Uveal melanoma (UM) is a rare ocular tumor. Up to 50% of the patients develop distant metastases predominantly targeting the liver. The median survival after diagnosis of patients with hepatic... Show moreUveal melanoma (UM) is a rare ocular tumor. Up to 50% of the patients develop distant metastases predominantly targeting the liver. The median survival after diagnosis of patients with hepatic metastases is approximately 4-6 months and hardly increased in the past decades due to lack of novel effective therapeutic options. Within the scope of this thesis we investigated the signaling landscape of metastatic UM and searched for novel avenues of therapy. In Chapter 2 we demonstrate that combinations of the multitarget drug Trabectedin with either the CK2/Clk double-inhibitor Silmitasertib or with the c-MET/TAM receptor inhibitors show synergistic growth inhibitory effects and induce apoptosis of UM cells in vitro. Chapter 3 describes the application of a CRISPR-Cas9 synthetic lethality screen for identification of molecular targets whose inhibition synergistically enhances the effect of the mTOR inhibitor everolimus in UM cells. In Chapter 4 we show that the combination of genetic depletion YAP1/TAZ together with Mcl-1 inhibition resulted in a synergistic inhibitory effect on the viability of UM cell lines. In Chapter 5 we analyzed the phospho-proteome of two UM metastatic cell lines and a primary tumor cell line from the same individual, and studied the role of MARK3 in YAP1/TAZ signaling. Show less
The past two decades have seen the growing development and consequent vast application of next-generation genome editing tools in fundamental and applied research. Nowadays GE based on RNA-guided... Show moreThe past two decades have seen the growing development and consequent vast application of next-generation genome editing tools in fundamental and applied research. Nowadays GE based on RNA-guided nucleases (e.g., engineered CRISPR-Cas9 nucleases) are the most common tools for targeted genetic modification. Nevertheless, these technologies are in need of increased efficiency and accuracy, especially looking forward to translation into diverse clinical applications. The work presented in this thesis focuses on improving the efficiency and accuracy of genome editing, particularly in cells with high therapeutic potential, such as induced pluripotent stem cells (iPSCs), by investigating the feasibility of using adenoviral vectors to test novel genome editing approaches and by exploring the possible applications of a scarless strategy. Show less
Sluimer, L.M.; Bullock, E.; Rätze, M.A.K.; Enserink, L.; Overbeeke, C.; Hornsveld, M.; ... ; Tavares, S. 2023
High expression of the non-receptor tyrosine kinase FER is an independent prognostic factor that correlates with poor survival in breast cancer patients. To investigate whether the kinase activity... Show moreHigh expression of the non-receptor tyrosine kinase FER is an independent prognostic factor that correlates with poor survival in breast cancer patients. To investigate whether the kinase activity of FER is essential for its oncogenic properties, we developed an ATP analogue-sensitive knock-in allele (FERASKI). Specific FER kinase inhibition in MDA-MB-231 cells reduces migration and invasion, as well as metastasis when xenografted into a mouse model of breast cancer. Using the FERASKI system, we identified Ski family transcriptional corepressor 1 (SKOR1) as a direct FER kinase substrate. SKOR1 loss phenocopies FER inhibition, leading to impaired proliferation, migration and invasion, and inhibition of breast cancer growth and metastasis formation in mice. We show that SKOR1 Y234, a candidate FER phosphorylation site, is essential for FER-dependent tumor progression. Finally, our work suggests that the SKOR1 Y234 residue promotes Smad2/3 signaling through SKOR1 binding to Smad3. Our study thus identifies SKOR1 as a mediator of FER-dependent progression of high-risk breast cancers. Show less
