2.2. c-Src/c-Abl inhibitor Si162 and primary melanoma cell treatment

Si162, a pyrazolo[3,4-d]pyrimidine derivative, was synthesized according to the method described by Kruewel et al. (2010), who also described detailed information about the structural and chemical characteristics of Si162.

To assess the cytotoxic response of melanoma cells to Si162, the cells were grown until they reached a specific density of 70%–80%. The cells were then detached using trypsin and seeded onto a sterile 96-well plate at a density of 1000 cells per well. To allow the cells to adhere to the bottom of the plate, they were incubated for 24 h at 37 °C in a 5% CO₂ environment.

After the 24-h incubation period, 3 wells were treated with 8 different concentrations of Si162, ranging from 0.02 μM to 20 μM, in 10 μL of medium. The treated cells were further incubated for 72 h at 37 °C in an environment with 5% CO₂. This experimental setup aimed to evaluate the cytotoxic effects of Si162 on the melanoma cells.