Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults, in which B-cell receptor signaling plays a key role in regulating the survival of leukemic cells. As the tyrosine kinases Lyn and Lck are important regulators of B-cell receptor signaling, they represent promising therapeutic targets for improving the efficacy of treatment with anti-CD20 monoclonal antibodies. The main aim of this master’s thesis was to evaluate the cytotoxic effects of saracatinib and HY-12072 in combination with rituximab and obinutuzumab in vitro and to determine whether these drug combinations exhibit synergistic cytotoxic effects.
Since the activity of anti-CD20 monoclonal antibodies is associated with activation of the complement system, we first investigated the effect of human serum on complement-mediated cytotoxicity induced by rituximab and obinutuzumab in MEC-1 cells. As the concentration of added human serum increased from 0% to 20%, the relative metabolic activity of MEC-1 cells decreased from 92.0% to 29.4% following treatment with rituximab and from 77.5% to 27.1% following treatment with obinutuzumab.
We subsequently evaluated the cytotoxic effects of saracatinib and HY-12072 in primary CLL cells and their combined effects with rituximab and obinutuzumab in MEC-1 cells. In primary CLL cells, saracatinib and HY-12072 induced concentration- and time-dependent decreases in relative metabolic activity. The median IC₅₀ value of saracatinib was 22.52 μM after 24 h and 8.86 μM after 48 h, whereas the median IC₅₀ value of HY-12072 was 3.78 μM after 24 h and 1.63 μM after 48 h. When evaluating the combined effects of saracatinib and HY-12072 with anti-CD20 monoclonal antibodies, relative metabolic activity of MEC-1 cells at saracatinib concentrations of 15 and 60 μM was 81.1% and 30.9%, respectively, in combination with rituximab, and 55.4% and 33.5%, respectively, in combination with obinutuzumab. At HY-12072 concentrations of 0.15 and 0.6 μM, relative metabolic activity was 79.5% and 59.7%, respectively, in combination with rituximab, and 56.3% and 49.0%, respectively, in combination with obinutuzumab. The combinations resulted in decreases in relative metabolic activity comparable to those observed with the individual agents.
These findings contribute to a better understanding of the response of CLL cells to treatment with saracatinib and HY-12072 in combination with rituximab and obinutuzumab and provide a basis for further investigation of novel combination therapeutic approaches.
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