3B as an endogenous source of mutation because it is overexpressed in cancerous tissue compared to normal tissue, it is active and generates detectable mutations in genomic DNA, and has clear associations with clinical outcomes. Here, we embarked on multiple approaches to identify therapeutically-relevant strategies of selectively killing A3B-expressing cancer cells. Overall, we are confident that a deeper understanding of the molecular mechanisms involved in processing A3B-catalyzed lesions will yield novel clinical opportunities.
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    APOBEC3B-driven mutagenesis in breast and other human cancers
    (2013-08) Burns, Michael Bradley
    Cancer is a disease that results from alteration of the cellular genome. The sources of these changes are multifarious, and in many cases unknown. This thesis focuses on the polynucleotide cytosine deaminase, APOBEC3B, as a newly discovered source of mutation in multiple human cancers. As a deaminase, APOBEC3B converts cytosines to uracils in single-stranded DNA. These uracil lesions are mutagenic as failure to properly repair them can result in a wide variety of mutation types. The initial discovery of this mutational phenomenon was described mechanistically using a variety of biochemical, genetic, and cellular assays in breast cancer cell lines. Follow-up work using publicly available next generation sequencing and clinical data indicated that this effect is operating in a large proportion of breast cancers. Expanded bioinformatic analysis that assessed APOBEC3B's potential impact was expanded to include 18 other human cancer types in addition to breast. This work shows that APOBEC3B is likely a significant contributor to the genetic heterogeneity in breast, head & neck, bladder, cervical, and lung (adeno- and squamous cell) carcinomas as evidenced by differential levels of expression in cancer tissues, increased mutation load, mutation clusters (kataegis), and an APOBEC3B mutation signature in tumors expressing high levels of this enzyme. Taken together, this thesis is a body of work describing a previously unappreciated source of genetic heterogeneity in several human cancers.
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    High-throughput Screening and Chemical Synthesis for the Discovery of APOBEC3 DNA Cytosine Deaminase Inhibitors
    (2015-09) Olson, Margaret
    APOBEC3 (APOBEC3A-APOBEC3H) enzymes catalyze single-stranded (ss)DNA cytosine-to-uracil (C-to-U) deamination as a function of innate immune defense against foreign DNA. Host cellular protection results from APOBEC3-catalyzed lethal mutagenesis of the offending genome. When misregulated, however, APOBEC3 enzymes have been demonstrated to drive the genetic evolution of numerous cancers and HIV-1. Specifically, sub-lethal levels of APOBEC3D/F/G/H-catalyzed mutation can enable HIV-1 escape from immune defense and antiretroviral therapies. Moreover, APOBEC3B over-expression in breast, bladder, cervical, lung, and head/neck cancers generates high levels of C-to-U mutation, which drives tumor formation, metastasis, and chemotherapeutic resistance. Thus, small molecule inhibition of APOBEC3-catalyzed deamination may provide a novel strategy for HIV-1 and cancer drug development. This thesis highlights efforts to discover small molecule inhibitors of the APOBEC3s through high-throughput screening (HTS) and chemical synthesis. Chapter 2 discusses an HTS of 168,192 compounds against APOBEC3B and APOBEC3G. In this effort, MN23 was discovered as a potent inhibitor of APOBEC3B (IC50 = 150 nM) and APOBEC3G (IC50 = 5.5 µM). Chapter 2 also reports a novel synthesis of MN23, and preliminary efforts to elucidate its mechanism of inhibition. Chapter 3 presents a class of covalent APOBEC3G-specific inhibitors based on a 1,2,4-triazole-3-thiol substructure. This compound class is predicted to inhibit APOBEC3G by covalently binding C321, forcing an inhibitory conformational change within the enzyme active site. Chapter 4 reports the discovery of a novel APOBEC3G inhibitory chemotype, which was discovered from the deconvolution of an impure HTS hit. In this effort, we also identified a previously unreported Pan Assay Interference Scaffold (PAINS), and characterized the mechanism by which compounds of this class undergo oxidative decomposition. Finally, Chapter 5 describes how benzthiazolinone-based APOBEC3 inhibitors are being developed into probes of APOBEC3 structure and function. Brief descriptions of two unrelated projects performed concurrent with these studies are also detailed in the Appendices.
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    Regulation of APOBEC3B catalyzed mutation in ovarian cancer
    (2015-08) Leonard, Brandon
    Cancer is the second highest cause of death in the United States. A greater understanding of the underlying causes of this disease is critical to improve patient outcomes. For years, researchers have known that cancer is primarily a genetic disease, caused by mutations that can activate oncogenes and inactivate tumor suppressors. Several studies have also shown that UV radiation, smoking and certain defects in DNA repair cause some of the mutations that lead to cancer, but the sources of mutations found in many tumor types are yet to be explained. Here, we build upon our initial finding that APOBEC3B is a source of mutation in breast cancer by defining its role in ovarian cancer. Parallel analyses looking globally at mutation in cancer have shown that APOBEC3B also contributes to mutation in several other tumor types. Additional studies have elucidated a major signaling mechanism that regulates APOBEC3B expression in cancer. While many efforts have been made to directly inhibit APOBEC3B enzymatic activity, the advances described here have the potential to inform alternative therapeutic strategies aimed at transcriptionally downregulating APOBEC3B to slow tumor evolution and improve the durability of conventional anti-cancer drugs. Ultimately, a more comprehensive understanding of the basic biology of APOBEC3B catalyzed mutagenesis in cancer will translate to larger impacts in the clinical arena.