published_or_final_version / Chinese Medicine / Master / Master of Philosophy

No description available.

published_or_final_version / Anatomy / Doctoral / Doctor of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / toc / Anatomy / Master / Master of Philosophy

Cytotoxic natural products represent a class of cancer drug candidates that have remained largely untapped as payloads in the antibody-drug conjugate (ADC) therapeutic modality. The epothilones, a class of exquisitely cytotoxic natural products and their synthetic analogs, are a prime example, and our work has focused on the development of epothilones as ADC payloads. Strategies toward this goal have included total synthetic efforts toward four structurally distinct epothilone analogs equipped with linker functionality and structural modifications designed to improve metabolic and chemical stability. In addition, we have pursued the synthesis of octreotide- epothilone conjugates, structures designed to target epothilones into cells that overexpress somatostatin receptors 2 and 5. Biological evaluation via in vitro cellular assays revealed one of our epothilone analogs as a promising epothilone-inspired ADC payload. Synthetic efforts toward these goals will be discussed.

Despite rapid advances in clinical oncology, acquired drug resistance still poses a significant threat to the long-term efficacy of current treatment regimens. Because most chemotherapy drugs aim to activate apoptosis in cancer cells, expansion of the pharmacopeia to include treatments targeting novel tumor cell death mechanisms is a promising anti-cancer strategy. Induction of ferroptosis, an iron-dependent form of regulated cell death, shows particular therapeutic potential as aggressive metastatic and drug-resistant cancer cell states have been demonstrated to possess an exquisite dependency on glutathione peroxidase 4 (GPX4), a key suppressor of the ferroptotic cell death pathway. However, current GPX4 inhibitors are limited by poor pharmacokinetic properties that preclude their clinical use. The development of novel drug-like GPX4 inhibitors would benefit from the discovery of new chemical scaffolds to both enhance our understanding of the structural basis of small molecule binding and inhibition as well as facilitate the rational design of future GPX4-targeted therapeutics. In this dissertation, we employed three high-throughput screening strategies to identify novel scaffolds of interest for GPX4 inhibitor development. First, a Lead-Optimized Compound (LOC) library was screened and we conducted further characterization and structure-activity relationship (SAR) studies on hit compound LOC880. Compared to the original hit, analogs QW-095 and QW-105 showed improved binding affinity and GPX4 inhibitory activity in vitro and also induced lipid peroxidation in cells suggestive of ferroptotic death. Further enhancement of the potency and ferroptosis specificity of this scaffold is still needed, but the potentially noncovalent and allosteric mechanism of action presents a novel approach for targeting GPX4. Second, we conducted extensive SAR analyses on another promising hit from the LOC library screen, LOC1886, which led to the identification of the lead compound QW-314. This analog showed significantly improved potency and ferroptosis specificity in multiple cancer cell contexts, including a drug-tolerant persister cell model of minimal residual disease. Characteristic markers of GPX4 inhibition and ferroptosis are also observed in cells treated with QW-314, including GPX4 protein degradation and induction of lipid peroxidation, and QW-314 exhibited excellent selectivity for GPX4 over another glutathione peroxidase family selenoprotein GPX1 in an in vitro assay using cell lysates. Moreover, we determined a baseline of pharmacokinetic measures including aqueous solubility and metabolic stability in human and mouse liver microsomes for further medicinal chemistry optimization. Lastly, we screened a DNA-encoded library (DEL) and an Enamine Diversity library, identifying 10 additional chemical starting points for future investigation.

Mounting evidence implicates persister cancer cells as the key element of minimal residual disease (MRD) from which cancer relapse occurs. The observation that persister cells are differentially and specifically sensitive to ferroptosis, a unique form of metabolically-linked cell death, presents a critical weak point through which identification and targeting of persister cells in MRD may become possible. To identify biomarkers for targetable cells, the drivers of ferroptosis sensitivity in persister cells must be identified. Using three chemotherapeutics and cell lines, we derived persister models across diverse tissues of origin and found that: 1) activating transcription factor 4 (ATF4), previously demonstrated as central to lung cancer persister state formation, is differentially expressed in prostate and fibrosarcoma persisters vs parentals, 2) proteins key to ferroptosis are underexpressed in persisters, and revert expression upon persister to parental reversion, 3) the lung persister lipidome is significantly rewired to drive ferroptosis and 4) upon persister to parental reversion and re-acquisition of ferroptosis resistance, the lipid signature also reverts back to a parental-like state, and 5) although ATF4 elimination in persisters does not revert ferroptosis sensitivity, mitochondrial elimination in persisters does abrogate ferroptotic sensitivity. Collectively, these findings reveal the mechanism of persister ferroptosis sensitivity across multiple cancer types, opening up the possibility of leveraging ferroptosis for elimination of minimal residual disease.

Indiana University-Purdue University Indianapolis (IUPUI) / Resveratrol has been shown to inhibit cell growth and induce apoptosis, as well as augment chemotherapeutics and irradiation in multiple cancer types. However, it is unknown if resveratrol is beneficial for treating drug-resistant cancer cells. To study the effects of resveratrol in triple negative breast cancer cells that are resistant to the common cancer drug, paclitaxel, a novel paclitaxel-resistant cell line was generated from the MDA-MB-231 breast cancer cell line. The resulting cell line, MDA-MB-231/PacR, exhibited a 12-fold increased resistance to paclitaxel but remained sensitive to resveratrol treatment. Resveratrol treatment reduced cell proliferation and colony formation and increased senescence and apoptosis in both the parental MDA-MB-231 and MDA-MB-231/PacR cell lines. Importantly, resveratrol treatment augments the effects of paclitaxel in both cell lines. The expression of the drug efflux transporter gene, MDR1, and the main metabolizing enzyme of paclitaxel gene, CYP2C8, was increased in the resistant cells. Moreover, pharmacological inhibition of the protein products of these genes, P-glycoprotein and CYP2C8, decreased paclitaxel resistance in the resistant but not in the parental cells, which suggests that the increase of these proteins are important contributors to the resistance of these cells. In conclusion, these studies imply that resveratrol, both alone and in combination with paclitaxel, may be useful in the treatment of paclitaxel-sensitive and paclitaxel-resistant triple negative breast cancers.