Pancreatic cancer is the fourth leading cause of cancer death in Canada. Significant advancements in chemotherapy for advanced pancreatic cancer have resulted in the need for a quantitative comparison between these treatments on a relative scale. Therefore, a systematic review and Bayesian network meta-analysis of randomized clinical trials was conducted using gemcitabine, the standard treatment, as the reference comparator. Based on results from the network meta-analysis, in which optimal treatments were identified and side effects of each treatment evaluated, an Internet-based patient decision aid was developed in order to present the benefits and risks of each therapy option: (1) Best supportive care (2) gemcitabine (3) FOLFIRINOX. The objective of the decision aid was to guide patients through the decision-making process based on their individual preferences and values. The decision aid was deemed to be acceptable and feasible based on results from a pilot study conducted at The Ottawa Hospital Cancer Centre.

With a five-year survival rate of less than 5%, pancreatic cancer is one of the deadliest cancers. The most common activating mutations in pancreatic cancer are found in the KRAS gene, causing a constitutively-active KRAS protein in approximately 90% of pancreatic ductal adenocarcinomas (PDAC). PDAC-derived cell lines that harbor oncogenic KRAS mutations can be divided into two classes, KRAS dependent (or addicted) cells and KRAS independent cells. Oncogene dependency (or addiction) is a phenomenon where tu-mors require sustained activity of a single aberrantly activated gene despite the accumulation of multiple oncogenic lesions. In the case of PDAC, the single aberrantly activated gene is KRAS. KRAS independent cells have acquired various other oncogenic lesions that confer alternative cell survival signaling pathways to bypass oncogenic KRAS dependency. The Spleen Tyrosine Kinase (Syk) is highly expressed in KRAS dependent cells, while KRAS independent cells have low Syk expression. This pattern suggests that in KRAS dependent cells, constitutively active KRAS and Syk play a role in stimulating pro-survival pathways. One of these pro-survival pathways is known as mTORC1, which causes increased anabolic processes like protein and lipid synthesis. Accordingly, mTORC1 causes suppression of catabolic processes like autophagy. The net effect is in-creased cellular growth and proliferation. However, mTORC1 inhibitors have limited clinical efficacy, and potential therapeutic targets upstream of mTORC1 have drawn interest. Syk is a non-receptor tyrosine kinase that is an upstream activator of the mTORC1 pathway in hematopoietic malignancies. Through Syk inhibition studies using the small molecule PRT062607 (SYKi), we demonstrated that Syk is also involved in activating the mTORC1 pathway in KRAS dependent PDAC cells. However, the mechanism by which Syk-mediated activation of mTORC1 occurs is currently unknown. Moreover, it is unclear whether SYK kinase activity is required for the activation of the mTORC1 pathway. To address this issue, we introduced a single nucleotide mutation in the kinase do-main of Syk to render it kinase-inactive and found that Syk requires its kinase function to activate mTORC1. Studies using Syki also revealed that mTORC1 activity was also inhibited in KRAS independent PDAC cells that lack significant Syk expression. Interestingly, substrate specificity studies indicate that Syki also binds to and inhibits structurally similar protein tyrosine kinases such as the SRC Family Kinases (SFKs). Therefore, we designed an experiment to look for Syk and SFK cooperativity in regards to mTORC1 activation in PDAC cells. Our results indicate that the SFKs, Yes1 and Src display the most significant cooperative effect with Syk in activating the mTORC1 pathway. Src and Yes1 may even be involved in the upstream activation of Syk. To establish the physiological significance of Syk signaling in pancreatic cancer, it is important to establish model organisms that could be used for future studies. Thus, we test-ed Syk expression and function in PDAC cell lines derived from genetically-engineered mouse models (GEMM), which develop pancreatic cancer via oncogenic mutations in KRAS and TP53. We found that Syk is indeed expressed in murine PDAC cell lines and that the use of Syki in the murine PDAC cell lines results in decreased mTORC1 activity. These results recapitulate those obtained in human KRAS dependent PDAC cell lines. In summary, our studies show that Syk is a key regulator of mTORC1 signaling in human and mouse-derived pancreatic cancer cells. Syk kinase activity is required for mTORC1 activation. Finally, SFKs cooperate with Syk to promote robust mTORC1 activation. The mechanisms of SFK and Syk cooperativity in mTORC1 pathway activation will require further investigation. Additionally, our findings provide a strong rationale to study the effects of Syk kinase inhibition in physiologically-relevant murine models of pancreatic cancer. / 2021-06-08T00:00:00Z

With new insights on mechanism, there is renewed interest in the use of pharmacological ascorbate (AscH-) in cancer therapy. The generation of H2O2 with AscH- acting as an electron donor to O2 is central to AscH- -induced cytotoxicity. We hypothesized that catalytic manganoporphyrins (MnPs) would increase the rate of oxidation of AscH- thereby increasing the flux of H2O2, resulting in increased cytotoxicity. We tested three different MnPs: MnTBAP, MnT2EPyP, and MnT4MPyP, which represent a range of physicochemical and thermodynamic properties. Of the MnPs tested, MnT4MPyP had the greatest effect on increasing the rate of oxidation of AscH-, as seen by the concentration of ascorbate radical [Asc*-] and rate of oxygen consumption. MnPs and AscH-, when combined at concentrations that had minimal effects alone, synergistically increased the cytotoxicity as seen by decreased clonogenic survival in human pancreatic cancer cell lines. Catalase, but not superoxide dismutase, reversed the cytotoxicity of AscH- and MnP, consistent with an H2O2-mediated mechanism. In addition, there was a marked increase in the steady-state concentration of ascorbate radical upon the addition of MnPs to whole blood ex vivo from mice infused with ascorbate as well as from patients treated with pharmacologic ascorbate. The combination of MnT4MPyP with ascorbate inhibited in vivo tumor growth. We conclude that MnPs can increase the rate of oxidation of AscH-, leading to an increased flux of H2O2 resulting in increased ascorbate-induced cytotoxicity

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abstract: Extracellular vesicles (EVs) are membranous particles that are abundantly secreted in the circulation system by most cells and can be found in most biological fluids. Among different EV subtypes, exosomes are small particles (30 – 150 nm) that are generated through the double invagination of the lipid bilayer membrane of cell. Therefore, they mirror the cell membrane proteins and contain proteins, RNAs, and DNAs that can represent the phenotypic state of their cell of origin, hence considered promising biomarker candidates. Importantly, in most pathological conditions, such as cancer and infection, diseased cells secrete more EVs and the disease associated exosomes have shown great potential to serve as biomarkers for early diagnosis, disease staging, and treatment monitoring. However, using EVs as diagnostic or prognostic tools in the clinic is hindered by the lack of a rapid, sensitive, purification-free technique for their isolation and characterization. Developing standardized assays that can translate the emerging academic EV biomarker discoveries to clinically relevant procedures is a bottleneck that have slowed down advancements in medical research. Integrating widely known immunoassays with plasmonic sensors has shown the promise to detect minute amounts of antigen present in biological sample, based on changes of ambient optical refractive index, and achieve ultra-sensitivity. Plasmonic sensors take advantage of the enhanced interaction of electromagnetic radiations with electron clouds of plasmonic materials at the dielectric-metal interface in tunable wavelengths. / Dissertation/Thesis / Doctoral Dissertation Biomedical Engineering 2020

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22351号 / 医博第4592号 / 新制||医||1042(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 妹尾 浩, 教授 溝脇 尚志, 教授 戸井 雅和 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(人間健康科学) / 甲第22390号 / 人健博第76号 / 新制||人健||5(附属図書館) / 京都大学大学院医学研究科人間健康科学系専攻 / (主査)教授 椎名 毅, 教授 精山 明敏, 教授 富樫 かおり / 学位規則第4条第1項該当 / Doctor of Human Health Sciences / Kyoto University / DFAM

Patients with pancreatic cancer in metastasis rarely survive, thus the need for diagnostic tools for early stage detection. Current techniques such as ELISA and SPR are complex and expensive and cannot detect cancer in its early stages. Cancer Antigen 242 (CA 242) is a potential protein biomarker of pancreatic cancer with high sensitivity and specificity. This thesis presents a negative Dielectrophoresis (DEP) based method of detecting pancreatic cancer protein biomarker CA 242 in serum. A spectrum of concentration levels was generated with a cut off level 20 U/mL using a transduction mechanism with negative DEP spectroscopy, light scattering, and image processing. This was a fast and cost-effective method to diagnose early stage pancreatic cancer. This thesis also presents the design and simulation of an electrode modified to increase the electric field gradient with reduced heat generation and a concentration prediction model to predict concentrations from the generated spectrum of experiments.

Pancreatic cancer is a malignant disease with an unfavorable prognosis. Currently, the diagnosis of pancreatic cancer is based mainly on CT, MR and EUS imaging methods, because a reliable biomarker of pancreatic cancer is not available. It is considered a success in the current clinical practice if patients suitable for surgical resection can be selected, because in this group the overall survival is slightly better than in the group where surgery is not technically possible. In the first part, the presented dissertation focuses on testing of already established imaging diagnostic methods used in common practice, i.e. EUS and EUS FNA, as well as modern examination possibilities, such as contrast endosonography, and the comparison of the accuracy of the methods used. The dominant role of the EUS FNA was confirmed. In the next phase of the work, the methodological procedure of EUS FNA collection and the preprocessing of pancreatic tissue samples was tested with regard to the quantity and quality for epigenetic examination and further testing using the prognostic role of KRAS and miR-21. It has been suggested that cytological smears are the most suitable source of biological material for DNA and miRNA analysis, where non-tumor tissue contamination is low. Although the prognostic role of KRAS is...

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