Design,Synthesis And Evaluation Of The Hypoxic Responsed Antitumor Prodrug

Chemotherapy is one of the important means of cancer treatment,however,the existing anti-cancer drugs generally have great side effects,in order to reduce these side effects,a large number of novel prodrugs have been developed.Diagnostic and therapeutic prodrugs,which have the function of both diagnosis and treatment,achieve a controlled release of drugs.Also,the released active drugs will be tested,so that the treatment effect can be timely feedback.Organic small molecule prodrugs have the characteristics of simple structure,easy modification and good biocompatibility,becoming a hot spot for research.The vast majority of cancer is the solid tumor.Hypoxia is one of the important features of solid tumors,leading to a situation that the tumor is not sensitive to chemotherapeutic drugs and produce drug resistance.However,tumor hypoxia also provides new idea for the treatment of cancer,that is,using the hypoxia reactive groups to modify the active drug,so that it can be activated only in the hypoxic region of the tumor,then release active drugs.Floxuridine?FDU?is an anti-metabolic anti-tumor drug,used for the clinical treatment of gastric cancer,breast cancer,and so on.But the side effects?such as nausea,vomiting,etc.?it caused limit its application.In order to reduce the side effects of FDU in the course of tumor therapy and to detect the distribution and accumulation of drugs in vivo,in this text,using nitrobenzyl as the hypoxia reaction group and 2-carboxy-2'-hydroxybiphenyl as a fluorescent dye precursor,coupling them with the active drug FDU by covalent bond,then the hypoxia reactive diagnostic prodrug FDU-BCM-NO₂ was designed and synthesized.The synthesized prodrug and intermediates were characterized by NMR,MS and IR.The reducing components of prodrug were identified by HPLC,and the drug release mechanism was explored.The fluorescence intensity of the prodrug changing with the reduction time was detected by fluorescence spectroscopy.MTT assay was used to detect the cytotoxicity of FDU and prodrug on MCF-7,MGC-803 tumor cells and BRL-3A hepatocytes.Flow cytometry was used to test the apoptosis of cancer cells induced by prodrug,and the in vitro therapeutic effect of prodrug was discussed.The images of hypoxic tumor cells and tumor globules incubated with prodrug were obtained by fluorescence microscopy,then they were used to study the diagnosis of prodrug and the tracing effect on FDU.The results showed that the prodrug was reduced under the condition of hypoxia to release the active drug FDU and the fluorescent molecule BCM.The release of the FDU can be monitored using the fluorescence signal from BCM.The longer incubation time in hypoxia condition,the more release of FDU.When MCF-7 cells,MGC-803 cells and BRL-3A cells were incubated with FDU under normoxic conditions for 48 h,cell survival rate were 41.43%,41.70% and 48.09%.When MCF-7 cells,MGC-803 cells and BRL-3A cells were incubated with FDU-BCM-NO₂ under normoxic conditions for 48 h,cell survival rate were 91.09%?91.22% and 99.08%.When MCF-7 cells and MGC-803 cells cells were incubated with FDU under hypoxia conditions for 48 h,cell survival rate were 22.57% and 41.70%.When MCF-7 cells and MGC-803 cells cells were incubated with FDU-BCM-NO₂ under hypoxia conditions for 48 h,cell survival rate were 44.76% and 51.27%.FDU has a cytotoxic effect on both normal cells and tumor cells,while the prodrug have hypoxia selectivity,only kill the hypoxic tumor cells,the normal cells will be safe.FDU-BCM-NO₂ can induce apoptosis of tumor cells,when MGC-803 cells were incubated with FDU-BCM-NO₂ under hypoxia conditions for 24 h,52.21% cells were induced apoptosis.BCM released from prodrug in the hypoxic condition has two-photon excitation characteristics,the fluorescence signal can be used for tumor cells and tumor globules hypoxia lever and drug release detection.Prodrug FDU-BCM-NO₂ has the dual function of diagnosis and treatment,the release of FDU can be controlled by using the hypoxic nature of solid tumors.This method successfully avoid the undesirable side effects on normal cells,provides a new way of thinking for the individualized treatment of cancer.