Synthesis, Activity And Fluorescence Imaging Of Novel Monofunctional Antitumor Platinum Complexes

Platinum anticancer agents are always one of the most attractive fields in inorganic medicinal chemistry. However, the current platinum agents are still suffering from their severe toxicity and distinct acquired/intrinsic resistance and limit their application. Many novel platinum anticancer compounds, which are not approved by the canonical structure-activity relationships, were developed to overcome these disadvantages of the current cisplatin analogues presumably due to the different antitumor mechanisms. On the other hand, the trial to track Ptâ…¡uptake, transportation, distribution and metabolism in real time is also a dependent approach to provide new clues to novel anticancer agents. Both the antitumor activity and in vitro/in vivo tracking of cationic monofunctinal platinum complexes were investigated in this thesis to explore their potential as the novel antitumor drugs.To explore the cellular process of monofunctional complex, two cationic fluorescent platinum complexes, I and II, were synthesized from two fluorescent ligands derived from 4-nitro-2,1,3-benzoxadiazole (NBD) fluorophore containing a pyridine group to coordinate to cis-[Pt(NH3)2]2+ motif. Fluorescence and ESMS determination showed that complex I of an additional methyl group results in the higher stability than complex II, which makes complex I be more suitable for fluorescence cell imaging. The confocal fluorescence imaging on HeLa cells incubated with complex I illustrates that the cell uptake of complex I is very slow and the fluorescent complex can finally get into the nucleus. Although the cellular uptake of the related ligand L1 is very quick and L1 accumulates around the nucleus, this ligand cannot get into the nucleus. It is obvious that the cationic monofunctional platinum complex is difficult in striding across the lipid bilayer, and the introduction of Ptâ…¡center endows the complex with the nuclear affinity. There is no distinct apoptosis phenomenon in the HeLa cells even after long time of incubation with I. However, the massive vacuoles were observed in the cytoplasm, suggesting that the monofunctional pyriplatin complex might lead to a nonapoptotic cell death, which is different from the apoptosis mechanism of cisplatin. MTT determination demonstrates that all the two free ligands are non-toxic towards Raji, BEL-7402, MCF-7 and HeLa cells, and the introduction of Ptâ…¡improves their cytotoxic activities to these tumor cell lines distinctly with complexâ… of even higher activity. These results demonstrate that monofuncitonal pyriplatin complex has distinct anticancer activity, which paves a new way to design new Ptâ…¡-based antitumor agents.After that, a new cationic 3N chelated fluorescent monofunctional platinum complexâ…¢was synthesized from the NBD derivative with a 2,2'-dipicolylamine (DPA) chelating group. The chelating effect makes the complex be more stable, and the Ptâ…¡coordination to DPA motif results in a 5-fold emission enhancement, favoring the desired cell imaging. Confocal fluorescence imaging of MCF-7 incubated with complexâ…¢disclosed that its cellular uptake is very slow. Besides the nucleolus preferential affinity, this positively-charged monofunctional complex displays also the accumulation on mitochondria in cytoplasm, which finally induces the mitochondria swelling to form vacuoles in cytoplasm after long time of incubation. Such a vacuole formation process implies the possibility of cell paraptosis. Moreover, its nucleolus binding ability is stronger than its mitochondria affinity. In addition, we have realized the in vivo fluorescence imaging of platinum complex for the first time with a stereomicroscope, profiting from the visible light excitability of complexâ…¢and the transparency of zebrafish larva. The in vivo imaging on zebrafish larva stained by complexâ…¢revealed that the complex could accumulate in the liver or cervicales/notochord with the variable incubation time and concentration. The current in vivo imaging provides a fine model for real time Ptâ…¡tracking in situ. CD spectra showed that complex III is able to affect the base stacking and decrease the helicity of CT-DNA by platination. All these results should not only promote to understand the antitumour mechanism of this kind of Ptâ…¡complexes but also favor to develop new drugs based on monofunctional chelated Ptâ…¡.Finally, three complexes IV, V and VI have been synthesized as a new kind of monofunctional Ptâ…¡complex of DNA intercalating nature by conjugating a pyriplatin moiety with the 1,8-naphthalimide group of different substituents. CD spectra and gel electrophoresis of CT-DNA showed the intensive DNA binding ability of the three complexes, which can be ascribed to the naphthalimide intercalation and platination, are able to induce the transition from B- to A-type DNA. Cytotoxicity assay against HeLa, BEL-7402 and MCF-7 cell lines indicates that both complex IV and V are more active than cisplatin. Although complex VI with a 4-nitro-1,8-naphthalimide group is more cytotoxic than cisplain against BEL-7402, MCF-7 cells, its IC50 value is higher than cisplatin towards HeLa cells. All these suggest that the antitumor activity of monofunctional platinum complexes could be improved by tethering with the DNA intercalator such as 1,8-naphthalimide group. These results have enlarged the field of platinum-based antitumor agents and provide some new clues to design Ptâ…¡-based drugs of higher activity and low side effects.