Abstract
Background
Our preliminary studies identified the arylpiperazine derivative NAF19 as a promising therapeutic agent against prostate cancer, though its precise mechanisms remained unclear.
Methods
Network pharmacology was utilized to pinpoint both therapeutic targets and disease-related targets. Functional assessments of cells were conducted using the CCK-8 method and wound-healing assays to analyze the effects of NAF19 on LNCaP cell proliferation and migration, respectively. The effect of NAF19 on the cell cycle progression of prostate cancer cells was examined using flow cytometry, while phosphorylation of AKT and PI3K was assessed using Western blotting.
Results
The anti-proliferative activity of NAF19 increased with higher doses and longer durations of treatment. The wound healing assay confirms its strong inhibitory effect on cell migration. Western blotting results indicate that NAF19 influences EMT-related protein expression, leading to elevated E-cadherin and decreased Vimentin and N-cadherin levels. Further mechanistic investigation showed that NAF19 significantly reduced the expression of CXCR4, as well as phosphorylated PI3K and Akt proteins. Western blot results confirmed that NAF19 downregulated Skp2 while increasing the levels of p27 and p21, suggesting involvement of the Skp2/p27/p21 signaling pathway. Molecular docking analysis demonstrated that NAF19 binds strongly to CXCR4, exhibiting a binding energy of -9.6 kcal/mol.
Conclusions
This investigation confirms that NAF19 acts as a new CXCR4 antagonist, effectively suppressing several cancer-promoting pathways in prostate cancer, such as PI3K/AKT signaling, cell cycle advancement, and EMT.