A streptomyces strain, and anti-kidney-cancer applications of piericidin metabolites thereof

A technology of streptomyces and compounds, which is applied in the field of application of penicillin compounds, can solve the problems of weak research on the mechanism of action and the need for further exploration of anti-tumor potential

Active Publication Date: 2017-12-15
SOUTH CHINA SEA INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

According to reports, fenopteromycin compounds have insecticidal and antibacterial activities, and also have inhibitory activity on some tumor cells, but the research on their mechanism of action is very weak, and the anti-tumor potential needs to be further explored

Method used

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  • A streptomyces strain, and anti-kidney-cancer applications of piericidin metabolites thereof
  • A streptomyces strain, and anti-kidney-cancer applications of piericidin metabolites thereof
  • A streptomyces strain, and anti-kidney-cancer applications of piericidin metabolites thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1 Streptomyces sp.HBERC-58855 cultivation, identification, fermentation and HPLC analysis

[0034] 1. Solid culture of Streptomyces sp.HBERC-58855

[0035] Streptomyces sp.HBERC-58855 is isolated from the mangrove bottom mud. The strain is stored on the slope of ISP-2 medium. The composition of ISP-2 medium is: yeast extract powder 4g, glucose 4g, malt extract powder 10g, Coarse sea salt 30g, agar powder 20g, water 1000mL, pH 7.2~7.4.

[0036] 2. Identification of 16s rRNA gene of Streptomyces sp.HBERC-58855

[0037] Genomic DNA of Streptomyces sp.HBERC-58855 was extracted by microwave method and part of it was used as a template. Using 16S universal primers, rTaqase from Trans Gene Company was used to perform PCR amplification at 58°C as the annealing temperature and other conditions. The amplified PCR products were subjected to TA cloning and chemical transformation, and positive clones were sent for sequencing. Then the sequence obtained was spliced ​​a...

Embodiment 2

[0043] Example 2 Separation, Purification and Structural Identification of Penicillins Compounds

[0044] Streptomyces sp.HBERC-58855 strain fermented dry extract was dissolved in a small amount of methanol, mixed with silica gel, and subjected to medium pressure silica gel column chromatography (200-300 mesh), eluent was petroleum ether: dichloromethane: methanol system Gradient elution. 8 elution sites S1-S8 were obtained (petroleum ether; petroleum ether: dichloromethane 2:1; dichloromethane; dichloromethane: methanol 200:1; 100:1; 50:1; 30:1; methanol) . S2 was repeatedly separated and purified by preparative HPLC to obtain purified compounds 1 (Piericidin A, 242 mg), 4 (112 mg), 5 (67 mg) and 6 (14 mg). S3 was repeatedly separated and purified by preparative HPLC to obtain purified compounds 1 (82 mg), 2 (24 mg) and 3 (32 mg).

[0045] High resolution mass spectrometry HRMS and figure 1 consistent. The NMR H and C spectrum assignments of the six compounds are shown i...

Embodiment 3

[0053] Example 3 Inhibitory Activity of Penicillins Compounds to Three Lines of Renal Cancer Cells

[0054] Three human renal cell lines were ordered from Shanghai Cell Resource Center, Chinese Academy of Sciences: 786-O human renal cell line (Cat#TCHu186); ACHN human renal cell line (Cat#TCHu199); OS-RC-2 human renal cell line strain (Cat#TCHu40).

[0055] The inhibitory activity of renal cancer cells was tested by CCK-8 assay. Collect the cells in the logarithmic growth phase, count them, resuspend the cells with complete medium, adjust the cell concentration to an appropriate concentration (determined according to the results of the cell density optimization test), inoculate a 96-well plate, and add 100 μl of cell suspension to each well. Cells were incubated for 24 hours at 37°C, 100% relative humidity, 5% CO2 incubator. Dilute the compound to be tested to an appropriate concentration with medium, and add 25 μl / well to the cells. For ACHN cells, the final concentration ...

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Abstract

A streptomyces strain, and anti-kidney-cancer applications of piericidin metabolites thereof are disclosed. The streptomyces strain is streptomyces sp.HBERC-58855, and is deposited in the China Center for Type Culture Collection, in Wuhan University, Wuhan, China on April 20, 2017. The accession number of the streptomyces sp.HBERC-58855 is CCTCC NO:M 2017186. The 16sRNA nucleotide sequence of the streptomyces sp.HBERC-58855 is shown as SEQ ID NO:1. Piericidin compounds including piericidin A, piericidin A2, piericidin C2, IT-143-A, IT-143-B and piericidin s3a are extracted from the streptomyces sp.HBERC-58855 through fermentation for the first time, and are applied in preparation of anti-kidney-cancer medicines. Candidate compounds are provided for development of novel anti-kidney-cancer medicines. The streptomyces and the piericidin compounds are of great significance for marine microorganism medicine resource development.

Description

technical field [0001] The invention belongs to the technical field of application of microorganisms and natural products thereof. More specifically, the invention relates to a Streptomyces sp. HBERC-58855 and the application of the fermentemycin compound obtained by fermenting and extracting the same. Background technique [0002] For a long time, malignant tumors have become one of the major diseases that seriously endanger human life and quality of life. For the treatment of malignant tumors, natural products and their derivatives play an important role. According to reports, from 1981 to 2008, anti-tumor drugs derived from natural products accounted for more than 60% of the anti-tumor drugs on the market, and the number of new natural products and their derivatives as new anti-tumor drugs is still increasing. [0003] Kidney cancer is one of the ten deadliest cancers in the world, and its incidence rate is second only to bladder cancer among urinary system tumors in my ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P17/12C12P17/16C07D213/69C07D405/06A61K31/44A61K31/4427A61P35/00A61P13/12C12R1/465
CPCC07D213/69C07D405/06C12P17/12C12P17/16C12N1/205C12R2001/465
Inventor 周雪峰方伟唐斓李坤龙刘永宏
Owner SOUTH CHINA SEA INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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