Study On The Screening, Identification And Anti-tumor Activity Of Inhibitors Of APE1

APE1is a multifunctional protein and a paradigm of biological macromolecularmultifunctional complex, which has functions such as DNA base excision repair and redoxactivity. APE1plays an importment role in in the repair of oxidative DNA damage andtranscriptional regulation. It is often overexpressed and cytoplasmic expressed in tumortissues,and considered as an important marker of maligant tumors.Inhibiting APE1significantly increases tumor cell killing and sensitizing to DNA damage challenge.Therefore, APE1could be a novel cancer therapeutic effective target which cutribute tocisplatin-based chemotherapy. Recent findings on redox inhibition of APE1have potentialclinical translational significance such that a redox inhibitor could be used as a single agent,in combination with current treatments or as a potential anti-growth, cytostatic agent. Anumber of small molecules have been described as Ape1inhibitors; however, thosecompounds are in the early stages of development. This study intend to develop morespecific and powerful small molecular redox inhibitor of APE1by virtual screening fromnature products in Chinese traditional medicine.The present study will provideexperimental foundation for cancer therapy targeted to APE1.Objectives:To Screen and identify inhibitors of APE1and investigate their anti-tumor activities.Methods:1. Discovering small-molecule inhibitors of APE1from nature products in Chinesetraditional medicine by computer virtual screening based on structure of APE1.2. Screening and identification small-molecule inhibitors of APE1from98small-molecule compounds using electrophoretic mobility shift assay3. To test inhibition of AP endonucleases by compounds, an oligonucleotide cleavageassay designed to monitor the cleavage of substrate to product through electrophoreticseparation was utilized. 4. Interactions between compounds and APE1were investigated using the dualpolarization interferometry.5. To investigate whether gossypol alone or combination with the clinically relevantagent DDP can inhibit tumor growth in vitro and in vivo, we tested effects of gossypolwith CCK-8array and xenograft assays.Results:1.61compounds were found that had potential inhibition to APE1,5of them werespecial to repair function,and56of them special to redox function.2. We have demonstrated that tanshinoneâ…¡ A, gossypol and ebselen effectivelyinhibit redox activities of APE1through a direct interaction using electrophoretic mobilityshift assays, AP endonuclease assays, dual polarization interferometry,and gossypol caneffectively inhibit both the repair and redox activities.3. Gossypol more effectively kills cancer cells and enhanced the killing effect of theclinically relevant agent DDP in vivo and in vitro.Conclusions:1. Tanshinoneâ…¡A and ebselen are novel inhibitors of redox function of APE1.2. Gossypol is a novel inhibitor of double-function of APE1, and it alone orcombination with the clinically relevant agent DDP can inhibit tumor growth in vitro andin vivo.