Apoferritin-benzothiazole compound complex as well as preparation method and application thereof

A kind of technology of apoferritin and benzothiazole, applied in the field of medicine

Active Publication Date: 2018-12-18
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Ineffective against normal breast cells and l

Method used

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  • Apoferritin-benzothiazole compound complex as well as preparation method and application thereof
  • Apoferritin-benzothiazole compound complex as well as preparation method and application thereof
  • Apoferritin-benzothiazole compound complex as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1: Preparation of apoferritin (Apoferritin, AFt)

[0039] Apoferritin is the iron-free form of ferritin, which is composed of 24 protein subunits and is a natural iron storage protein; in order to obtain apoferritin, we need to dialyze ferritin.

[0040] (1) Prepare dialysis buffer: dissolve sodium acetate in water at room temperature, adjust the pH to 5.5 with hydrochloric acid, and make a sodium acetate dialysis buffer with a concentration of 0.1mol / L;

[0041] (2) Prepare the dialysis reaction: put 1.5L of dialysis buffer into a 2L beaker, add 1.95mL of thioglycolic acid drop by drop in a dark environment, and then slowly fill with nitrogen for 2 hours before use;

[0042] (3) Prepare ferritin: Dilute ferritin (SIGMA, CAS: 9007-73-2) 10 times with 0.1mol / L sodium acetate dialysis buffer, and put it into a dialysis bag wetted with sodium acetate dialysis buffer in advance (MWCO 12000-14000), immerse the dialysis bag in the beaker containing the sodium acet...

Embodiment 2

[0053] Example 2: The steps of this example are the same as in Example 1, except that step (6) prepares the coating reaction: dilute the apoferritin mother solution with the coating buffer to a final concentration of 2 mg / mL, take 10 mL and put it into the pre-used In a dialysis bag (MWCO 12000-14000) wetted with coating buffer, put it into a beaker containing coating buffer, and place it in the dark for 12 hours at 4°C for dialysis; in addition, use coating buffer to prepare a final concentration of 10mM Benzothiazole compound solution and add the sodium hypochlorite that final concentration is 0.02mmol / L, stand-by, wherein the mol ratio of apoferritin and benzothiazole compound is 1000:1; Benzothiazole compound is respectively 2-(4-amino -3-methylphenyl)-5-fluorobenzene (compound 1), 2-(4-methoxy-3-methoxyphenyl)-5-fluorobenzene (compound 2), L-lysine -(4-(5-fluorobenzo[d]thiazol-2-yl)-2-methoxy)-phenyl ester (compound 4);

[0054] Table 2 Loading efficiency (%) of apoferri...

Embodiment 3

[0056] Embodiment 3: The compound of embodiment 1 and embodiment 2 inhibits tumor growth experiment

[0057] Tumor cells MKN-45, NCI-N87, KATO-III, SGC-7901 and BGC-823 cell lines were cultured in RPMI-1640 medium containing 10% fetal bovine serum at 37°C and 5% CO2 box.

[0058] Take the above cells in the logarithmic growth phase (cell confluency 80%) for experiments; plant 5000-6000 cells in a 96-well plate, set up a drug-dosing group and a control group in each well plate, and add the concentration of the reagent in terms of the compound For 0.001 μM, 0.01 μM, 0.1 μM, 1 μM and 50 μM, place the 96-well plate in the incubator and incubate for 72 h, then add 25 μL of 5 mg / mL thiazolyl blue (MTT) into the incubator and incubate for 4 h. After the MTT incubation, take out the 96-well plate and carefully remove the liquid in the 96-well plate with a vacuum pump to avoid sucking away the blue formazan crystals formed by the MTT incubation at the bottom; after pipetting, add 150 ...

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Abstract

The invention discloses an apoferritin-benzothiazole compound complex as well as a preparation method thereof. Liquid ferritin is loaded into a dialysis bag, the dialysis bag is placed in mercaptoacetic acid-containing sodium acetate dialysis buffer liquid with pH of 4.5 to 5.5 for dialysis, in a dialysis process, nitrogen is introduced continuously to maintain an oxygen-free environment, the mercaptoacetic acid-containing sodium acetate dialysis buffer liquid with the pH of 4.5 to 5.5 is replaced every 20-25 min until the liquid in the dialysis bag is colorless, after the liquid in the bag issubjected to sterilize filtering with a 0.22 mu m filter, apoferritin mother liquor is prepared, and a BCA method is used for measuring the apoferritin mother liquor concentration; the apoferritin and a benzothiazole compound are subjected to an in-situ production method to make the benzothiazole compound loaded into the apoferritin, so that the apoferritin-benzothiazole compound complex is obtained. The prepared complex has a potential targeted anti-tumor effect.

Description

technical field [0001] The invention belongs to the field of medicine, and relates to a preparation method of a benzothiazole compound compound loaded with apoferritin and its application in targeted antitumor. Background technique [0002] Drug delivery system is one of the hotspots in tumor research, and the research and application of tumor-targeted drug delivery system is a part that has attracted much attention in the field of cancer treatment research. Apoferritin is the iron-free form of ferritin, composed of 24 protein subunits, and is a natural iron storage protein. It is known that a large number of ferritin-specific receptors are expressed on the surface of many tumor cells, which can mediate the endocytosis of ferritin and apoferritin through the binding of ferritin receptors and ferritin (Zhang, L. et al. Receptor- mediated cellular uptake of nanoparticles: a switchable delivery system. Small 2011, 7 (11), 1538−41.). Highly expressed ferritin-specific receptor...

Claims

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Application Information

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IPC IPC(8): A61K47/64A61K31/428A61P35/00
CPCA61K31/428A61K47/644A61P35/00
Inventor 张继虹杨帆罗瑛王玉玲俞飞段文芳宋斌苏永南
Owner KUNMING UNIV OF SCI & TECH
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