Pyrazolo[3,4-d]pyrimidine tyrosine kinase inhibitors induce oxidative stress in patient-derived glioblastoma cells

Background: Glioblastoma (GBM) highly expresses Src tyrosine kinase involved in sur-vival, proliferation, angiogenesis and invasiveness of tumor cells. Src activation also reduces reactive oxygen species (ROS) generation, whereas Src inhibitors are able to increase cellular ROS lev-els. Methods: Pro-oxidative effects of two pyrazolo[3,4-d]pyrimidine derivatives—Src tyrosine kinase inhibitors, Si306 and its prodrug pro-Si306—were investigated in human GBM cells U87 and patient-derived GBM-6. ROS production and changes in mitochondrial membrane potential were assessed by flow cytometry. The expression levels of superoxide dismutase 1 (SOD1) and 2 (SOD2) were studied by Western blot. DNA damage, cell death induction and senescence were also ex-amined in GBM-6 cells. Results: Si306 and pro-Si306 more prominently triggered ROS production and expression of antioxidant enzymes in primary GBM cells. These effects were followed by mitochondrial membrane potential disruption, double-strand DNA breaks and senescence that eventually led to necrosis. Conclusion: Src kinase inhibitors, Si306 and pro-Si306, showed significant pro-oxidative potential in patient-derived GBM cells. This feature contributes to the already demonstrated anti-glioblastoma properties of these compounds in vitro and in vivo and encour-ages clinical investigations.