Efficient Asymmetric Synthesis Of α-CF₂H Spiro[Pyrrolidine-Oxindole] And Its In Vitro Antitumor Activity

p53 plays an important role as an anti-oncogene in the process of apoptosis.Deletions or mutations in the p53 gene are present in about 60%of human cancers.MDM2,the most important negative regulator,can form a complex with p53 to inactivate the p53 gene.Researchers have identified several novel inhibitors of p53-MDM2 through extensive drug screens,and among these novel inhibitors,small molecule inhibitors are the most important component.Among the small-molecule compounds that have entered preclinical studies,spirooxindoles have received a lot of attention from the medicinal chemistry community due to their significant anticancer activity and unique structural features.In the past few years,spirocyclic oxindole derivatives containing trifluoromethyl（-CF₃）have been designed and successfully synthesized for application as p53-MDM2 inhibitors,while little has been reported on the application of difluoromethyl（-CF₂H）in p53-MDM2 inhibitors compared to the widely studied-CF₃.Related studies have shown that the introduction of-CF₂H into spirocyclic oxindole compounds can not only serve as bioelectronic equivalents of sulfhydryl（-SH）,hydroxyl（-OH）,and amino（-NH₂）groups to optimize drug properties,but also improves the metabolic stability of the drug by enhancing the lipid solubility and bioavailability of the drug.More importantly,its extra H-bond is beneficial to improve the affinity between drug and protein targets,which is of great importance in improving protein-protein interactions as a modulator.In the present study,we developed a ketimine containing-CF₂H and successfully applied it to the reaction to obtain 45 chiralα-spiro[pyrrolidine-oxindole]analogues in relatively mild conditions with good yields（70%-99%）and excellent stereoselectivity（all dr values>20:1,ee values of 73%-99%）.All compounds were characterized and selected for in vitro antitumor activity studies on this series of compounds in colorectal cancer cells（SW620,HCT-116）,gastric cancer cells（AGS,HGC-27）and pancreatic cancer cells（Bx Pc-3）,and some of the compounds showed good antitumor activity,among which,compound 3an showed the best antitumor activity against SW620 and HGC-27,the Then,we further chemically modified compound 3an to obtain compounds 4an～6an,and the conformational studies showed that compounds5an and 6an had the best activity.Then,we further investigated the effects of these two drugs by MTT assay,cloning,scratching and Transwell migration assay using HGC-27 and SW620 as the subjects,which laid a good foundation for the next mechanism study.