DNA/TOP2A-IN-1 functions as an inhibitor of DNA and TOP2A. It binds selectively to TOP2A instead of TOP2B and interacts with DNA to form a stable DM1-TOP2A-DNA ternary complex. This compound induces DNA damage, elevates reactive oxygen species (ROS), and triggers apoptosis in triple-negative breast cancer (TNBC) cells. Additionally, DNA/TOP2A-IN-1 disrupts microtubule distribution and induces cell cycle arrest while exhibiting potent antiproliferative activity and inhibiting cell migration. It effectively suppresses tumor growth and is a valuable agent for TNBC research.

DNA/TOP2A-IN-1 (Compound DM1) interacts with TOP2A to disrupt the DNA secondary structure. At concentrations of 0-1.6 μM for 24 hours, it binds to DNA within 4T1 cells, forming a ternary DM1-TOP2A-DNA complex, and induces cellular damage. DM1, in the range of 10-100 μM and at 37 ℃ for 30 minutes, inhibits TOP2A activity by preventing TOP2A-mediated DNA relaxation. DNA/TOP2A-IN-1 suppresses the proliferation of various cancer cells with IC50 values of 1.12 μM (4T1), 2.53 μM (MDA-MB-231), 2.68 μM (MDA-MB-468), 5.02 μM (MCF-7), 3.80 μM (Hela), 3.44 μM (SiHa), 4.78 μM (PC9), 3.68 μM (A549), 2.7 μM (ID8), 2.69 μM (PANC1), and 9.03 μM (BEAS-2B) over 48 hours. It inhibits cell viability, proliferation, and migration in a dose-dependent manner in 4T1 and MDA-MB-231 cells, with ranges of 0-4 μM (24-72 hours), 0-3 μM (3 days), and 0-12 μM (24 hours), respectively. At 0-2.4 μM for 24 hours, DNA/TOP2A-IN-1 causes microfilament collapse and disintegration, along with changes in actin and microtubule distribution in 4T1 and MDA-MB-231 cells. Additionally, it promotes apoptosis and increases intracellular reactive oxygen species in 4T1 and MDA-MB-231 cells at 0-7.2 μM over 0-48 hours. This compound induces cell cycle arrest at the G2/M phase in these cells at 0-12 μM for 24 hours. Moreover, DNA/TOP2A-IN-1 at concentrations of 2 and 10 μM over 6 days significantly inhibits breast cancer growth in organoids.

DNA/TOP2A-IN-1 (Compound DM1), administered via intraperitoneal injection at doses of 20 and 30 mg/kg every two days over a 14-day period, demonstrates antitumor activity in a 4T1 tumor model in BALB/c mice.