Significant progress has been made in the diagnosis and treatment of cancer in recent years. However, many patients still suffer from treatment resistance and cancer recurrence. This can be attributed to a small population of cancer stem cells (CSCs), a subpopulation of cells that are intrinsically resistant to most existing therapeutic approaches. Targeting CSCs could be an effective strategy to overcome the limitations of current cancer therapies and prevent cancer recurrence. The lysosomal cysteine peptidases, cathepsins B and X, represent promising molecular targets as they are involved in key processes of cancer development and progression. Increased expression of cathepsins B and X in cancer is associated with a less differentiated cellular phenotype and the transition of cancer cells between epithelial and mesenchymal phenotypes. To successfully incorporate cathepsins B and X into novel therapeutic approaches for cancer treatment, it is critical to understand the effects of their inhibition on CSC phenotype changes.
We isolated CSCs from the MCF7, MDA-MB-231, and MCF-10A neoT cell lines based on their ability to form tumor spheres. Using RT-qPCR with specific oligonucleotide primers, we demonstrated that the expression of cathepsin B and X is increased in CSCs isolated by tumor sphere formation compared to differentiated cancer cells prior to CSC isolation. By inhibiting cathepsins B and X with specific inhibitors, we then investigated the effects on the expression of genes for cancer markers differentially expressed in CSCs using appropriate oligonucleotide primers. We showed that inhibiton of cathepsins B and X affects the phenotype of CSC, as it leads to a reduction in the expression of CSC markers. We also evaluated the effects of combined therapy with cathepsin inhibitors and the conventional chemotherapeutic 5-fluorouracil on the expression of CSC markers in isolated CSCs. We showed that with some combinations of 5-fluorouracil and cathepsin B or X inhibitors, a synergistic effect occurs that further reduces the expression of CSC markers.
In this master thesis, we demonstrated that inhibition of cathepsin B and X reduces the expression of CSC markers, thereby reducing the stem cell capability of CSCs and targeting CSCs that are resistant to most existing therapeutic approaches. This confirms cathepsins B and X as potential therapeutic targets in cancer therapy.
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