Novel cancer-related regulatory targets for the signalling sphingolipids sphingosine-1-phosphate and sphingosylphosphorylcholine

The signalling sphingolipid sphingosine-1-phosphate (S1P) is necessary for development of the immune system and vasculature and on a cellular level regulates migration, proliferation and survival. Due to these traits S1P has an important role in cancer biology. It is considered a primarily cancer-promoting factor and the enzyme which produces it, sphingosine kinase (SphK), is often over-expressed in tumours. S1P is naturally present in the blood, lymph, tissue fluids and cell cytoplasm and functions through its cell surface receptors (S1P1-5) and as an intracellular second messenger. Sphingosylphosphorylcholine (SPC) is closely related to S1P and has similar regulatory functions but has not been extensively studied. Both S1P and SPC are able to evoke either stimulatory or inhibitory effects on cancer cells depending on the context. The aim of this thesis work was to study novel regulatory targets of S1P and SPC, which mediate the effects of S1P/SPC signalling on cancer cell behaviour. The investigated targets are the transcription factor hypoxia-inducible factor 1 (HIF-1), the intermediate filament protein vimentin and components of the Hippo signalling pathway. HIF-1 has a central role in cancer biology, as it regulates a multitude of cancer-related genes and is potently activated by intratumoural hypoxia through stabilization of the regulatory subunit HIF-1α. Tumours typically harbour high HIF-1α levels and HIF-1, in turn, facilitates tumour angiogenesis and metastasis and regulates cancer cell metabolism. We found S1P to induce follicular thyroid cancer cell migration in normal oxygen conditions by increasing HIF-1α synthesis and stability and subsequently HIF-1 activity. Vimentin is a central regulator of cell motility and is also commonly over-expressed in cancers. Vimentin filaments form a cytoskeletal network in mesenchymal cells as well as epithelial cancer cells which have gone through epithelial-mesenchymal transition (EMT). Vimentin is heavily involved in cancer cell invasion and gives tumours metastatic potential. We saw both S1P and SPC induce phosphorylation of vimentin monomers and reorganization of the vimentin filament network in breast and anaplastic thyroid cancer cells. We also found vimentin to mediate the anti-migratory effect of S1P/SPC on these cells. The Hippo pathway is a novel signalling cascade which controls cancer-related processes such as cellular proliferation and survival in response to various extracellular signals. The core of the pathway consists of the transcriptional regulators YAP and TAZ, which activate predominantly cancer-promoting genes, and the tumour suppressive kinases Lats1 and Lats2 which inhibit YAP/TAZ. Increased YAP expression and activity has been reported for a wide variety of cancers. We found SPC to regulate Hippo signalling in breast cancer cells in a two-fold manner through effects on phosphorylation status, activity and/or expression of YAP and Lats2. In conclusion, this thesis reveals new details of the signalling function of S1P and SPC and regulation of the central oncogenic factors HIF-1 and vimentin as well as the novel cancer-related pathway Hippo.

Crowdfunding as a new funding alternative for early stage start-ups

Attracting outside capital is a common problem for start-up companies. Capital markets define the funding options for companies and firms which suffer the most from these capital market imperfections are small start-up companies. Therefore it is important to study any new funding model which can offer a new solution to this inefficiency of capital markets. This study explains the traditional funding models for start-ups such as founders, friends & family, banks, business angels and venture capitalist. After giving background to traditional start-up funding this study delves into crowfunding (CF) and introduces it as a new funding method. The objective of the thesis is to answer one broad research question: Why and how should start-up companies use CF as an alternative funding method? To properly delve into this, this question has the following sub-questions: What kind of funding alternatives do start-up companies have? What are the pros and cons of CF compared to other funding options? How can start-ups benefit from CF? This study gives background on the rise of CF and the reasons why this new model is needed. Author will explain the different components of CF such as platforms, crowdfunders and projects. Also benefits and challenges of the crowdfunding model are investigated. As a new funding model CF has had to clear out many obstacles from its way. These are, for example, legal and regulatory issues as well education of crowd investors to understand this new investment option. . Start-up entrepreneurs can gain valuable insight from this study. The author has attempted to form best practices and guidelines of how to operate in the CF environment. This study was conducted by performing expert interviews, collecting data from previous studies and performing a content analysis of successful crowdfunding cases. Main findings from the study were that CF has huge potential in funding entrepreneurial projects. It is still a niche way for funding but growing rapidly. CF is earning its place among traditional funding options and has potential to fund projects which otherwise would struggle to find funding. With CF entrepreneur can tap into geographically diverse audience. It is a powerful validation tool for products and ideas and has the power to bring democratic elements to entrepreneurial funding.