Browsing by Subject "Androgen Receptor"

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Detection of AR gene rearrangements and regulation of AR variant-driven prostate cancer models
    (2022-07) Daniel, Mark
    Prostate cancer is one of the leading causes of death for men in the United States. The androgen receptor (AR) plays a central role in normal prostate development and homeostasis, but prostate cancer cells are also dependent on AR signaling for growth and survival. AR is the target for most prostate cancer therapies. Inhibiting this signaling axis is associated with progression to the lethal form of the disease, castration-resistant prostate cancer (CRPC). The development of CRPC is usually due to a variety of AR reactivation mechanisms including AR amplification, AR gene rearrangements, AR ligand binding domain mutations, altered expression of AR coregulatory factors, and the emergence of AR variants (AR-V). These resistance mechanisms allow AR to function in the presence of AR inhibitors, which creates challenges for development of prostate cancer therapies. The studies performed in this dissertation were aimed at expanding our ability to detect AR gene rearrangements that encode AR-Vs and understanding coregulatory proteins that regulate AR-V-driven prostate cancer models. In this thesis, we review AR-Vs, alterations in AR, and alterations in the broader AR pathway examined in the context of primary prostate cancer tissue, metastatic CRPC tissue, circulating tumor cells (CTCs), and circulating cell-free tumor DNA (ctDNA). Additionally, we profile a technique we developed for detection of AR gene rearrangements from CTC DNA. Using this approach, we analyzed matched CTC DNA and ctDNA from a cohort of ten CRPC patients and found that genomic alterations in CTC DNA and ctDNA were discordant. This proves the feasibility and utility of analyzing genomic alterations from a single blood sample. Finally, we characterized the role of TLE3 in AR-V-driven prostate cancer models. We found that TLE3, a transcriptional co-repressor, supports anchorage- independent growth and restricts migration in prostate cancer cell lines that express AR-Vs. Mechanistically, we found that TLE3 restricts cell migration by negatively regulating mTOR. This study enhances our understanding of the cross connectivity of AR, Wnt, and mTOR signaling that occurs in prostate cancer.
  • Thumbnail Image
    Item
    Synthesis of Photoresponsive Nucleosides and Their Incorporation into Oligonucleotides: Targeting Androgen Receptor and NF-κB Transcription Factors
    (2016-02) Struntz, Nicholas
    The sequencing of the human genome suggests that transcription factors (TFs) make up one of the largest classes of human proteins, revealing that there are over 2000 genes that code for transcription factors. The pivotal roles of TFs in cell biology become quite apparent when one or more of these regulatory mechanisms becomes mutated or altered. For example, the androgen receptor (AR) transcription factor plays a pivotal role in prostate carcinogenesis and progression. Additionally, the inflammatory response of the NF-κB transcription factor proteins results in the transcription of many genes, which play pivotal roles in carcinogenesis. There are several approaches to modulate and study transcription factor activity and biochemistry. Utilizing cis element DNA decoys to sequester TFs is one approach to directly modulate transcription factors. Introducing these synthetic double-stranded DNA decoys containing TF binding sites into cells effectively sequesters TFs and inhibits their target gene expression. Over the past couple of decades, numerous reports have validated utilizing this approach. For example, a phosphorthioate STAT3 DNA decoy has entered the “first-in-human” Phase 0 clinical trials for the treatment of head and neck squamous cancer. STAT3 expression and cell viability was reduced in the head and neck cancers injected with the decoy compared to the saline control. Combining the spatial and temporal resolution of caging technology with the DNA decoy strategy for the inhibition of transcription factor activity can yield an approach for the very precise ability to photochemically regulate gene expression, which has potential as a therapeutic agent and tool for probing biological pathways. This thesis will focus on efforts to develop several novel DNA-based and small molecule-based probes to investigate the biochemistry of TFs and their signaling pathways. Chapter 2 discusses the synthesis and characterization of caged DNA decoys that target the Androgen Receptor (AR). Caged DNA decoys successfully captured AR in LNCaP lysate when irradiated with light. Chapter 3 introduces the complement to caging technology, which is catch and release DNA decoys (CRDDs). CRDDs capture transcription factors, by binding and sequestering them, and then a pulse of light photochemically destroys the CRDD, permitting release of the TF. Several 7-nitroindole (7-NI, 1.47) nucleobase mimics were incorporated into NF-κB-directed DNA decoys, which still allowed the capture of the p50-p65 NF-κB proteins. Irradiation with 350 nm light drives the release of the p50-p65 NF-κB. The capture and photochemical release of an endogenous transcription factor is demonstrated for the first time. Chapter 4 continues the work of Chapter 3 by developing second-generation nucleobase mimics for use in CRDDs. Addition of molecular recognition properties on a photo-responsive monomer is hypothesized to increase binding affinity to capture endogenous TFs. 8-Nitroguanosine contains this added molecular recognition, is more stable within duplex DNA, and also displayed similar photochemical depurination properties. Chapter 5 outlines work developing photoswitchable nucleobases that transpose their hybridization properties upon photolysis. Chapter 6 highlights work to determine the mechanistic NF-κB inhibitory properties of several Cryptocaryone analogues, which were found to inhibit the NF-κB translocation to the nucleus. Appendix A focuses on the characterization of the enantioselectivity of guanosine monophosphate synthetase (GMPS), a crucial enzyme in nucleotide biosynthesis.
  • Thumbnail Image
    Item
    Therapeutic Targeting of Intrinsically Disordered Androgen Recptor Functional Domains in Prostate Cancer
    (2015-04) Brand, Lucas
    Prostate cancer (PCa) is a leading cause of morbidity and mortality in the United States, and contributes to a significant healthcare burden due to an overall lack of curative interventions for advanced-stage disease. Because PCa is largely insensitive to cytotoxic chemotherapy, the androgen receptor (AR) has long been the primary therapeutic target for the clinical management of locally advanced and metastatic PCa. Problematically, targeting AR signaling via androgen deprivation or treatment with AR antagonists is associated with progression to lethal, castration-resistant prostate cancer (CRPC) via a variety of molecular mechanisms that alter AR expression and function. However, CRPC is marked by a continued reliance on AR expression and activity. Thus, new modes of intervention with ability to durably repress AR activity in advanced CRPC are an unmet clinical need. In Chapter 1, we review the problem of castration resistance through a new paradigm of genetic rearrangements that produce truncated AR variants (ARV), which confer resistance to all current forms of AR-based PCa therapy. In Chapter 2, we discuss a novel AR inhibitor that directly targets the AR NH2-terminal transcriptional activation domain (NTD), but with significant off-target effects due to the lack of specificity for the intrinsically disordered NTD. In Chapter 3, we characterize the differences in NTD utilization between full length AR and ARV. Finally, in Chapter 4, we discuss a brief history of AR targeting in PCa, and offer a perspective on how future translational studies can approach the problem of intrinsic disorder in the NTD to develop new interventions with more durable and lasting mechanisms of action.