Design,Synthesis And Antitumor Activity Evaluation Of Small Molecule Inhibitors Derived From Naphthoquinone And Indole Structures Of Traditional Chinese Medicine

As an environment for the survival and development of tumor cells,the tumor microenvironment can not only promote the growth of tumors,but also provide favorable conditions for the invasion,metastasis and immune evasion of tumor cells.Through their own specific immune response and special metabolic regulation mechanisms,tumor cells maintain a dynamic balance in a hypoxic,low p H and nutrient-poor microenvironment,regulate their own life cycle,and proliferate indefinitely.Therefore,targeting the tumor microenvironment will block the nutrient acquisition and energy source of tumor cells,thereby preventing the occurrence and development of tumors.Among them,STAT3,a tumor immune target,and PHGDH,a tumor metabolic target,are closely related targets to the tumor microenvironment.1.Design,synthesis and biological activity evaluation of STAT3 inhibitors derived from naphthoquinone structure of traditional Chinese medicineAberrant activation of signal transducer and activator of transcription 3（STAT3）plays an important role in the development of tumors.In order to find novel and effective STAT3 inhibitors,we used a ring-opening strategy to structurally engineer BBI608,an active monomer with naphthoquinone structure isolated from the traditional Chinese medicine,to obtain a novel class of STAT3 small molecule inhibitors with backbone.Compound B31 and positive drug BBI608 had equivalent inhibitory activity.Through more systematic in vitro and in vivo experimental studies,compound B31 could inhibit the proliferation of breast cancer MDA-MB-231,MDA-MB-468 and hepatoma Hep G2 cells with IC₅₀of 0.67±0.02μM,0.77±0.01μM and 1.24±0.16μM,respectively.Preliminary mechanistic studies showed that compound B31 was able to selectively inhibit the phosphorylation of Try705 in STAT3 protein,which in turn inhibited the expression of downstream target proteins Cyclin D1 and C-myc,without affecting the phosphorylation of STAT1 protein.Flow cytometry showed that compound B31 could promote breast cancer cell apoptosis and block S-phase cell replication.In vivo experiments demonstrated that compound B31significantly inhibited the growth of tumors at a low dose of 10 mg/kg and showed good antitumor activity without significant toxic side effects.Molecular docking studies further elucidated the binding mode of B31 at the STAT3 SH2 domain.This study not only further enriches the diversity based on structural modification of BBI608,but also provides more valuable reference for the development of more natural STAT3 small molecule inhibitors.2.Design,synthesis and structure activity relationship of PHGDH inhibitors based on indole structurePhosphoglycerate dehydrogenase（PHGDH）is a key enzyme of intracellular serine synthesis pathway,which is highly expressed in many malignant tumor cells and has become a key drug target for anti-tumor therapy in recent years.In view of the reported indole-competitive PHGDH inhibitors,3D-QSAR,molecular docking and molecular dynamics methods were used to carry out a more systematic theoretical study at the molecular level,the relationship between structure and activity was discussed,and the necessary conditions for the binding of inhibitors to proteins were explored to provide a theoretical basis for guiding the discovery of new compounds.In addition,we constructed Co MFA and Co MSIA models that are reliable and have good predictive ability,and in the color block diagram,electrostatic and hydrogen bond fields were found to have a large effect on the activity.Molecular docking and kinetics revealed that hydrogen bonding between the compound and key residues Asp174,Gly153,Arg154,Ile155,Gly156 and hydrophobic interaction with residues Leu150、Pro175、Leu192、Thr212、Leu215 could improve the binding stability of the ligand to the protein,and at the same time,hydrogen bonding with residue Asp174was essential for the compound to produce activity.The R1 region should introduce groups with hydrophobic properties and a flat structure,and substituents can introduce halogen atoms.R2 is a key region affecting the strength of the activity,where the group should be positively charged and have a large polarity and can act as a hydrogen bond donor or acceptor.Based on the above results,a compound ZINC56603336 was found in the ZINC database using virtual screening technology,and through structural optimization,synthesis,and enzyme activity tests,it was found that compound E26 could be used as a seedling compound for subsequent studies.The results enrich the molecular level theoretical study of competitive PHGDH inhibitors and provide important theoretical guidance and reference value for the discovery of novel PHGDH inhibitors.