A kit for the treatment of colorectal cancer

Abstract

The application discloses a colorectal cancer treatment kit. The application discloses a colorectal cancer treatment kit. The kit comprises a WWP1 gene inhibiting reagent. Preferably, the kit comprises a reagent for inhibiting the expression level of a WWP1 gene protein. More preferably, the kit comprises a reagent for inhibiting the expression level of a WWP1 gene protein, and the reagent is composed of the following nucleotide: Si-WWP1: 5'-GACCAAAGCUUUCCUUGAUTT-3'. The application provides a colorectal cancer treatment kit and application of a reagent for inhibiting the expression level of a WWP1 gene protein by determining the correlation between the expression level of a WWP1 gene protein and colorectal cancer, and provides a new method for treating colorectal cancer.

Description

Technical Field

[0001] This invention belongs to the field of medical diagnostics, and more specifically, relates to a colorectal cancer treatment kit. Background Technology

[0002] Colorectal cancer (CRC) is a collective term for colon cancer and rectal cancer. It is a type of malignant tumor of the digestive tract with high incidence and mortality. Thoroughly investigating the underlying pathogenic factors of colorectal cancer and improving its prevention, diagnosis, treatment, and prognosis remains a significant challenge for researchers.

[0003] The ubiquitin-proteasome system (UPS) is the main degradation pathway for clearing misfolded proteins within cells. In the process of degrading misfolded proteins, UPS first requires the ubiquitination of the substrate. Ubiquitin ligase E3, by specifically recognizing the substrate, mediates the transfer of activated ubiquitin from ubiquitin-conjugating enzyme E2 to the substrate's activated lysine residue, and is one of the key enzymes in the ubiquitination process. E3 mainly contains the HECT domain family, the RING-finger domain family, and the U-box domain family, among which the HECT family E3 itself has the function of elongating polyubiquitinated chains. WWP1, an E3 ligase containing the WW domain, is an E3 ligase in the HECT domain family. It determines the linkage form of polyubiquitinated chains and performs different forms of ubiquitination modifications on different substrates, including monoubiquitination, S-48 cleavage, and S-63 cleavage polyubiquitination. Its expression is significantly increased in various diseases, such as lung cancer, gastric cancer, Huntington's disease, and colorectal cancer. Lowering the expression level of the WWP1 gene can significantly inhibit the proliferation and migration of cancer cells, increase cancer cell apoptosis, and inhibit the growth of cancer tissue. Based on current research on this disease and the WWP1 gene, there are currently no clear drugs targeting this gene. Summary of the Invention

[0004] In view of the above-mentioned defects or improvement needs of the existing technology, the present invention provides a colorectal cancer treatment kit. Its purpose is to solve the technical problem of the lack of effective clinical treatment methods for colorectal cancer by using a reagent that inhibits WWP1 protein expression to prepare the colorectal cancer treatment kit, since the exact relationship between the WWP1 gene and colorectal cancer has been clarified.

[0005] To achieve the above objectives, according to one aspect of the present invention, a colorectal cancer treatment kit is provided, comprising a reagent for inhibiting the WWP1 gene.

[0006] Preferably, the colorectal cancer treatment kit includes a reagent that inhibits the expression level of the WWP1 gene protein.

[0007] More preferably, in the colorectal cancer treatment kit, the reagent for inhibiting the expression level of the WWP1 gene protein is composed of the following nucleotides:

[0008] Si-WWP1:5'-GACCAAAGCUUUCCUUGAUTT-3'

[0009] In summary, compared with the prior art, the above-described technical solutions conceived by this invention can achieve the following beneficial effects:

[0010] This invention clarifies the association between WWP1 gene protein level expression and colorectal cancer, and provides a colorectal cancer treatment kit and the application of reagents that inhibit WWP1 gene protein level expression, offering a novel approach to the treatment of colorectal cancer. Attached Figure Description

[0011] Figure 1 Effects of an inhibitory agent for WWP1 gene protein expression (siWWP1) on the proliferation capacity of cancer cells.

[0012] Figure 2 Effects of an inhibitory agent for WWP1 gene protein expression (siWWP1) on the migration ability of cancer cells.

[0013] Figure 3 Effects of a reagent that inhibits WWP1 gene protein expression (siWWP1) on tumor growth in mice. Detailed Implementation

[0014] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0015] Furthermore, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0016] Example 1

[0017] A colorectal cancer treatment kit includes: a Lipofectamine 2000 liposome transfection kit and an siRNA reagent to inhibit the expression of the WWP1 gene protein: Si-WWP1:5'-GACCAAAGCUUUCCUUGAUTT-3'

[0018] Example 2

[0019] Cell transfection:

[0020] Lipofectamine 2000 was used as the transfection reagent. Taking 6-well plate transfection as an example: Cells were first evenly seeded into 6-well plates and transfected when they reached 60%-90% confluency. First, an appropriate amount of Lipofectamine 2000 (5-7.5 μL / well) was diluted with Opti-MEM medium and allowed to stand for 5 minutes. During these 5 minutes, the nucleic acid to be transfected was diluted with Opti-MEM. The amount of siRNA used was 100-200 pmol per well, and the amount of plasmid used was 0.5-1.0 μg / well. After 5 minutes, the diluted Lipofectamine 2000 and nucleic acid were mixed and allowed to stand for 20 minutes. Near the end of the 20 minutes, the 6-well plate was removed from the incubator, the medium was aspirated, and replaced with 2 mL of serum-free Opti-MEM medium. When the 20 minutes were finished, the Lipofectamine 2000 coated with nucleic acid was added to the 6-well plate and gently shaken to mix. Then the plate was incubated. After 4-6 hours in the incubator, the serum-free Opti-MEM should be replaced with pre-prepared culture medium containing 2% FBS.

[0021] Animal experiments:

[0022] A. SW480 cells were evenly seeded in 100mm culture dishes; B. When the cell density reached 60-80%, siWWP was transfected; C. 48 hours after transfection, the cells were digested with trypsin, washed twice with PBS, and resuspended in PBS to adjust the concentration to 1.5 x 10⁷ cells / mL; D. PBS-resuspended cells (200 μL / mouse) were implanted into the axilla of BALB / c–nu nude mice. The mice were then fed normally; tumor volume was measured periodically; E. On day 24, when the largest tumor reached approximately 1 cm in diameter, the mice were euthanized by cervical dislocation, and the tumors were removed, photographed, and weighed. Results are shown below. Figure 3 .

Claims

1. A colorectal cancer treatment kit, characterized in that, The invention includes a reagent for inhibiting the expression of the WWP1 gene, wherein the reagent for inhibiting the expression of the WWP1 gene is composed of the following nucleotides: Si-WWP1: 5'- GACCAAAGCUUUCCUUGAUTT-3'.

2. The use of an agent that inhibits WWP1 gene expression in the preparation of a reagent for treating colorectal cancer, characterized in that, The reagent for inhibiting WWP1 gene expression consists of the following nucleotides: Si-WWP1: 5'-GACCAAAGCUUUCCUUGAUTT-3'.

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