Synthesis,Evaluation And Proteomic Analysis Of Protac Based On Parthenolide

Parthenolide,extracted from the western plant asteraceae(Daisy),is a sesquiterpene lactone natural product,which was first used in Europe to relieve fever,migraine,rheumatoid arthritis and other diseases.It is a natural product with a long history of medicinal use.However,the content of parthenolide in daisies is low,which makes it difficult to develop and utilize.The researchers found that parthenolide content was as high as 9.6 percent in the root bark of a plant endemic to China.Xinyi is a traditional Chinese medicinal material.It has a strong taste,a warm nature,a return to the lung and stomach meridian,and a function of dispelling wind cold and clearing nasal passages.It is used for wind cold headache,nasal congestion and runny nose,nasal deep,etc.The earliest written records can be traced back to Shennong Baicao Classic.Because of this,parthenolide is of great interest to chemists and biologists.Existing studies have shown that parthenolide has anti-inflammatory,anti-tumor and anti-leukemia pharmacological activities.It can act on FAK1,NF-κB,DNMT1 and Wnt/β-catenin signaling pathways and functional proteins,but the exact biological targets are still to be clarified systematically.Therefore,this study aimed to modify the structure of parthenolide,optimize its biological activity,and explore its interacting proteins by combining proteomics.Protein degradation via proteolysis-targeting chimeras(PROTAC)provide a new strategy for target discovery of natural products,which can be used to explore the panorama of protein changes in cells through proteomic investigation to analyze their potential targets.Based on this idea,this study used parthenolide as the parent structure,selected PEG,alkyl,saturated heterocyclic fragments linkers,designed and synthesized20 parthenolide degraders,confirmed their structure by MS and ~1H NMR.The antitumor proliferative activity of all parthenolide degraders in vitro was determined by SRB method.The results showed that compounds K1,K9,K10,K15,K16,K17,K18 and K20 showed better biological activity on colon cancer cells HCT-116 and HT29 than FT671.We further selected the compound K10 with the best anti-cell proliferation activity to carry out proteomic experiments,and preliminarily explored interacting proteins of parthenolide.The results showed that the saturated heterocyclic fragment chains with strong rigid structure could significantly enhanced the anti-cell proliferation activity of parthenolide degraders.The Alkyl chains was more conducive to the inhibitory activity of parthenolide on HCT-116 and HT29 cells.We selected K10,the most active compound in anti-cell proliferation,to carry out proteomic experiments,and preliminarily explored interacting proteins of parthenolide.165 up-regulated differentially expressed proteins(DEPs)and139 down-regulated DEPs including CYP24A1,DHX36,RPS10,DNMT1 and CLK3were identified.Online enrichment analysis of all down-regulated proteins showed that their functions were concentrated in mitochondria,ribosomes and RNA-binding regions.These findings suggested that parthenolide may have a proliferative inhibitory activity on tumor cells by influencing intracellular energy metabolism and provided novel ideas for the discovery of parthenolide targets.