Design, Synthesis And Biological Applications Of H 2 Se Near-infrared Fluorescent Probes

There are several therapeutic methods for cancer in recent years such as surgical treatment,chemotherapy, radiotherapy and more. The chemotherapy is that cancer patients are treated with some anti-cancer medicine which could inhibit cancer cell proliferation or initiate cancer cell apoptosis pathway. Different kinds of anti-cancer medicine could effectively kill cancer cells at each phase of the cell cycle.Sodium selenite becomes the focus of anti-cancer medicine because of economy, low toxicity and effectivity. Sodium selenite could inhibit tumor cell proliferation and induce tumor cell apoptosis, but the mechanism of inducing tumor cell apoptosis is not really clear. A lot of research work on sodium selenite were carried out under normoxic condition. Hydrogen selenide generated by the metabolic process of sodium selenite could react with oxygen to produce a large number of reactive oxygen species?ROS?. Thus it is generally accepted that tumor cell apoptosis induced by sodium selenite is due to oxidative stress. However it is well known that solid tumor microenvironment is hypoxic condition, previous research work about sodium selenite were carried out under normoxic condition. So we designed our research work under the 1% oxygen concentration condition, novel small-molecule fluorescent probes were used to monitor hydrogen selenide and active oxygen during sodium selenite induced tumor cell apoptosis. We observed the redox state of tumor cell during sodium selenite-induced tumor cell apoptosis and tried to provide more accurate research conclusion for the study of sodium selenite anti-cancer mechanism.Our research work mainly include the following two parts:1. We developed the design, synthesis and biological application of the near infrared fluorescent probe NIR-H₂Se which can image the endogenous hydrogen selenide at the first time. The merocyanine dye acts as fluorescent matrix of the near infrared fluorescent probe NIR-H₂Se and the Se-N bond structure is designed as probe report group while the Se-N bond structure can specifically react with structure of-HSe. The near infrared fluorescent probe NIRH₂Se could image hydrogen selenide instantaneously, high selectively and high sensitively in living cell and in vivo. Near infrared fluorescent probe NIR-H₂Se can avoid the interference of spontaneous background fluorescence and reduce the damage of tissue. The novel small-molecule fluorescent probe NIR-H₂Se is used to monitor the content of hydrogen selenide during sodium selenite induced HepG2 cell apoptosis under the 1% oxygen concentration condition and applied in tumor bearing mice imaging successfully. This probe should serve as a powerful tool for exploring the sodium selenite anti-cancer mechanism.2.On the basis of the first half of our research work about sodium selenite we further explored the redox state of tumor cell during the sodium selenite-induced apoptosis under hypoxic condition. We used probe NIR-H₂Se and commercial probe DHE to monitor the intracellular hydrogen selenide and superoxide anion under the 1% and 5% oxygen concentration condition, meanwhile added rhodamine 110 into the simultaneous detection system as reference fluorophore and depended on the fluorescence ratio method to eliminate the interference of external factor. The emission of rhodamine 110 fluorescent dye is 532 nm which can effectively be separated with the emission 638 nm of probe DHE reaction product and the emission 735 nm of probe NIR-H₂Se, so as to achieve three optically independent channels at the same time. The molecular structure of the reaction products of two probes and reference fluorescent dye are similar because of containing two amino groups, thus this advantage improve the sensitivity of this fluorescence ratio method.Our research showed the higher content of superoxide anion yet selenium hydrogen content is low under normoxic condition. Under hypoxic condition hydrogen selenide cannot react with oxygen to produce a large number of superoxide anion rapidly due to lacking of oxygen, so hydrogen selenide accumulate and superoxide anion content is low. These findings indicate that under hypoxic condition the anti-cancer mechanism of sodium selenite maybe via non-oxidative stress.