Targeting Mammalian Thioredoxin System By Small Molecules Based On Phenylarsenoxide(PAO) Scaffold

Arsenic trioxide(As₂O₃),which is a traditional Chinese medicine,has attracted worldwide interest because of its relatively safe and effective treatment for relapsed and refractory acute promyelocytic leukemia(APL).The success of As₂O₃ in APL treatment has revived research on arsenic-based drugs.Thioredoxin system is a novel target for cancer chemotherapy,which has an important relationship with cancer.Inhibition of thioredoxin system and alteration of cellular redox homeostasis are promising in treatment of cancer.We designed and synthesized a series of PAO derivatives,and evaluated their anticancer activity in cellular models.Considering the important role of the thioredoxin system,the development of a more convenient method for determinating redox states of Trx R/Trx is a research priority.The thioredoxin system is composed of thioredoxin reductase(Trx R),thioredoxin(Trx),and NADPH.The C-terminal active site of mammalian Trx R shares a unique sequence of-Gly-Cys-Sec-Gly while the active site of Trx contains a sequence of-Cys-Gly-Pro-Cys-.Both can be regarded as closely spaced vicinal dithiol,which can specifically bind with PAO.So a series of PAO derivatives was developed to determin the redox states of Trx R/Trx.Protein vicinal dithiols play fundamental roles in redox regulation through the reversible vicinal dithiol oxidation to disulfide.Thus,vicinal-dithiol-containing proteins(VDPs)are attracting more and more attention.Since trivalent arsenicals selectively interact with protein vicinal dithiols,we developed a probe to recognition of VDPs based on fluorescence resonance energy transfer(FRET)mechanism.In summary,based on the high affinity between PAO and vicinal dithiols,we explored anticancer activity of PAO and its analogues via targeting Trx R in cells,and developed novel probes for determination of redox states of Trx R/Trx and for recognition of VDPs.The main contents are summarized as follows:Chapter 1.This chapter reviewed therapeutic and analytical applications of arsenic compounds,including cancer chemotherapy,molecular imaging and protein purification.Specific examples including arsenic-based drugs,the imaging of cellular events using arsenic-based dyes and the affinity purification of arsenic binding proteins are given.In addtion,the structure and function of thioredoxin system were introduced,and strategies for determining redox states of Trx R/Trx were discussed.Chapter 2.This chapter described the synthesis,biological evaluation of PAO and its derivatives.One of the derivatives,PAO-PDT,displays highest cytotoxicity toward human HL-60 cells among all tested cell lines.The action mechanism of PAO-PDT was studied.The simplicity of the structure and the presence of an easily manipulated amine group will facilitate the further development of PAO-PDT as a potential cancer chemotherapeutic agent.Chapter 3.With PAO as a recognition group,we synthesized fluorescence-labeled PAO(Nap-PAO),immobilized PAO(Seph-PAO)and biotinylated PAO(Biotin-PAO).Nap-PAO can determine redox states of Trx in vitro and in cells while Biotin-PAO can determine redox states of Trx R in vitro and in cells.Chapter 4.We developed a probe COAzo B-PAO based on fluorescence resonance energy transfer(FRET)mechanism for exogenous VDPs and endogenous VDPs.