Comet Assay to Quantify DNA Damage in FLT3 Mutant-expressing 32D Cells after Exposure to Type I and Type II FLT3 Inhibitors.

FMS-like tyrosine kinase 3 (FLT3), a class III receptor tyrosine kinase, exhibits a mutation frequency of approximately 30% in acute myeloid leukemia (AML) patients and constitutes a critical therapeutic target. As representative type I and type II FLT3 inhibitors, respectively, gilteritinib and Quizartinib (AC220) are clinically employed in FLT3-mutant AML management. Gilteritinib inhibits both activated and inactivated FLT3 proteins, with additional inhibition of AXL targets to help overcome resistance. AC220 inhibits activated FLT3. The comet assay was systematically employed to quantify and compare DNA damage patterns induced by type I versus type II FLT3 inhibitors in mutant cells. The experimental results revealed that the DNA damage caused by AC220 was significantly higher than that caused by Gilteritinib. For strong DNA damage, FLT3 inhibitors can be combined with DNA damage repair inhibitors to target DNA repair defects. The results provide experimental support for the rational combination strategy of DNA damage-targeting drugs.