1,2,3-Triazole-substituted cabotegravir analogues, i.e., KJ-9, have been developed as lead structures to explore their potential as antitumor agents. The new analogue exhibited significant anti-proliferative activity against various human cancer cell lines, with particularly strong effects on the HepG2 and HCCLM3 liver cancer lines, as it induced a marked loss of colony-forming ability and triggered apoptosis upon KJ-9 exposure. Furthermore, treatment with KJ-9 increased the Bax-to-Bcl-2 protein ratio and activated cleaved caspase-9, caspase-3, and poly(ADP-ribose) polymerase (PARP). Meanwhile, KJ-9 treatment induced a blockage in the cell cycle (G2/M), increased DNA damage levels, and induced the accumulation of histone variant H2AX (γ-H2AX) protein. Furthermore, there was strong induction of p-ATM and p-ATR proteins, along with their downstream effectors p-CHK1 and p-CHK2. Additionally, KJ-9 treatment increased phosphorylation levels of the tumor suppressor protein p53 and inhibited components of the PI3K/AKT pathway. Although it did not significantly affect AKT phosphorylation, the ATM/ATR inhibitor CGK733 significantly reversed KJ-9-induced upregulation of p-ATM, p-ATR, p-p53, γ-H2AX, and activated caspase-3. Moreover, reactive oxygen species (ROS) were generated in greater quantities by KJ-9 treatment. After KJ-9 treatment, ROS were suppressed by the addition of the antioxidant N-Acetylcysteine (NAC), leading to increased levels of p-AKT and reduced levels of p-ATM, p-ATR, p-p53, cleaved caspase-3, and γ-H2AX. These findings suggest that KJ-9 promotes oxidative stress, which further inhibits AKT activation while activating the ATM/ATR pathway, leading to p53 accumulation, sustained DNA damage responses, G2/M-phase cell cycle arrest, and apoptosis in both HepG2 cells and HCCLM3 cells.
