Breast cancer is a significant global health issue, impacting millions of women worldwide. ER-positive breast cancer, the most common subtype, accounts for over 70% of cases. While hormone-based therapies like tamoxifen are effective, resistance develops in many patients, posing clinical challenges. Recent research has focused on the molecular mechanisms behind this resistance, aiming to identify new therapeutic targets.
This thesis investigates the regulatory role of microRNAs (miRNAs) in ER-positive breast cancer, particularly their impact on ER expression, cell proliferation, and tamoxifen sensitivity. Through a genome-wide miRNA high-throughput screen, we analysed regulatory networks involving miRNAs to understand their mechanisms better.
Two miRNAs affected ER expression in MCF7 cells, including miR-130a and miR-18b/miR-106a. MiR-18b was highlighted as a key regulator, influencing ER expression and cell proliferation. Analysis of potential targets of miR-18b identified amongst others target genes like ESR1, HMGCS1, and SON, supporting a role of miR-18b in these processes.
Additionally, we identified five miRNAs associated with tamoxifen resistance and validated their role in response to tamoxifen for miR-130a, let-7g, and miR-15b/miR-16-2. For miR-130a, 20 genes, including ESR1 and AIB1, were identified as predicted targets influencing tamoxifen sensitivity. Downregulation of ESR1 and AIB1 by miR-130a was confirmed via Western blotting.
These findings underscore the importance of miRNA regulation in ER-positive breast cancer and pave the way for potential miRNA-based therapeutic strategies to improve treatment outcomes. The research also highlights the complexity of miRNA-mediated regulation and the need for further studies to validate these findings across different breast cancer models.

Classic Hodgkin lymphoma (cHL) is a B-cell malignancy marked by Hodgkin and Reed/Sternberg (HRS) cells that highly express CD30. We explored the roles of microRNAs (miRNAs) in cHL pathogenesis and CD30-targeting therapy using Brentuximab Vedotin (BV) through high-throughput screening techniques. Review of miRNA-related studies on known and potential miRNA-target gene interactions identified multiple miRNA-gene pairs with potential relevance for cHL pathogenesis. This highlighted the relevance of miRNAs in cHL, warranting further studies to deepen our understanding.
A microRNA overexpression library screen revealed 11 miRNA-target gene pairs implicated in cHL pathogenesis and 66 miRNA constructs that regulate BV treatment response. Among these, miR-590 was found to inhibit CD30 expression and induce BV resistance. Additionally, miR-191-5p not only conferred BV resistance but also promoted cell growth. By using AGO2-immunoprecipitation, we identified a network of involved target genes, including PTEN.
To investigate endogenous miRNAs involved in BV resistance mechanisms, we established two BV-resistant cHL cell lines and observed dynamic changes in CD30 expression. Moreover, we utilized a miRNA-focused CRISPR-Cas9 screen to identify endogenous miRNAs that regulate CD30 expression and/or BV sensitivity. To this end, we generated two inducible Cas9 monoclonal cell lines and optimized the entire screening process. These screens await further data analysis.
In summary, this thesis highlights the functional importance of specific miRNAs in cHL and provides detailed procedures of the miRNA-focused screening approaches, from initial setup to optimization and validation of results.

Parasporal proteins from Bacillus thuringiensis (Bt) are well studied in various fields of environmental biotechnology and recently in medical biology. Correspondingly, this dissertation presents the use of site-directed mutagenesis technologies to obtain mutant cry and parasporin proteins with different effects against vector-borne diseases and human cancer cell lines. The use of in silico modeling offers a possible explanation of how the altered residues are involved in changing their structure and interacting with cell membrane receptors. The proteins from this work can be used as targets for new studies that will provide more information about the mechanism of action of Bt proteins, and open the door to the employment of these biomolecules in combating diseases.

The goal of this thesis was to investigate the role of circRNAs in B-cell lymphoma and breast cancer. We focused on the characterization of their expression patterns, functional mechanisms, and regulatory roles. In the first part, we conducted RNA sequencing to characterize the circRNA landscape of three main B-cell lymphoma subtypes and normal B-cells. This was followed by more in-depth studies of the roles of circPVT1 and circZDHHC11 in B-cell lymphoma. CircPVT1 and the linear PVT1 transcripts showed opposite differential expression patterns and both supported cell growth. In contrast to other cancers, we showed that the effect on growth was independent of their ability to bind miRNAs. CircZDHHC11 was strongly enriched in the AGO2-IP fraction upon miR-150 overexpression and supported the growth of B-cell lymphoma. However, its growth-supporting effect was independent of its ability to bind miR-150. In the second part, we investigated the role of circ-NOL10 in breast cancer. Circ-NOL10 was selected based on its most pronounced downregulation in triple-negative breast cancer (TNBC) compared to control tissue. Upon binding of MTDH and CASC3, circ-NOL10 levels were strongly reduced, resulting in the release of miR-149-5p, miR-330-3p, and miR-452-5p. These miRNAs subsequently targeted PDCD4 and this promoted progression of breast cancer. Overall, our study extends current knowledge of circRNAs and highlights specific functions of selected circRNAs in B-cell lymphoma and breast cancer.

Unfortunately, lymphoma (Hodgkin’s and Non-Hodgkin’s lymphoma) is still an incurable disease for many people and new and smart treatment techniques are needed to improve the prognosis of these patients.
The goal of the research described in this dissertation was to find pre-existing medications used in other cancers, using a technique that allows to test the dependence on anti-apoptotic proteins (proteins important for the survival of cancer cells). This technique is called BH3 profiling.
By applying this technique, it has been found that tamoxifen, an affordable, anti-hormone drug that has been used in breast cancer for more than 40 years, can kill lymph node cancer cells in laboratory experiments and in animal experimental models. If tamoxifen is combined with a drug that blocks anti-apoptotic proteins, the anticancer effect is enhanced. This has led to a clinical trial in patients with untreatable aggressive lymphoma.
In addition to these important findings and applications, this thesis describes the efficacy of combinations of different drugs in lymphoma and whether these agents alter the dependence on these anti-apoptotic agents.
This project was carried out through a KWF grant.