In vitro cytotoxicity evaluation of triphenylphosphonium (TPP) conjugates of some acetylenated nucleic bases and their analogues

In this work, we designed and synthesized 12 triphenylphosphonium (TPP) conjugates of acetylenated nucleic bases (uracil, thymine) and their analogues (6-methyluracil, quinazoline-2,4-dione) and evaluated their in vitro cytotoxicity against 9 human cancer cell lines M-HeLa, HuTu 80, MCF-7, T 98 G, A 549, DU-145, SK-OV-3, PC-3, A-375 and two lines of normal human cells RPMI 1788 and WI-38. All synthesized TPP-conjugates showed high cytotoxicity (IC₅₀ values in the range of 0.1–7.3 µM) against all used human cancer cell lines. The mechanisms of cytotoxic action were studied for the lead compounds 2c,d, 4c,d which exhibited very high cytotoxicity (IC₅₀ = 0.2–0.3 μM) against PC-3 cancer cells. The flow cytometry method using Annexin V and propidium iodide (PI) has shown that the lead compounds cause apoptosis of PC-3 cells. With the help of flow cytometry using cationic carbocyanine dye JC-1, it was found that the lead compounds cause a significant dose-dependent decrease in the mitochondrial membrane potential of PC-3 cancer cells, that induces apoptosis along the mitochondrial pathway. Significant ROS production in PC-3 cells after their treatment with the lead compounds 2c,d was detected by flow cytometry using CellROX® Deep Red fluorogenic probe. Enzyme-linked immunosorbent assay (ELISA) found that the lead compounds activated apoptosis-initiating caspase-9 and blocked anti-apoptotic Bcl-2 protein in PC-3 cancer cells. This experimental fact was explained by molecular docking.