Design,Synthesis And Antitumour Activity Of 6-(Imidazo[1,2-a]pyridin-6-yl) Quinazoline Derivatives

Objective:Malignant tumors are one of the major diseases that threaten human lives in the world today.Research on molecularly targeted drugs has become a hot topic in drug development,and these drugs are expected to have better efficacy and lower toxic side effects.Quinazolines are a class of nitrogen-containing heterocyclic compounds with good biological activities,such as anti-inflammatory,antibacterial,anti-human immunodeficiency virus（HIV）,and anti-cancer effects.In this work,a series of6-（imidazo[1,2-a]pyridin-6-yl）quinazoline derivatives were designed and synthesized based on a selective inhibitor of 4-aminoquinazoline PI3Kαidentified in the previous study 6b,combined with a backbone combination strategy,and their preliminary activity studies were carried out.In order to discover novel antitumor lead compounds with safe and effective physical and chemical properties.Methods:Main research methods:1.Based on the PI3Kαinhibitor 6b synthesized in the previous study,the 4-aminoquinazoline structure of compound 6b was analyzed based on the structure-activity relationship,in which the 4-aminoquinazoline structure of compound 6b was the key pharmacodynamic group for inhibiting PI3Kαactivity.Therefore,we identified this pharmacodynamic group as one of the basic backbones of the target compounds.It was shown that imidazo[1,2-a]pyridine is the key pharmacodynamic group of PI3Kαinhibitor HS-173,and based on the skeleton association strategy,we introduced it into the C⁶position of 4-aminoquinazoline by Suzuki–-Miyaura cross-coupling reaction and modified its structure to explore its conformational relationship.2.The molecular structures of all compounds were characterized by ¹H-NMR,¹³C-NMR,¹⁹F-NMR,and HR-MS.3.Test the IC₅₀values against various cancer cell lines,including HCC827（human non-small cell lung cancer cells）,A549（human non-small cell lung cancer cells）,SH-SY5Y（human neuroblastoma cells）,HEL（Human red leukocytes leukemia cells）,and MCF-7（human breast cancer cells）,using the thiazole blue method（MTT）.4.Molecular docking simulation by MOE（Molecular Operating Environment）.5.Detect kinase activity by ADP-Glo ^TMMax Assay.6.Verify the cycle,apoptosis of target compounds by flow cytometry.7.Verify the effect of target compounds on PI3K signaling pathway,other target protein pathways,and cell cycle by immunoblotting.8.Analyze the long-term efficacy of compounds using 3D cell culture assay.Results:A series of 32 6-（imidazo[1,2-a]pyridin-6-yl）quinazolines were synthesized,of which 21 were derivatives 10a-u containing ester substituents and 11 were derivatives13a-k containing amide substituents.The results of anti-proliferative activity tests showed that most of the compounds had good proliferation inhibitory effects.For example,compounds 10b,10s,13a,13f and 13k showed IC₅₀values less than 1μM against HCC827 cells,especially compound 13k showed an IC₅₀value of 0.09μM against HCC827 cells,and 13k inhibited the proliferation of HCC827 cells in a time-dependent manner.In addition,13k showed good inhibitory activity against the other four tumor cell lines tested with IC₅₀values of 0.18-0.43μM,meanwhile the IC₅₀of 13k against normal cells MRC-5（human embryonic lung fibroblasts）was 1.98μM,indicating that the compound is selective for tumor cells.Molecular docking simulations showed that compound 13k could stably bind to the ATP-binding pocket of its target protein PI3Kα.In addition,13k induced G2/M phase block and eventually apoptosis in HCC827 cells by effectively inhibiting the kinase activity of PI3Kα(IC₅₀value of 1.94 n M).Immunoblot analysis showed that compound 13k inhibited PI3K and its downstream AKT/MAPK signaling pathway.In addition,we found that compound 13k was able to reduce the protein levels of BRD4 and its related downstream proteins c-Myc and various RTKs（such as Er BB2 and c-Met）as well as their phosphorylation levels.The related cyclins cyclin B1,CDK1,and p-CHK1 were found to be reduced in a dose-dependent manner in cells treated with compound 13k,and the related apoptotic proteins cl-caspase9 and cl-PARP were correspondingly upregulated.Conclusions:In this thesis,we designed and synthesized a series of6-（imidazo[1,2-a]pyridin-6-yl）quinazoline derivatives with promising antitumor activity.Preliminary activity studies revealed that compound 13k could further inhibit cell proliferation and promote apoptosis by inhibiting the signaling pathways of PI3K and BRD4.It provides a lead compound for the design and development of new targeted antitumor drugs.