Photoaffinity-based Proteome Profiling To Identify Cellular Targets Of Betulinic Acid And ABT-199

Modern drug discovery mainly includes the identification of target,the discovery of lead compounds,structural modification and clinical research and other stages.Accurately,confirming the target of active molecules is one of the most important stage to affect the whole process of research.The betulinic acid extracted from natural medicinal birch is a natural drug molecule with many pharmacological effects and has good antitumor activity and very low biotoxicity.In this work,we mainly use the PAL-ABPP?Photoaffinity-based proteome profiling?technology to synthesize some active based betulinic acid probe molecules BA-1,BA-2,BA-3.And around these small molecule probes,we do a variety of biological target identification and validation.The work on this section mainly includes the following parts:1.Two kinds of modified parts of betulinic acid were constructed by using betulinic acid BA as the parent compound.The molecular probe BA-1/2 with diazirine and tetrazole as photoaffinity groups and a fluorescent molecular probe BA-3 carrying coumarin group were synthesized.2.The inhibition of tumor cell proliferation was carried out on the three probes BA-1/2/3,and the bioactivity of the molecular probe was confirmed by reference of the parent drug betulinic acid.The results showed that IC₅₀ of BA and BA-1/2/3 on MCF-7 cell line was 10.5?M,11.6.?M,8.7?M,15.0?M,respectively.3.In situ labeling profile was carried out by probe molecule BA-1/2 to verify the availability of probes study on betulinic acid.With the above molecular probes,we carried out cell imaging experiments to determine the drug molecules in the cell sub-structural distribution.After confirming the biological activity of the molecular probe,we carried out the Bio-Mass?LC-MS?experiment to identificate the sample of pull down.In addition to the confirmation of the biological target of betulinic acid,we also carried out the biologically side-effect work and target verification for the currently listed Bcl-2 inhibitor ABT-199.With this work we can more conveniently understand the in vivo mechanism of these drugs and generation mechanism of side effects to provide the basic information.It has been confirmed that some molecular probes are effective,the biological mass spectrometry verification work is still in progress.