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125i labeled caerin polypeptide and its application

A labeling and polypeptide solution technology, applied in the biological field, to achieve good in vitro stability, good radiochemical purity, and high labeling rate

Active Publication Date: 2019-02-19
WNL生物医学科技公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Existing studies have shown that F1 and F3 polypeptides have inhibitory effects on the growth of HPV-transformed cells (TC-1 cells) and human cervical cancer cells (Hela cells) in vitro, but the effect of polypeptides on different tumor cells has a concentration-effect relationship. The inhibitory effect of high-concentration peptides on different tumor cells is dose-dependent. As the concentration of peptides increases, the inhibitory effect is gradually enhanced. Medium and low-concentration peptides have no direct killing effect on tumor cells, no obvious activation effect, and no time dependence. and dose-dependent

Method used

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  • 125i labeled caerin polypeptide and its application
  • 125i labeled caerin polypeptide and its application
  • 125i labeled caerin polypeptide and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1125I

[0042] Example 1 125 Preparation of I-labeled F3 polypeptide

[0043] S1. Take lodogen, add chloroform to dissolve it at a solid-to-liquid ratio of 1 mg / mL, take 100 μL and add it to the bottom of the reaction tube, dry it under negative pressure, seal it, and store it at -15°C to -20°C to prepare a reaction coated with lodogen. Tube;

[0044] S2. Take the reaction tube coated with lodogen prepared in step S1, and add 40 μL of F3 polypeptide solution with a concentration of 0.4 μg / μL and 20 μL of Na with a concentration of 25 μCi / μL in sequence. 125 1, stirring reaction 15min under the condition of 25 ℃ at temperature;

[0045] S3. After the reaction time is over, immediately add 300 μL of phosphate buffer solution with a concentration of 0.05 mol / L and a pH of 7.4 to terminate the reaction, and let stand for 5 minutes to obtain a reaction solution;

[0046] S4. Purify the reaction solution described in step S3 with a SephadexG-25 chromatography column, elute with a phospha...

Embodiment 2125I-F3

[0050] Example 2 125 I-F3 Stability Investigation

[0051] (1) take 125 I-F3, placed under the condition of 4°C for 60h, measured respectively after 0h, 5h, 10h, 15h, 20h, 25h, 30h, 45h, 60h, 125 The radiochemical purity of I-F3, see image 3 . Depend on image 3 The results show that the present invention provides 125 After I-F3 was stored at 4°C for 60 hours, the radiochemical purity was still as high as 98.25%.

[0052] (2) take 125 I-F3, placed under the condition of 25 ℃ for 24h, measured respectively after 2h, 4h, 8h, 12h, 16h, 20h, 24h, 125 The radiochemical purity of I-F3, see Figure 4 . Depend on Figure 4 The results show that the present invention provides 125 After I-F3 was stored at 25°C for 24 hours, the radiochemical purity was still as high as 98.01%.

[0053] (3) Take 50 μL of 125 I-F3 was added to 100 μL of normal saline and 100 μL of human plasma, and placed at a temperature of 37°C for 48 hours. After 10 minutes, 20 minutes, 30 minutes, 1 hour...

Embodiment 3125I-F3

[0054] Example 3 125 Inhibitory effect of I-F3 on the growth of MCF-7 cells in vitro

[0055] Take the MCF-7 cells (human breast cancer cells) in the logarithmic growth phase, adjust the cell concentration to 1×10 with RPMI1640 medium containing 10% volume fraction calf serum 5 / mL, seeded in a 96-well plate, 100 μL per well, and 6 parallel wells for each group. After inoculation, different concentrations of 125 I-F3 medium 10 μL / well, 125 The radioactivity of I-F3 was 1, 10, 37, 100, 200 and 500kBq / well respectively, and the control group Na 125 The I radioactivity is 1, 10, 37, 100, 200, 500kBq / well, and the blank group is an equal-volume culture medium without cells, placed at 37°C and 5% CO 2 After culturing in the incubator for 24 and 48 hours, add 10 μL / well of MTT (5 mg / mL), continue to incubate for 4 hours, then terminate the culture, add 10% SDS-HCl 100 μL / well, and act at 37°C for 12 hours. A wavelength of 570nm was selected, and the optical absorption value (OD...

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Abstract

The invention belongs to the technical field of biology and provides a 125I-labeled Caerin polypeptide and an application thereof to preparation of a drug for treating tumors and particularly benign and malignant tumors relevant to HPV infection. The 125I-labeled Caerin polypeptide provided by the invention is prepared by using an optimized direct labeling method, and the preparation method is simple and convenient to operate, high in labeling rate, short in reaction time and good in in-vivo and in-vitro stability. The 125I-labeled polypeptide prepared after a Caerin polypeptide is 125I-labeled has a remarkable effect on inhibiting tumor cells and is capable of remarkably improving the treatment effect of the Caerin polypeptide, reducing the use dosage of the Caerin polypeptide and reducing the preparation cost and chemical toxicity of the drug for treating the tumors and particularly tumors relevant to HPV infection, so that the treatment cost of the tumors is reduced, and the medication safety of the tumors is improved.

Description

technical field [0001] The invention belongs to the field of biotechnology, and specifically relates to 125 I-marked Caerin polypeptide and its application in the preparation of medicines for treating tumors, especially tumors related to HPV infection. Background technique [0002] Caerin polypeptides are a series of biologically active polypeptides extracted and identified from the secretions of the back glands of tree frogs (Litoria) distributed in the coastal rainforest of eastern Australia. cells have anticancer activity. F1 polypeptide and F3 polypeptide are two types of Caerin polypeptides, and their sequences were first published in the paper "New antibiotic caerin 1 peptides from the skinsecretion of the Australian tree frog Litoria chloris. Comparison of the activities of the caerin 1 peptides from the genus Litoria", wherein the F1 sequence is: GLLSVLGSVAKHVLPHVVPVIAEHL-NH2; the F3 sequence is: GLFGVLGSIAKHLLPHVVPVIAEKL-NH2. [0003] The paper "Host-defence pept...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/46C07K1/13A61K38/17A61K51/08A61P35/00A61P31/20A61K101/02
CPCA61K38/00A61K51/08C07K14/463
Inventor 袁建伟王天放潘宣倪国颖刘晓松朱然
Owner WNL生物医学科技公司
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