Complexes for target killing tumor cell, preparation and application thereof
A technology for tumor cells and compounds, which is applied in the field of preparation of compounds that target and kill tumor cells. It can solve the problems of restricting the use of ricin and high toxicity, and achieve good aqueous phase dispersion, low toxicity, and good biological phase. capacitive effect
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Embodiment 1
[0031] Example 1 Preparation of MWNT-RTA complex
[0032] (1) Purification of MWNT
[0033] Put 1g MWNT in a round bottom flask, add 16ml concentrated HNO 3 , refluxed in an oil bath at 140° C. for 4 hours, centrifuged, discarded the supernatant, repeated washing and centrifugation until the supernatant pH ~ 6, and filtered with a microporous membrane with a pore size of 0.45 μm to obtain a black solid.
[0034] (2) Preparation of truncated MWNT
[0035] The purified MWNTs were further truncated by ultrasonication at 40°C for 8 hours in concentrated sulfuric acid and concentrated nitric acid with a volume ratio of 3:1, so that a large number of carboxyl groups appeared at both ends of the tube and part of the tube wall. The cut-off carbon tube dispersion is suction-filtered with a microporous membrane of 0.45 μm in diameter, washed with a large amount of distilled water until the filtrate is close to neutrality, and then the obtained filtrate is centrifuged at 9000 rpm for 1...
Embodiment 2
[0038] Example 2 Broad-spectrum killing effect of MWNT-RTA complex on numerous cell lines
[0039] (1) Cell culture
[0040] The cells used in the present invention are all preserved by the laboratory of Professor Zhong Jiang, School of Life Sciences, Fudan University. Among them, the culture medium used for human liver cell HL7702, human cervical cancer Hela cell, and African green monkey kidney cell COS-7 is 10% fetal RPMI 1640 of bovine serum; human breast cancer cell MCF-7 uses DMEM culture medium containing 10% fetal bovine serum and 1% penicillin-streptomycin. The cells used were cultured in a saturated humidity incubator with a volume fraction of 5% CO2 at 37°C, and cells in the logarithmic growth phase were selected for the experiment.
[0041] (2) Determination of cell death rate
[0042] Add 150 μL of the prepared MWNT-RTA complex to the above cells and continue to culture for 22 hours. The cultured cells were removed from the culture medium, digested with trypsin...
Embodiment 3
[0043] Example 3 Targeted Killing Effect of MWNT-RTA-anti-HER-2 Complex on Human Breast Cancer Cell MDA-MB-453
[0044] Add 250 μL MWNT (0.05 mg / mL), 50 μL RTA (2 μM) and 50 μL anti-HER-2 protein (1 μM) into 150 μL PBS (pH=7.4), stir in ice bath for 2-3 hours, at this time the toxin protein and target Proteins can be adsorbed on the surface of carbon nanotubes due to electrostatic, hydrophobic and other weak interactions.
[0045] The culture medium of human breast cancer cell MDA-MB-453 and Chinese hamster ovary cell CHO is DMEM medium. Add 150 μL of the prepared MWNT-RTA-anti-HER-2 complexes to breast cancer cells and normal cells, and continue to culture for 22 hours. The cultured cells were quickly detected by flow cytometry according to the method in Example 2(2). Column 7 in Figure 4 is the death data of two strains of cells caused by MWNT-RTA, wherein the ratio of the death rate of cancer cells to normal cells is 1:1, and MWNT-RTA-anti-HER-2 (column 8) The killing ra...
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