Cancer Biology Commons^™

The Loss Of Atrx Creates Susceptibility To Kras-Mediated Pancreatic Damage, Claire C. Young

Electronic Thesis and Dissertation Repository

Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with a five-year survival rate of 8%. Oncogenic KRAS mutation is found in greater than 95% of PDAC cases, but additional mutations or injury are required for disease initiation and progression. Chromatin remodeling protein ATRX has been previously implicated in DNA repair, replication and maintaining genomic stability. I hypothesized that loss of ATRX could increase the susceptibility of pancreatic tissue to pancreatic injury or KRAS-mediated damage. In this study, combination of inducible ATRX loss in adult mice with recurrent pancreatic injury or oncogenic KRAS activation resulted in increased pancreatic damage, including fibrosis, …

Nuak1 Has Tumor Suppressive Activity In Epithelial Ovarian Cancer, Parima Saxena

Electronic Thesis and Dissertation Repository

NUAK1, a downstream substrate of the stress metabolism regulator Liver kinase B1 (LKB1), has been implicated as an oncogene and tumor suppressor, with distinct roles in cell cycle, senescence and cell adhesion. Epithelial ovarian cancer (EOC) spheroids remain dormant during intraperitoneal metastasis with reduced proliferation and metabolism to survive metabolically harsh conditions, possibly implicating a role of NUAK1 in EOC spheroid biology. I hypothesize that NUAK1 is regulated by LKB1 to promote dormancy in EOC. I demonstrate that NUAK1 expression and phosphorylation is regulated by LKB1 in EOC cell lines. NUAK1 is largely under-expressed in many established and new ascites-derived …

Regulation Of Smad Anchor Necessary For Receptor Activation (Sara) Protein Levels By The E3 Ubiquitin-Protein Ligase Smurf2 And Smad7, Sanghyun Lee

Electronic Thesis and Dissertation Repository

The TGFβ pathway, which regulates cell proliferation and differentiation, has also been shown to induce non-small cell lung cancer cell (NSCLC) migration and invasion. The TGFβ pathway is initiated through the binding of TGFβ to cell surface Ser/Thr kinase receptors. Activated receptors then phosphorylate intracellular signaling proteins, termed Smads, which translocate into the nucleus to regulate transcriptional responses. The protein Smad anchor for receptor activation (SARA) facilitates the phosphorylation of Smads and allows for efficient signal transduction. On the other hand, the inhibitory Smad, Smad7, recruits the E3 ubiquitin ligase, Smurf2, to catalyze the degradation of TGFβ receptors. Since the …

The Effects Of Acetylenic Tricyclic Bis-(Cyano Enone) On Cell Migration, Eddie Chan

Electronic Thesis and Dissertation Repository

Although cancer survival rates have significantly improved over the past few decades, the improvements are primarily due to early diagnosis and inhibiting cancer growth. Limited progress has been made in the treatment of cancer metastasis, which contributes to 90% of cancer related deaths, and therapeutic agents targeting the various aspects of metastasis are lacking. One potential approach is to utilize small pharmacological compounds to inhibit tumour cell motility, as a strategy against tumour cell migration, invasion, and metastasis. The acetylenic tricyclic bis-(cyano enone), TBE-31, has been shown to be a promising chemopreventative compound. However, its effects on cell migration are …

Mt1-Mmp Mediates The Migratory And Tumourigenic Potential Of Breast Cancer Cells Via Non-Proteolytic Mechanisms, Mario Cepeda

Electronic Thesis and Dissertation Repository

Membrane Type-1 Matrix Metalloproteinase (MT1-MMP) is a multifunctional protease that affects cell function via proteolytic and non-proteolytic mechanisms such as promoting degradation of the extracellular matrix (ECM) or augmentation of cell migration and viability, respectively. MT1-MMP has been implicated in metastatic progression ostensibly due to its ability to degrade ECM components and to allow migration of cells through the basement membrane. Despite in vitro studies demonstrating this principle, this knowledge has not translated into the use of MMP inhibitors (MMPi) that inhibit substrate catalysis as effective cancer therapeutics, or been corroborated by evidence of in vivo ECM degradation mediated by …

Discovery Of Novel Diagnostic Biomarkers On Prostate Tumor Microparticles For Discriminating Between Low And High Risk Prostate Cancer And Improving Prostate Cancer Screening, Sabine Brett

Electronic Thesis and Dissertation Repository

There are few protein-based biomarkers to accurately distinguish between patients with low risk prostate cancer from those with high risk disease in a non-invasive manner. Prostate specific antigen (PSA) is used for clinical follow-up of prostate cancer; however, it is not effective as a screening tool. As a result, many men with non-life threatening disease having to undergo unnecessary and painful biopsies. Therefore, there is a dire need for minimally invasive platforms for monitoring patients with clinically significant prostate cancer. Prostate cell microparticles (PCMPs) released by prostate epithelial cells into plasma are a potential source of biomarkers specific for prostate …

Regulation Of E2f1 In Keratinocytes During Uv-Damage And Differentiation, Randeep K. Singh

Electronic Thesis and Dissertation Repository

The E2F1 transcription factor regulates the expression of key genes involved in cell proliferation and differentiation to maintain skin homeostasis. The expression of E2F1 is tightly regulated during cell cycle progression and when cells are committed to differentiate, as well as in response to DNA damage. In keratinocytes, E2F1 protein and transcript levels increase following UV-induced DNA damage, whereas, in response to Ca2+-induced differentiation, both E2F1 protein and transcript levels decrease. In this thesis, I examined in detail the mechanism that modulates E2F1 stability following DNA damage and during keratinocyte differentiation. I show that E2F1 associates with hHR23 and together …

Characterization Of The Catalytic Ck2 Subunits With Substitutions At Residues Involved In Inhibitor Binding, Paul Desormeaux

Electronic Thesis and Dissertation Repository

CK2 is a constitutively active, ubiquitously expressed and pleiotropic serine/threonine protein kinase that is implicated in many cellular functions including tumorigenesis. CK2 has two catalytic subunits, CK2a and CK2a’, that carry out its function in the cell. Previous studies have indicated that inhibitor-refractory mutants have been effective in recovering residual CK2 activity, in the presence of inhibitors, when compared to wild type CK2. Based on these observations, inhibitor-refractory mutants were created for both CK2a and CK2a’ and tested with various concentrations with two CK2-specific inhibitors, CX-4945 and inhibitor VIII. The CK2a triple mutant (V66A/I174A/H160D) was tested in inducible U2OS Flp-In …

Inhibition Of Mt1-Mmp Proteolytic Function And Erk1/2 Signalling Influences Breast Cancer Cell Migration And Invasion Through Changes In Mmp-2 And Mmp-9 Expression, Caitlin L. Evered

Electronic Thesis and Dissertation Repository

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is a multifunctional protease that invokes changes extracellularly via cleavages of ECM substrates, and intracellularly through induction of cell signalling cascades, both to influence cell behaviour and motility. The effects of MT1-MMP activation, MMP catalytic activity, and ERK1/2 signalling were examined by chemically or genetically altering each parameter. Regardless of treatment, expression of the two gelatinases, MMP-2 and -9, were always altered in an inverse relationship. This work proposes a pathway through active MT1-MMP initiation of ERK1/2 phosphorylation and subsequent targeting the NF-κB transcription factor, to ultimately influence MMP-9 expression. Modulation of MT1-MMP activation, …

Characterization Of Numb As A Regulator Of Anaplastic Lymphoma Kinase, Ran Wei

Electronic Thesis and Dissertation Repository

Cellular events rely on protein-protein interactions that are often mediated by modular domains which recognize particular sequence motifs in binding partners. The NUMB protein is the first described cell fate determinant and multifaceted adaptor that is involved in a wide variety of cellular events. NUMB mainly mediates protein interactions via its modular PTB domain. Here we present a systematic investigation of the NUMB-PTB interactome by employing an integrative strategy combining both protein and peptide arrays. We profiled NUMB-PTB binding specificity and interacting proteins genome-wide. The receptor tyrosine kinases (RTKs) are found highly enriched in the interactome, raising the possibility that …

The Cytoplasmic Domain Of Membrane-Type 1 Matrix Metalloproteinase Is Required For Its Survival-Promoting, But Not Its Migration-Promoting Function In Mcf-7 Breast Cancer Cells, Jacob Jh Pelling

Electronic Thesis and Dissertation Repository

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is a multifunctional protease that degrades proteins during cell migration, and influences cell survival. Both the protein localization and signal transduction capabilities of MT1-MMP depend on its cytoplasmic domain (CD), indicative of a diverse regulatory function. The effects of CD mutations on cell migration and survival were examined by ectopically expressing MT1-MMP variants in MCF-7 cells. CD alteration by substitution or deletion did not abolish the migration-promoting effects of MT1-MMP, but did decrease cell survival and increase apoptosis. Expression of CD-altered MT1-MMP resulted in a protrusive cell morphology in 3D culture that was lost upon …

Oncolytic Virus Therapy For The Treatment Of Metastatic Ovarian Cancer, Jessica Tong

Electronic Thesis and Dissertation Repository

The management of patients with epithelial ovarian cancer (EOC) faces two major challenges which standard treatments fail to effectively address: 1) Diffuse metastasis as a consequence of late stage diagnosis and 2) intra-tumoral heterogeneity, which fuels tumor evolution and drives the acquisition of chemotherapeutic resistance. In this thesis, we tested new therapeutic strategies using a 3-dimensional in vitro spheroid culture model that mimics key steps of epithelial ovarian cancer metastasis; and another model that mimics both temporal and cellular heterogeneity by establishing multiple cell lines from a single patient over the course of disease progression. Using these models, we investigated …

Exploring The Regulation And Function Of Epithelial-Mesenchymal Plasticity In Ovarian Cancer Spheroids, Samah Rafehi

Electronic Thesis and Dissertation Repository

Epithelial-mesenchymal transition (EMT) serves as a key mechanism driving tumour cell migration, invasion, and metastasis in many carcinomas. Transforming growth factor-beta (TGFβ) signalling is implicated in several steps during cancer pathogenesis and acts as a classical inducer of EMT. Since epithelial ovarian cancer (EOC) cells have the potential to switch between epithelial and mesenchymal states during metastasis, we predicted that modulation of TGFβ signalling would significantly impact EMT and the malignant potential of EOC spheroid cells. Ovarian cancer patient ascites-derived cells naturally underwent an EMT response when aggregating into spheroids, and this was reversed upon spheroid re-attachment to a …

Characterizing The C-Terminal Region Of Human Adenovirus E1a: An Undiscovered Country, Michael J. Cohen

Electronic Thesis and Dissertation Repository

Human Adenovirus (HAdV) E1A is the first protein expressed during viral infection. The primary function of E1A is to reprogram the cell for viral replication, but it is additionally capable of transforming primary rodent cells in co-operation with other oncogenes such as HAdV E1B. Despite extensive study, little is known about the function and cellular targets of the C-terminal region of E1A. Importantly, this region is required for the transforming ability of E1A with E1B, but can also suppress transformation with Ras. Previous studies showed that interaction with the C-terminal Binding Protein (CtBP) plays a role in both functions described …

Clinical And Experimental Importance Of Circulating Tumor Cells In Prostate Cancer, Lori Lowes

Electronic Thesis and Dissertation Repository

Prostate cancer (PCa) remains a leading cause of death in men, primarily due to ineffective treatment in the metastatic setting. During this phase of PCa, circulating tumor cells (CTCs) are shed into the bloodstream and their presence and number are important in patient prognosis. The CellSearch® system (CSS) is the only U.S. Food and Drug Administration (FDA) and Health Canada approved instrument for detection of CTCs, making it the current clinical gold standard in CTC technology. Although the CSS provides a minimally invasive means of patient monitoring in the metastatic setting, little is known about the role of CTCs in …

The Role Of Connexins And Pannexins In Mammary Gland Development And Tumorigenesis, Michael K. G. Stewart

Electronic Thesis and Dissertation Repository

The identification of key regulators of breast cancer onset and progression is critical for the development of targeted therapies. Connexins and pannexins are characterized by their ability to form large-pore channels and are frequently dysregulated in cancer. However, their role in breast cancer progression remains poorly understood due to a lack of in vivo models capable of assessing the proposed and opposing roles of connexins and pannexins as both tumor suppressors and/or facilitators in multiple stages of the disease. Using 2 previously uncharacterized genetically-modified mice, connexin43 (Cx43) and connexin26 (Cx26) were evaluated for their role in normal mammary gland development …

The Lung Microenvironment Influences The Metastatic Behaviour Of Breast Cancer Cells In An Innovative 3d Ex Vivo Pulmonary Metastasis Model, Matt M. Piaseczny

Electronic Thesis and Dissertation Repository

Lung metastasis remains a leading cause of death in breast cancer patients. This study established an innovative 3D ex vivo pulmonary metastasis assay (PuMA) to test the hypothesis that the lung microenvironment promotes metastatic behaviour of whole population and stem-like ALDH^hiCD44⁺ breast cancer cells. Following in vivo delivery of breast cancer cells to mice, lungs were excised, maintained in culture and imaged to observe breast cancer growth over time. We observed metastatic progression of breast cancer cells in the PuMA, most notably of ALDH^hiCD44⁺ cells which progressed rapidly from single cells to multicellular colonies …

Investigating The Role Of Hydrogen Sulfide In The Survival, Growth And Angiogenic Potential Of Clear Cell Renal Cell Carcinoma Cell Lines And Xenografts, Eric Sonke

Electronic Thesis and Dissertation Repository

Clear cell renal cell carcinoma (ccRCC) is characterized by Von Hippel-Lindau (VHL)-deficiency, resulting in pseudohypoxic, angiogenic and glycolytic tumours. Hydrogen sulfide (H₂S) is an endogenously-produced gasotransmitter that accumulates under hypoxia and has been shown to be pro-angiogenic and cytoprotective in cancer. It was hypothesized that H₂S levels are elevated in VHL-deficient ccRCC, contributing to survival, metabolism, and angiogenesis. Using H₂S-specific probes, it was found that H₂S levels were higher in VHL-deficient ccRCC cell lines compared to cells with wild-type VHL. Inhibition of H₂S-producing enzymes could reduce the proliferation, metabolism …

Atrx Loss-Of-Function In Mouse Neuroprogenitor Cells As A Model Of Early Events In Gliomagenesis, Hannah E. Goldberg

Electronic Thesis and Dissertation Repository

ATRX is a chromatin remodeling protein important for neural development, and ATRX inactivation leads to genomic instability, mitotic defects and TP53-mediated apoptosis. In the last few years, ATRX mutations were identified in a large proportion of paediatric and adult gliomas that often coincide with mutations in the tumor suppressor TP53. The present work shows that combinatorial loss of ATRX and TP53 function in vitro causes genomic instability while improving cell viability, identifying potential early events in gliomagenesis. Furthermore, several gene transcripts associated with glioma development and known oncogenic pathways were significantly upregulated in the Atrx-null neonatal mouse forebrain. …

Characterizing The Interaction Between Ranbpm And C-Raf, Wesley Berube-Janzen

Electronic Thesis and Dissertation Repository

RanBPM/RanBP9 is a multi-domain nucleocytoplasmic protein which has been linked to numerous cellular processes including cell adhesion, migration, transcription and apoptosis. Although RanBPM is a member of the mammalian CTLH complex, the counterpart of a conserved yeast E3 ubiquitin ligase complex, its exact function remains unknown. Previous work in our laboratory has shown that RanBPM inhibits the ERK pathway by interacting with the kinase c-Raf and downregulating c-Raf levels. Here, we show that the N-terminus, LisH/CTLH and CRA domains of RanBPM are required for downregulation of c-Raf and that RanBPM interacts directly with c-Raf through its CRA domain. We also …

Genomic Predictors Of Drug Response To The Alpha-Specific Phosphoinositol 3-Kinase (Pi3ka-Alpha) Inhibitor Byl719 In Head And Neck Cancers, Giananthony T. Rizzo

Electronic Thesis and Dissertation Repository

PIK3CA is the only frequently mutated, druggable oncogene in head and neck squamous cell cancer (HNSCC), with PIK3CA point mutations and gene amplification rates of 17.5% and 40% respectively, with higher rates in HPV-positive disease. The objective of this research was to determine the effects of BYL719, an α-specific PI3K inhibitor in HNSCC cell lines.

All cell lines with PIK3CA hotspot point mutations or gene amplifications will be sensitive to BYL719.

Twenty-eight HNSCC cell lines were subjected to increasing concentrations of BYL719 and cell viability was measured over time. Cell lines were screened for activating PIK3CA hotspot mutations and amplifications …

Investigating Microenvironmental Regulation Of Human Chordoma Cell Behaviour, Priya Patel

Electronic Thesis and Dissertation Repository

The tumor microenvironment is complex and often includes matricellular proteins and regions of hypoxia, which can promote stem and progenitor properties that regulate cancer cell biology. We hypothesized that hypoxia and CCN2 would promote notochord progenitor-like characteristics in human chordoma (U-CH1) cells, and assessed cell phenotype using Real-time qPCR and in vitro functional assays. We found the expression of CCN family members CCN1, CCN2, CCN3 and CCN5 in U-CH1 cells. We demonstrate that hypoxia and CCN2 promoted progenitor-like characteristics specific to the notochordal tissue of origin. Specifically, hypoxia had the greatest ability to promote progenitor characteristics (increase in notochord markers …

Kiss1r Signaling Promotes Breast Cancer Metastasis, Cameron G-F Goertzen

Electronic Thesis and Dissertation Repository

Kisspeptins, peptide products of KISS1, are endogenous ligands for KISS1R, a G protein-coupled receptor. In numerous cancers, KISS1 acts as a metastasis suppressor. However, studies have revealed that patients with elevated KISS1 and KISS1R breast tumor expression have increased tumor grade, increased lymph node metastases and poor survival. We hypothesize that depletion of KISS1R inhibits breast cancer cell metastasis. In order to assess the role of KISS1R in breast cancer metastasis, we used a pre-clinical orthotopic xenograft mouse model using MDA-MB-231 breast cancer cells for breast tumor establishment. We discovered that depletion of KISS1R decreased primary tumor growth and …

Characterization Of The Anti-Apoptotic Function Of The Lysine Demethylase Plant Homeodomain Finger Protein 8 (Phf8), Kimberly Muranko

Electronic Thesis and Dissertation Repository

Apoptosis is an essential process in development and tissue maintenance. The tumor suppressor protein p53 initiates apoptosis through transactivation of pro-apoptotic genes when cellular stress is detected. This study identifies a regulatory role for the lysine demethylase, PHF8, in the p53-mediated apoptosis pathway. We initially suspected PHF8 of demethylating the adaptor protein Numb, however found this to be untrue. PHF8 has been found to have oncogenic properties including an anti-apoptotic effect, however how PHF8 negatively affects apoptosis has not been previously investigated. We found PHF8 inhibits translation of the pro-apoptotic genes TP53, BAX and CASP3. Chromatin immunoprecipitation revealed …

The Role Of Mir-526b In Cox-2 Mediated Human Breast Cancer Progression And Induction Of Stem-Like Phenotype Via Ep4 Receptor Signaling, Erin O. Landman

Electronic Thesis and Dissertation Repository

Our laboratory previously established that aberrant expression of cyclo-oxygenase (COX)-2 promotes breast cancer progression and metastasis via multiple mechanisms, including stem-like cell (SLC) induction, owing to activation of the prostaglandin E2 receptor EP4. COX-2 expression was linked to up-regulation of miRNA-526b. We hypothesized that miR-526b is regulated by EP4 activity, and that miR-526b supports breast cancer progression and induction of SLCs. Using stably miR-526b transfected MCF-7 and SKBR-3 cells in functional assays, including tumorsphere formation in vitro and lung colony formation in vivo, we observed enhanced migration, invasion, proliferation, tumorsphere formation, and in vivo tumorigenecity compared to controls. EP4 …

Sirna Targeting Of Thymidylate Synthase, Thymidine Kinase 1 And Thymidine Kinase 2 As An Anticancer Therapy: A Combinatorial Rnai Approach, Christine Di Cresce

Electronic Thesis and Dissertation Repository

Thymidylate synthase (TS) is the only de novo source of thymidylate (dTMP) for DNA synthesis and repair. Drugs targeting TS protein are a mainstay in cancer treatment but off-target effects and toxicity limit their use. Cytosolic thymidine kinase (TK1) and mitochondrial thymidine kinase (TK2) contribute to an alternative dTMP-producing pathway, by salvaging thymidine from the tumour milieu, and may modulate resistance to TS-targeting drugs. We have previously shown that TS antisense molecules (oligodeoxynucleotides, ODNs, and small interfering siRNA, siRNA) sensitize tumour cells, both in vitro and in vivo, to TS targeting drugs. As both TS and TKs contribute to cellular …

Distinct Roles Of Bmp And Lkb1/Ampk Signalling Impacting Ovarian Cancer Spheroid Biology, Teresa M. Peart

Electronic Thesis and Dissertation Repository

High-grade serous (HGS) carcinoma, the most prevalent and most deadly subtype of epithelial ovarian cancer (EOC), presents unique therapeutic challenges since the majority of cases are diagnosed at advanced, metastatic stage. At this point widespread intraperitoneal metastatic lesions are numerous, which is why models that recapitulate disease dissemination are critical to uncover novel therapeutic targets. One of the initiating events in ovarian cancer metastasis is shedding from the primary tumour into the peritoneal cavity where cells must survive in suspension in order to seed secondary tumours. This non-adherent population of cells exists as multicellular aggregates, or spheroids; data from our …

The Effects Of Atypical Protein Kinase C On Tgfβ Signalling, Adrian D. Gunaratne

Electronic Thesis and Dissertation Repository

The transforming growth factor beta (TGFβ) signalling pathway is an essential regulator of many cellular processes including epithelial growth control, epithelial to mesenchymal transition (EMT), apoptosis, and the establishment of developmental fate. Alterations in TGFβ signalling patterns are associated with various pathological disorders such as fibrosis and cancer. In recent years it has become clear that regulation of TGFβ signalling is dependent on the trafficking and endocytosis of the TGFβ receptors, however, the factors that control these processes are still under investigation.

In this thesis, I examined the role of Protein Kinase C (PKC) in the regulation of TGFβ signalling …

Transforming Growth Factor-Beta Receptor Signalling Is Modulated By Integrin-Linked Kinase, Stellar H. Boo

Electronic Thesis and Dissertation Repository

Transforming growth factor-beta 1 (TGF-β1) modulates regeneration after injury through induction of fibroblast proliferation, migration, and differentiation into myofibroblasts. Induction of myofibroblast differentiation by TGF-β1 requires expression of integrin-linked kinase (ILK). I now show that ILK interacts with TGF-β receptor type II (TβRII) in primary dermal fibroblasts. Further, colocalization of ILK and TβRII can be observed at the cell membrane and in intracellular vesicles. The association of TβRII and ILK does not require TGF-β1 stimulation, kinase activity of TGF-β1 receptor type I or TβRII, and it does not involve interactions between ILK and focal adhesion-associated proteins. When this interaction is …

Cx43 Reduces Melanoma Growth Within A Keratinocyte Microenvironment And During Tumorigenesis In Vivo, Mark J. Ableser

Electronic Thesis and Dissertation Repository

Connexins have been frequently identified as tumor suppressors in many cancers, however, their role in melanoma tumorigenesis remains controversial. Here, we show that B16-BL6 mouse melanoma cells express low levels of Cx26 and Cx43, rendering them gap junctional intercellular communication (GJIC) deficient. Following ectopic expression of Cx26 and Cx43, gap junction-like plaques were evident at the cell surface and the incidence of dye transfer was significantly increased similar to connexin-rich keratinocytes. The expression of Cx43, but not Cx26, significantly reduced proliferation and anchorage-independent growth relative to controls, whereas migration was unaffected. Additionally, Cx43-expressing melanoma cells displayed significantly reduced growth amongst …