Design,Synthesis And Antitumor Activity Evaluation Of Novel Chlorin E6-DCA Conjugates

Photodynamic therapy（PDT）is a relatively non-invasive cancer treatment that selectively kills cells in lighted areas.PDT utilizes light to excite photosensitizers,and performs energy transfer with molecular oxygen to generate cytotoxic reactive oxygen species,which ultimately lead to tumor cell death and tumor elimination.However,the therapeutic efficiency of PDT is limited due to the tumor hypoxic microenvironment.Ferroptosis is a mode of cell death caused by lipid peroxidation,which also produces reactive oxygen species（ROS）cytotoxicity.Therefore,the combined therapy of PDT and ferroptosis is expected to produce a synergistic effect and enhance the effect of tumor therapy.Dichloroacetic acid（DCA）acts as a pyruvate dehydrogenase kinase（PDK）inhibitor,which induces ferroptosis.In this project,chlorin e6（Ce6）was used as the parent,and different numbers of DCA groups were introduced into its three free carboxyl groups at positions 13¹,15²,and 17³.By exerting the combined effect of PDT and ferroptosis,the tumor-killing ability of the photosensitive compound is improved.In this project,three target compounds were synthesized and characterized by nuclear magnetic resonance and mass spectrometry.The photophysical and chemical properties test results show that the spectral absorption and singlet oxygen yield of the target compound are similar to those of Ce6,but the photostability is significantly enhanced.The results of in vitro antitumor evaluation showed that in Hela and MCF-7 cells,the cellular uptake rate of the target compounds was increased by 2 to 18 times,and the photodynamic activity was enhanced by 2 to 21 times,except for 8c,compared with the parent.The highest activity was found in MCF-7 cells(IC₅₀1.398μM).It was found by scratch experiment that compound 4b could significantly inhibit the migration of MCF-7 and MDA-MB-231 cells.The DCFH-DA probe was used to detect the intracellular ROS content.Compared with the blank control and Ce6,the conjugate induced increased cellular ROS level.Compounds trigger cell death through mixed apoptotic and ferroptotic pathways.When testing MCF-7 cell viability,the addition of the ferroptosis inhibitors fer-1 and DFO inhibited the cytotoxicity induced by these compounds,while the addition of the ferroptosis inducers Fe³⁺and erastin increased the cytotoxicity of these compounds.Further investigation of cellular redox status found that compound 4b reduced intracellular GSH levels by 80%and GPX4 activity by 90%.In addition,the lipid peroxidation was studied,and the level of C11-BODIPY-labeled lipid ROS was increased by 1 to 2 times,and the content of lipid peroxidation product MDA was increased by 1 to 4 times.By increasing the concentration of intracellular active ROS,depleting GSH and inactivating GPX4,it leads to peroxidation of cellular lipids,induces ferroptosis in tumor cells,and improves the inhibitory activity of tumor cell proliferation.The iron content of cells was detected by colorimetry.Compared with the blank control,the iron content of cells in the compound group increased to different degrees.Combined with the experimental results that exogenous addition of Fe³⁺can increase the cytotoxicity of these compounds,it is suggested that the enhanced activity may be related to the compounds promoting the uptake of Fe³⁺by cells,resulting in the accumulation of intracellular Fe²⁺.In summary,the conjugates of Ce6 and DCA designed and synthesized in this paper can synergistically induce tumor cell apoptosis and ferroptosis,thereby enhancing the inhibitory activity of tumor cell proliferation.At the same time,this thesis also provides a new strategy for the design of efficient antitumor active compounds.