Characterisation of Cholesterolcontaining Microparticles in Autoimmunity

In recent years, it has become apparent that sub-micron scale remnants of cells, called microparticles (MPs), are present in human circulation. MPs contain DNA, RNA, protein and lipids (such as cholesterol). Increased numbers of MPs are observed in autoimmune conditions such as Systemic Lupus Erythematosus (SLE) and Rheumatoid Arthritis (RA). This work aims to address the deficiency in the literature concerning MP cholesterol (MPC) by establishing reference values for MPC in a representative healthy population group, and by investigating the potential of MP cholesterol as a biomarker in autoimmunity

Discovery and Mechanistic Studies of Novel Redox Modulators for Treatment of Pancreatic Cancer

Pancreatic cancer remains a devastating disease and conventional chemotherapy shows modest efficacy because of drug resistance and systemic toxicity. The reprogramming of energy metabolism and oxidative stress are two hallmarks of cancer, and redox modulators have been developed as an attractive approach to treat cancer. At low or moderate levels, reactive oxygen species (ROS) serve as signaling molecules to mediate cellular functions; while at high levels, ROS induce oxidation of lipids, proteins, and DNA, ultimately leading to cell death. In this dissertation project, I aimed to identify novel redox modulators and provide a preclinical characterization of their mechanisms of action (MOAs) in pancreatic cancer cells. Through lead optimization of a previously studied quinazolinedione-based redox modulator, we identified QD394 with significant cytotoxicity in pancreatic cancer cells. Bru-seq technique and clustering analysis revealed remarkably similar post-treatment transcriptomic profiles between QD394 and napabucasin. Both compounds inhibited STAT3 phosphorylation, induced DNA damage, increased cellular ROS, and decreased the GSH/GSSG ratio. Moreover, QD394 caused an iron- and ROS-dependent, GPX4-mediated cell death, suggesting ferroptosis as a major mechanism. QD394 also decreased the expression of mitochondrial proteins, including LRPPRC and PNPT1 involved in mitochondrial RNA catabolic processes. A derivative QD394-Me was synthesized with improved plasma stability and reduced toxicity in mice compared to QD394. These results demonstrate that QD394 and QD394-Me represent novel ROS-inducing drug-like compounds warranting further development for the treatment of pancreatic cancer. Mito-Chlor, a mitochondrial-targeting triphenylphosphonium derivative of the nitrogen mustard chlorambucil, was identified to inhibit transcription of the mitochondrial genome through Bru-seq analysis, which is similar to a new ROS inducer SQD1 featuring a styrylquinoline-5, 8-dione core. Both Mito-Chlor and SQD1 decreased the mRNA levels of mitochondrial genes. However, only Mito-Chlor reduced their protein expression, and interfered with mitochondria membrane potential and oxidative phosphorylation. Both compounds increased cellular and mitochondrial ROS and stimulated similar downstream signaling related to oxidative stress and AP-1 transcription factors. These results establish SQD1 and Mito-Chlor as novel mitochondrial transcription inhibitors and redox modulators that may be applied to study cancer cell death related to mitochondrial function and redox signaling. Finally, a medium-throughput phenotypic screen of 20,000 diverse drug-like compounds produced a quinolin-chlorobenzothioate, QCBT7, as a potent hit with submicromolar cytotoxicity in cancer cells. Its structure is similar to 8-quinolinethiol hydrochloride (8TQ), a proteasome inhibitor. Proteasome inhibitors have shown anticancer efficacy. As a more stable derivative of 8TQ, QCBT7 caused the accumulation of ubiquitylated proteins, indicating its proteasome inhibitory activity. Additionally, QCBT7 increased the expression of a set of genes (PFKFB4, CHOP, HMOX1, and SLC7A11) at both nascent RNA and protein levels, similar to the known proteasome inhibitors MG132 and ixazomib. We have also identified PFKFB4 as a potential biomarker of proteasome inhibitors that can be used to monitor treatment response. Together, this study discovers that QCBT7 induces proteasome inhibition, hypoxic response, endoplasmic reticulum stress, and glycolysis, leading to cell death. In summary, the work as a whole provides a detailed characterization of redox modulators and their effects on cell death, mitochondria, or proteasome activity. We also identify novel ROS-related genes and pathways that could be beneficial for pancreatic cancer therapeutics. This thesis contributes to the overall understanding of ROS signaling in pancreatic cancer and the validity of ROS-modulating therapies. This collective work provides the foundation to improve the redox modulators discovered for testing in vivo.PHDMedicinal ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/163190/1/shuaihu_1.pd

Propagating functional dependencies with conditions

The dependency propagation problem is to determine, given a view defined on data sources and a set of dependencies on the sources, whether another dependency is guaranteed to hold on the view. This paper investigates dependency propagation for recently proposed conditional functional dependencies (CFDs). The need for this study is evident in data integration, exchange and cleaning since dependencies on data sources often only hold conditionally on the view. We investigate dependency propagation for views defined in various fragments of relational algebra, CFDs as view dependencies, and for source dependencies given as either CFDs or traditional functional dependencies (FDs). (a) We establish lower and upper bounds, all matching , ranging from PTIME to undecidable. These not only provide the first results for CFD propagation, but also extend the classical work of FD propagation by giving new complexity bounds in the presence of finite domains. (b) We provide the first algorithm for computing a minimal cover of all CFDs propagated via SPC views; the algorithm has the same complexity as one of the most efficient algorithms for computing a cover of FDs propagated via a projection view, despite the increased expressive power of CFDs and SPC views. (c) We experimentally verify that the algorithm is efficient. </jats:p

CD51/CD61+ Endothelial Microparticles Decrease in Diabetes Patients with Hypertension

Backgrounds: Type 2 Diabetes Mellitus (T2DM) and hypertension are commonly co-occurred and both diseases are related to endothelial dysfunction. Endothelial microparticles (EMPs) are shed from endothelial cells and can be found in condition of endothelial dysfunction. This study aimed to evaluate the circulating endothelial MPs (CD51/CD61+) levels in T2DM patients with or without hypertension and the correlation between endothelial MPs and clinical parameters. Methods and Results: 20 healthy control, 16 T2DM patients without hypertension and 11 T2DM patients with hypertension were recruited. CD51/CD61+ EMPs from all subjects were analyzed by flow cytometry. We found that, in the group of T2DM patients with hypertension, the absolute median number of CD51/CD61+EMPs was significantly decreased, compared with that in the healthy control and T2DM without hypertension groups. Furthermore, we conducted receiver operating characteristics (ROC) analysis to examine the accuracy of CD51/CD61+EMPs in the discrimination&nbsp; between T2DM patients with hypertension and healthy control, showing the accuracy was 76.4%. In addition, we also found that the accuracy of CD51/CD61+EMPs was 83.5% in the discrimination between T2DM patients with hypertension or without hypertension Conclusion: These findings identify CD51/CD61+EMPs as a potential biomarker to monitor endothelial dysfunction in T2DM patients with hypertension