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Reagent or kit for early diagnosis of hepatocellular carcinoma and application of reagent or kit

A technology for early diagnosis and liver cancer, applied in the direction of microbial determination/testing, biochemical equipment and methods, etc., can solve the problem of few tumor markers

Active Publication Date: 2017-05-31
SHENZHEN KIVITA INNOVATIVE DRUG INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are very few specific tumor markers for the diagnosis and treatment of liver cancer at present, and it is urgent to find more liver cancer markers to provide new targets and new ways for the diagnosis and treatment of early liver cancer

Method used

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  • Reagent or kit for early diagnosis of hepatocellular carcinoma and application of reagent or kit
  • Reagent or kit for early diagnosis of hepatocellular carcinoma and application of reagent or kit
  • Reagent or kit for early diagnosis of hepatocellular carcinoma and application of reagent or kit

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Experimental program
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Embodiment approach

[0028] According to one embodiment of the present invention, the sequence of the primer is:

[0029] UBE2J2 forward primer: 5'-ATGACCCCTTATGAAGGTGG-3' (SEQ ID NO: 1);

[0030] UBE2J2 reverse primer: 5'-TCACTCCTGCGCGATGCT-3' (SEQ ID NO: 2).

[0031] According to one embodiment of the present invention, the sequence of the primer is:

[0032] ATP6VOD1 forward primer: 5'-CGGAGCTTTACTTTAACGT-3' (SEQ ID NO: 3);

[0033] ATP6VOD1 reverse primer: 5'-AGGGATGTAGTTGTCGATTTT-3' (SEQ ID NO: 4).

[0034] According to one embodiment of the present invention, the sequence of the primer is:

[0035] STXBP2 forward primer: 5'-TGAAGGCGGTGGTGGGGGA-3' (SEQ ID NO: 5);

[0036] STXBP2 reverse primer: 5'-AATGTCCTCCAGCTTCTTGT-3' (SEQ ID NO: 6).

[0037] According to the present invention, preferably, the liver cancer is metastatic liver cancer.

[0038] According to the third aspect of the present invention, there is provided a kit for early diagnosis of liver cancer, which includes the reagent...

Embodiment 1

[0043] Boyden's modified method was used to isolate the mRNA of the protrusion and cell body of highly metastatic hepatocellular carcinoma cells and perform RNA-seq sequencing.

[0044] Transfer the high-metastatic HCCLM3 and low-metastatic smmcl7721 liver cancer cells to a suspended cell culture dish with a pore size of 1.0 μm incubated with type I collagen, and let them stand at 37°C for 24 to 30 hours. Wash with 1×PBS, invert the Boyden Petri dish, use a pipette gun to suck off the residual medium on the upper side of the Boyden Petri dish, and then use a cell scraper soaked in Trizol to scrape the cell protrusions on the upper side of the Boyden Petri dish. When isolating the RNA in the cell body, lay the Boyden dish flat, use a pipette gun to suck off the medium in the Boyden dish, and then use a cell scraper soaked in Trizol to scrape the cell body inside the Boyden dish.

[0045] The resulting mRNA was extracted and analyzed by total mRNA sequencing (DRS). More than 5,...

Embodiment 2

[0057] Cell invasion assay.

[0058] Highly metastatic HCCLM3 liver cancer cells were pre-transfected with UBE2J2 siRNA for 24 hours, and then transferred to a hanging cell culture dish with a pore size of 8.0 μm incubated with type I collagen and left at 37°C for 24 hours. Wash with 1×PBS, fix with 3% formalin for 15 minutes, and gently wipe off the upper layer of cells that have not penetrated the small hole with a cotton swab. After washing with PBS, stain with crystal violet, take pictures, and count the number of cells that penetrated.

[0059] The result is as figure 2 A- figure 2 As shown in B, it shows that UBE2J2 has the ability to regulate the invasion of HCCLM3 liver cancer cells, where con represents the control group. ATP6VOD1 and STXBP2 also exhibit similar cell invasion abilities.

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Abstract

The invention provides a reagent or a kit for early diagnosis of hepatocellular carcinoma and an application of the reagent or the kit. Specifically, the invention provides an application of at least one of UBE2J2, ATP6V0D1 and STXBP2. The UBE2J2, ATP6V0D1 and STXBP2 show high expression in highly metastatic hepatocellular carcinoma cells. Tests on a plurality of hepatocellular carcinoma and para-carcinoma specimens prove that expression of the UBE2J2, ATP6V0D1 and STXBP2 in most hepatocellular carcinoma specimens is higher than that in para-carcinoma specimens. Based upon cell invasion assay, it proves that the UBE2J2, ATP6V0D1 and STXBP2 can regulate and control invasion capacity of the hepatocellular carcinoma cells. Therefore, the UBE2J2, ATP6V0D1 and STXBP2, as new targets for hepatocellular carcinoma diagnosis and treatment, have a potential prospect of clinical application.

Description

technical field [0001] The present invention relates to the field of disease diagnostic reagents, and more specifically, relates to the use of at least one of UBE2J2, ATP6VOD1 and STXBP2 as a molecular marker for early diagnosis of liver cancer, and a reagent or kit for early diagnosis of liver cancer. Background technique [0002] Liver cancer is one of the most common malignant tumors in humans, and its incidence rate ranks fourth in the world. About 745,000 people die of liver cancer every year. At present, the routine monitoring methods for hepatocellular carcinoma (HCC) in clinical practice are ultrasound scanning and determination of serum alpha-fetoprotein (alpha-fetoprotein AFP) level. The sensitivity, specificity, and positive predictive value of abdominal ultrasound scan for HbsAg carriers in the diagnosis of HbsAg carriers can reach 71%, and the specificity can reach 93%, but the positive predictive value is only 14%. Only 39%-64%, 76%-91% and 9%-32%. About 40% ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6886C12Q2600/158
Inventor 张晶珠陈宇综陈少鹏
Owner SHENZHEN KIVITA INNOVATIVE DRUG INST
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