Molecular marker for diagnosing and treating multiple myeloma

A multiple myeloma, material technology, applied in the field of tumor diagnosis, prognosis prediction, treatment, can solve the problem of no patient

Active Publication Date: 2016-12-14
GUAN BOJIAN BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the main treatment for multiple myeloma (MM) is still combined chemotherapy and autologous hematopoietic stem cell transplantation. Because the average age of onset of MM is r...

Method used

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  • Molecular marker for diagnosing and treating multiple myeloma
  • Molecular marker for diagnosing and treating multiple myeloma
  • Molecular marker for diagnosing and treating multiple myeloma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Example 1 Gene Chip Screening for Differentially Expressed Genes

[0075] 1. Materials:

[0076] Multiple myeloma tissue: A total of 10 bone marrow biopsy specimens were collected from confirmed MM patients. The diagnostic criteria for MM refer to the "Chinese Guidelines for the Diagnosis and Treatment of Multiple Myeloma 2011 Revised Edition". Among the patients, there were 5 males and 5 females, with a median age of 59 years.

[0077] Normal bone marrow tissue: 10 bone marrow biopsy specimens were collected from patients with malnutrition anemia at the same period as the control group, including 5 males and 5 females, with a median age of 53 years.

[0078] 2. Obtaining tissue RNA

[0079] Total tissue RNA was extracted using the Trizol one-step method.

[0080] 3. Determination of RNA purity and concentration

[0081] Take 1 μl of RNA solution, measure OD260 and OD280 with the instrument, the RNA concentration is OD260 value × dilution factor × 40 / 1000, calculate...

Embodiment 2

[0094] Example 2 Large sample verification screened out differentially expressed genes

[0095] Considering the gene that has not been studied in the prior art on the correlation between this gene and multiple myeloma as a candidate gene, and considering the results of gene sequencing, the CCDC72 gene (its expression is up-regulated in multiple myeloma tissues) was selected for verification .

[0096] 1. Sample collection

[0097] According to the method of Example 1, 50 cases of multiple myeloma tissues and 60 cases of normal bone marrow tissues were collected.

[0098] 2. Validation at the mRNA level

[0099] 2.1 Extract tissue RNA

[0100] Step is with embodiment 1.

[0101] 2.2 Reverse transcription

[0102] A total of 20 μL of reverse transcription system, including 2 μg / 2 μL of total cell RNA, 1 μL of 50 U / μL Rnasin, 4 μL of 5× reverse transcription reaction buffer, 2 μl of 10 mM d NTP, 2 μL of 50 μg / mL random primer (promega), 200 U / μL M-MLV Reverse transcriptase ...

Embodiment 3

[0125] Embodiment 3 inhibits CCDC72 gene expression

[0126] 1. siRNA design and synthesis

[0127] siRNA sequences against CCDC72:

[0128] siRNA-CCDC72:

[0129] The sense strand is 5'-UUUGUUUCUGCUUGAAAGCCU-3' (SEQ ID NO.5)

[0130] The antisense strand is 5'-GCUUUCAAGCAGAAACAAAAA-3' (SEQ ID NO.6);

[0131] The above siRNA sequences and negative control siRNA sequences (siRNA-NC) were provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd.

[0132] 2. Culture and transfection of multiple myeloma cells

[0133] 2.1 Cell culture

[0134] RPMI8226 cells were placed in RPMI 1640 medium containing 15% fetal bovine serum (FBS), penicillin 100 U / ml, and streptomycin 100 μg / ml at 37°C, 5% CO 2 , cultivated in a saturated humidity environment.

[0135] 2.2 Cell transfection

[0136] (1) 24 hours before transfection, inoculate 0.5-2*10 5 cells, the cell confluency was 30-50% at the time of transfection. When plating, the cells should be digested and mixed completely t...

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Abstract

The invention discloses a molecular marker with a CCDC72 gene for diagnosing the multiple myeloma. An experiment shows that compared with a normal myeloid tissue, the CCDC72 gene has a large expression quantity in a multiple myeloma tissue. The invention further discloses application of the CCDC72 gene in preparation of a medicine for treating the multiple myeloma. The research achievement provides a novel clinical multiple myeloma diagnosis method, and also provides a novel medicine target for treating the multiple myeloma.

Description

technical field [0001] The present invention relates to the fields of tumor diagnosis, treatment, and prognosis prediction. More specifically, the present invention relates to a method for tumor diagnosis and prognosis prediction by detecting abnormality of CCDC72; and a tumor therapeutic agent for inhibiting CCDC72 gene or protein. Background technique [0002] Multiple myeloma (MM) is a malignant tumor with abnormal proliferation of plasma cells in the terminal stage of B cell differentiation. Abnormal differentiation of plasma cells leads to bone damage and changes in metabolism and immune function. With the aging of my country's population, the incidence of MM is on the rise. In my country, the annual incidence rate of MM is 1 / 100,000, while that in western industrialized countries is 4 / 100,000. In the United States, the annual incidence of MM is 14,000. Although people have known about the malignant hyperplasia of MM for 150 years, MM is still an incurable tumor so fa...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N33/68G01N33/574A61K45/00A61P35/00
Inventor 杨承刚林慧芳
Owner GUAN BOJIAN BIOTECH CO LTD
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