Expression Of Aromatase(CYP19) In Pichia Pastoris And Synthesis Of The Aromatase Inhibitors

Breast cancer,as the most common cancer among women worldwide,it continues to be an main cause of female mortality.In China,because of the changes in living environment and Lifestyles,the incidence of the breast cancer turns to rise rapidly.Research found that almost two-thirds of breast cancer patients are estrogen-dependent,and we can inhibit the development of breast cancer by reduced the estrogen content.Aromatase,otherwise called CYP19,is a rate-limiting enzyme in the biosynthesis of estrogens.Reduction of aromatase and deprivation of estrogens can inhibit the growth of breast cancer cells and extend the patients’ lifespan.That makes aromatase to the ideal target in the research of pharmacotherapy for breast cancer.Nowadays,Aromatase inhibitors(Als)have become the first-line drugs for hormonally-responsive breast cancer in postmenopausal women.However,with the wide application of AIs in clinical practice,some problems gradually show up,such as non-response in many patients and resistance to AIs treatment.Hence,there is a need to discover and develop the new generation of AIs to overcome these defects.In the research of the aromatase inhibitors,the aromatase which is used to determination of reactivity are extract from the placenta or the AIs test kit.The source of placenta was restricted and AIs test kit is very expensive,so we need to find a new way to obtain the aromatase.In this study we describe the expression of a recombinant derivative of P450arom in pichia pastoris of which the first 45 amino acids were deleted from the N-terminus.The plasmid pPIC 9K-aromatase was constructed by DNA recombination and the recombinant protein was expression in the pichia pastoris.This recombinant protein has activity similar to that of the wildtype protein.Based on the analysis of 45 aromatase inhibitors which have high inhibitory activity a common feature Pharmacophore model was established,15 compounds were designed and synthesized.The structures of all compounds and the intermediate compounds have been characterized by 1H NMR and 13C NMR and the inhibitory activity test was performed.It is noteworthy that solvent,temperature and base of the reaction were optimized,and the ideal reaction condition was found.The test result shows that 2 of the 15 compounds are of good inhibitory activity which are compound 2-4(IC50=1.009 μM)and compound 2-5(IC50=l.703μM),on the whole,the 3H-2-Benzopyran-3-one ramification have good inhibition activity of the aromatase.AIs based on a 3H-2-Benzopyran-3-one scaffold have never been reported before.The discovery of new compounds based on a novel scaffold will help in discovering the next generation of AIs.