Research On The Design, Synthesis, Antitumor Activity And Mechanism Of ?-N-heterocyclic Chelate Gallium(?) And Copper(?) Complexes

Gallium???is the second metal element used in tumor chemotherapy after platinum???.It has a broad spectrum of anti-tumor activity,especially for breast cancer,lymphoma,osteosarcoma,non-small cell lung cancer,etc.However,gallium???is easily hydrolyzed to form gallium hydroxide,generating a low solubility polymer,resulting in very poor absorption of gallium???by the human body.Coordination with organic ligands not only prevents their hydroxylation,but also increases the lipophilicity of gallium???,further improving the bioavailability and antitumor activity of gallium???.Therefore,this paper studied the design,synthesis and mechanism of?-N-heterocyclic chelated gallium???antitumor drugs.Firstly,two types of 2-pyridinecarboxaldehyde thiosemicarbazones ligands?L1 and L2?and four Ga???complexes?C1-C4?have been synthesized,and determined by X-ray single crystal diffraction which are 2:1?C1 and C3?and 1:1?C2 and C4?ligand/Ga???complexes.The antiproliferative activity of these Ga???complexes have been examined to illuminate the structureactivity relationships essential to form Ga???complexes with remarkable anticancer activity.In addition,Ga???complexes where the metal/ligand ratio was 1:1?C4?had observably higher antiproliferative activity than 1:2?C3?.Ga???complexes caused a marked increase of caspase-3 and9 activity in NCIH460 cells compared to the metal free ligand.Caspase activation was somewhat mediated by the release of Cyt C from mitochondria after incubation with selected agents.Both types of Ga???complexes showed more effective in inhibition of the G1/S transition than the ligand alone.In order to study the mechanism of Ga???complex inhibiting cell cycle,We have synthesized and characterized five thiosemicarbazone Ga???complexes?C5�C9?were by X-ray single crystal diffraction,and they were all 1:1 ligand/Ga???complexes.The antiproliferative activity of these Ga???complexes was tested against three cancer cell lines,demonstrating that Ga???complexes showed about3�10 folds more anticancer activity than their ligands alone.Importantly,thiosemicarbazones and Ga???complexes have a low toxicity to human fetal lung fibroblast cells?MRC-5?and exhibit a high therapeutic index for tumor cells.The results of UV�visible spectroscopy showed that the binding constant of C8 with Topo-I-DNA was significantly higher than that of L5.The Ga???complex?C8?caused Topo-I inhibition and distinct DNA cleavage.Moreover,Ga???complex and thiosemicarbazone ligand prolonged the G1 phase in NCI-H460 cell cycle,which might be depended on the ability of these compounds to affect the expression of cell cycle related proteins.In order to study the mechanism of Ga???complex promoting apoptosis,were synthesized and characterized five thiosemicarbazone ligands by condensation with different aldehydes or ketones by 4-phenylthiosemicarbazone.C13 was characterized by X-ray single crystal diffraction,which was 1:1 ligand/Ga???complexes.The structure-activity relationship of these ligands and Ga???complexes have been investigated,and the results demonstrate that the formation of Ga???complexes have significant antiproliferative activity over the corresponding ligands.The anticancer mechanism of gallium???complexes has been studied in detail,which is typical agents that effect on the mitochondrial apoptotic pathway.The ability of gallium???complexes to inhibit the cell cycle does not enhanced with the increasing concentrations,whereas the ability to promote apoptosis is concentration-dependent.To investigate the effects of Ga???complexes on ROS levels,we synthesized three bis-ligated gallium???�2-benzoylpyridine-thiosemicarbazone complexes and studied their antitumor mechanisms.The structures of the Ga???complexes were identified by X-ray singlecrystal diffraction.Cytotoxicity analysis demonstrated that the ligands and gallium complexes exerted a higher antitumor activity and a lower cytotoxicity than those of normal cells.The most active complex was C17,which exhibited a better antitumor viability than its related ligands and the anticancer agent 3-AP.Thus,the Ga???complexes not only transmitted the iron ions but also induced intracellular Ca²⁺release.As a result,the ROS standards in redox-active iron complexes were increased.The mechanism involved the release of Cyt C from the mitochondria which lack membrane potential,and then the activation of the caspase family proteins stimulated cell apoptosis.Copper is an essential element in most aerobic organisms and serves as a structural and catalytic cofactor,so it is involved in many biological pathways and its role in the development of cancer and other diseases.Copper???complexes efficiently catalyze hydrogen peroxide to generate reactive oxygen species?ROS?,which is the major reason for its significant anti-tumor activity.We synthesized three Cu???piperidylthiosemicarbazone complexes and examined their structures by X-ray single crystal diffraction.These Cu???complexes have significant apoptosis-promoting activity at nanomolar concentrations.The antitumor activity of these Cu???complexes is increased by more than 40-fold relative to that of the ligand.The binding experiment results demonstrated that the Cu???complexes interact with DNA with moderate binding affinity by in situ intercalation.Gel electrophoresis analysis indicated that DNA is degraded when the copper complex catalyzes hydrogen peroxide to produce reactive oxygen species?ROS?.Apoptosis mechanism results showed that excessive ROS leads to mitochondrial membrane potential dissipation and promote the release of apoptotic factors from mitochondria.