ATM/ATR and SMAD3 pathways contribute to 3-indole-induced G₁ arrest in cancer cells and xenograft models

Background: 3-Indole inhibits lung cancer growth by apoptosis. Here, the growth inhibition mechanism besides apoptosis was further characterized. Materials and Methods: The Comet assay was used to examine 3-indole-induced DNA damage. Cell cycle distribution and protein expression were analyzed using flow cytometry, Western blotting and immunohistochemistry in cell and animal models. Results: 3-Indole induced dose-dependent DNA damage, which was reversed by reactive oxygen species (ROS) inhibitor in lung cancer cells. Cell cycle G ₁ arrest was observed in the 3-indole-treated cells. DNA damage-responsive proteins involved in the ataxia telangiectasia mutated/ataxia telangiectasia and Rad3-related (ATM/ATR) pathway and G₁ regulation proteins such as p21 and SMA- and MAD-related protein 3 (SMAD3) were induced in the cell models. The altered expression of ATM, ATR, checkpoint kinase 2 (CHK2), and cell division cycle 25 homolog A (CDC25A) were confirmed in xenograft models. Importantly, the 3-indole-induced ATM/ATR and transforming growth factor (TGF)-β/SMAD pathways were attenuated by ROS inhibitor. Conclusion: 3-Indole causes DNA damage and triggers ATM/ATR and SMAD3 signaling pathways to arrest lung cancer cells at the G₁-phase.