Design,Synthesis And Mechanistic Studies Of 1-N-substituted Acridone Derivatives Based On Antitumor Activity

Cancer is one of the major diseases that seriously threaten health and life of human.Cytotoxic drugs,especially many DNA-targeted drugs,have been widely used in clinic as first-line chemotherapy drugs,such as carboplatin,adriamycin and amsacrine.However,owing to the lack of selectivity towards normal cells and cancer cells,the resulting serious side effects of these cytotoxic drugs have greatly limited their clinic use.Therefore,further development of cytotoxic drugs with high effectiveness and low toxicity is of great significance.Given the fact that many acridine and acridone entities from natural source have been proved to be promising antitumor agents,in our previous work,we have synthesized a series of acridone-based derivatives as potent anticancer agents and the preliminary structure-activity relationship(SAR)investigation was conducted.Among these compounds,E17 and E24 displayed the most potent antiproliferative activits.In particular,E17 strongly inhibited the activity of topo Ⅱ without causing significant degradation of topo Ⅱ level in contrast to the controls,VP16 or ICRF-187,implying E17 serves as a topo Ⅱ inhibitor and might not induce MDR.Interestingly,the structural difference between these two hits is very small,a difference of one carbon atom in the linkers between 1-N nitrogen and piperazine motif.This means the follow-up structural optimization could be concentrated on the linker variations.So,a series of N-substituded acridone derivatives with different length of linkers were designed and synthesized.In the first round of chemical optimization,we focused on the linker length between 1-NH and distal N-methyl piperazine fragment,and compounds 6a-g were synthesized herein.As a result,compound 6b with an ethyl linker gave the most potent anti-proliferative activity and it provided about 4-fold enhancement compared with the positive control Etoposide.However,the prolongation of the carbon length led to the slowly decreased activity,further confirming the size and disposition of linker greatly affected the bioactivity.The second round structural optimization was carried out based on 6b.To detect whether the replacement of the three nitrogen atoms(a,b and c)of 6b with other smaller sized groups would affect the bioactivity,compounds 6h-61 were further prepared.Surprisingly,compounds 6h,6l and 6i significantly elevated the antiproliferative activities.Compound 6h gave the most potent anti-proliferative activity among all the test compounds.This results proved the importance of three nitrogen atoms to bioactivity,with the influencing sequence of Nc>Nb>Na.After the completion of two rounds of chemical optimizations,further bioevaluation on the most potent compound 6h was systematically carried out.It revealed that 6h is a strong topo Ⅱ poison which could trap ’DNA-topo Ⅱ’ cleavage complex and cause obvious DNA double strand breaks(DSBs).DNA intercalating and DNA binding assays further proved 6h could strongly bind to DNA.The topoisomerase I and Ⅱα/β inhibition of 6h were subsequently determined by DNA relaxation assay,and the results were represented by IC50 values.It showed that 6h exhibited superior selective inhibition towards topo Ⅱ over topo I,but gave similar inhibitory activities towards two topo Ⅱ isoforms,topo Ⅱα and topo Ⅱβ,implying that 6h may be a favorable topo Ⅱα/β inhibitor.In addition,6h could apparently induce cell cycle arrest in two cancer cells and induce serious apoptosis in KG1 cells.Finally,considering a majority of reported topo Ⅱ inhibitors,e.g.doxorubicin,with cardiac toxicities,the cardiac toxicity of 6h in rat myocardial H9C2 cells was thereby checked.It evinced that in 6h treated H9C2 cells,the cell viability and mitochondrial membrane potential were almost unchanged,while the control doxorubicin changed obviously,suggesting 6h might not induce serious cardiotoxicity in comparison with doxorubicin.In summary,in this thesis,we have conducetd two rounds of chemical optimizations based on hit E17 with the expectation of enriching the SAR outcomes of acridone-derived anticancer agents.Among all these synthesized acridone derivatives,compound 6h with the most anticancer activity was submitted to the mechanism of action study.Collectively,our results confirmed 6h is an invaluable candidate that is worthy of further in-depth development.