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Recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application

A human lysozyme, tumor cell technology, applied in the field of medicine, can solve the problems of non-uniformity, other characteristic differences, and anti-tumor research on the direct killing effect of human lysozyme on tumor cells has not been found.

Pending Publication Date: 2013-09-25
SHANGHAI TRANSGENIC RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in addition to having common bacteriolytic properties, different lysozymes have very different or different properties
Although there are studies on the use of lysozyme in the treatment of tumors, these pharmacological studies are mainly limited to egg white lysozyme extracted from egg whites, and the results of the study only found that egg white lysozyme can be used as a strong immune regulator for the prevention and treatment of malignant tumors. Tumors, no direct inhibitory effect on tumor cell growth
[0007] In conclusion, so far, there has been no report that human lysozyme has a direct killing effect on tumor cells and its application in anti-tumor research, let alone the report that human lysozyme can be used to directly prevent and treat tumors

Method used

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  • Recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application
  • Recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application
  • Recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0169] Human lysozyme (abbreviated as ghLZ) produced by transgenic sheep and its purification

[0170] Human lysozyme transgenic goat milk (the preparation process is the same as CN200510110772.1) 400mL, refrigerated and centrifuged at 4°C, 8000-10000rpm, centrifuged for 15-30min to defat. Remove the precipitated impurities and the fat in the upper layer, get the middle layer liquid and filter it with skimmed milk gauze to obtain the skimmed whey (SDS-PAGE analysis such as figure 1 Shown in lane 2) 300mL. An equal volume of 20 mM pH7.4 PB buffer was added thereto, the pH was adjusted to 4.6, and the mixture was bathed in water at 40° C. for 30 min. Refrigerate and centrifuge at 4°C, 8000-10000rpm, and centrifuge for 15-30min to remove casein. Collect the whey (SDS-PAGE analysis such as figure 1 Shown in swimming lane 3), the pH was adjusted to 7.4 with NaOH, and the precipitate was removed by centrifugation again, and collected after gauze filtration.

[0171] SP column (c...

Embodiment 2

[0174] Inhibitory effect of ghLZ on human gastric cancer MGC-803 cells in vitro

[0175] Adjust the density of human gastric cancer MGC-803 cells to 1×10 4 / mL, add 100 μL to 96-well culture plate, that is, 1000 cells / well. Dilute the ghLZ lyophilized powder with RPMI 1640 culture medium to a final concentration of 8 mg / mL, and filter to sterilize with a 0.22 μm filter membrane. Dilute 4-fold with RPMI 1640 culture medium to 4 mg / mL, 1 mg / mL, 0.25 mg / mL, 0.0625 mg / mL and 0.016 mg / mL, 50 μL / well. Each concentration was performed in triplicate.

[0176] At the same time, cisplatin (1mg / mL) was used as a positive control, the RPMI 1640 group without ghLZ was used as a negative control, the culture medium (no cells) and the drug group (no cells) were used as blank controls, and water was added to the surrounding wells of the 96-well culture plate to reduce The liquid in the edge hole evaporates. 37°C, 5% CO 2 Cultivate, pay attention to observation during the period. After 7...

Embodiment 3

[0179] Inhibitory effect of ghLZ on human gastric cancer BGC-823 cells in vitro

[0180] The method is the same as in Example 2, except that the cells are human gastric cancer BGC-823 cells.

[0181] The result is as Figure 4 As shown, the inhibition rate was 50.24% at 1 mg / mL, and it was determined to be active if it was greater than 50%.

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Abstract

The invention relates to a recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application. Specifically, the invention discloses new application of the recombinant human lysozyme (ghLZ). The recombinant human lysozyme is completely consistent with natural human lysozyme, and has no variant. The ghLZ not only retains the bactericidal activity of broad gram positive bacteria, but also has unique broad spectrum in vitro tumor cell killing activity at the same time.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to a recombinant human lysozyme with broad-spectrum tumor cell killing activity and its application. Background technique [0002] Tumor cells have three remarkable basic characteristics: immortality, migration and loss of contact inhibition. In addition, tumor cells have many physiological, biochemical and morphological characteristics different from normal cells. The tumor cell cycle is not controlled by the normal growth regulation system, and can continue to divide and proliferate. The benign and malignant tumors and the malignant degree of the tumor are usually determined according to the degree of differentiation and maturity of the tumor's solid morphology and atypia. Malignant tumors grow rapidly and can be transferred to other parts of the body. They also produce harmful substances that destroy normal organ structures, make the body dysfunctional, and even threaten lif...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/47A61K45/00C12N5/09C12Q1/34A61K35/00
Inventor 成国祥刘思国陈建泉
Owner SHANGHAI TRANSGENIC RES CENT