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Application of microRNAs in preparation of reagent or kit for early screening or diagnosing Brachyury positive tumors

A kit, the technology of hsa-mir-449c, is applied in the field of medical biological detection to achieve the effects of prolonging the survival period of patients, preventing metastasis and reducing mortality

Active Publication Date: 2015-04-08
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The current research has not yet seen the microRNAs associated with tumor-specific targets as markers for early diagnosis of tumors (including lung cancer, etc.), and there is no kit for detecting the content of this series of microRNAs in serum to target the overexpression of some Brachyury Screening of patients with tumors (including lung cancer, etc.)

Method used

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  • Application of microRNAs in preparation of reagent or kit for early screening or diagnosing Brachyury positive tumors
  • Application of microRNAs in preparation of reagent or kit for early screening or diagnosing Brachyury positive tumors
  • Application of microRNAs in preparation of reagent or kit for early screening or diagnosing Brachyury positive tumors

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1: microRNAs regulated by tumor-specific transcription factor Brachyury

[0051] In order to find the microRNAs regulated by the tumor-specific transcription factor Brachyury, the inventors used lentivirus infection to transfer the cds sequence of exogenous Brachyury in a lung cancer cell line H1299 and 293T as a normal human control cell to overexpress it Then microRNA chip technology was used to screen out several microRNAs that were significantly up-regulated after being regulated by Brachyury (see figure 1 , figure 2 ).

[0052] The expression changes of these microRNAs were verified by Real-Time PCR to be consistent with the chip results. Subsequently, in the clinical plasma collection and detection of lung cancer patients, chronic obstructive pneumonia patients and normal people, it was found that hsa-miR-4286 and hsa-miR-449c in the microRNAs group showed significant high expression in lung cancer patients, while in the microRNAs group There was no s...

Embodiment 2

[0053] Embodiment 2: the preparation of kit of the present invention

[0054] The kit of the present invention consists of:

[0055] A. 2 tubes of specific reverse transcription primers, 10uM concentration, 200uL;

[0056] B. Real-Time PCR amplification upstream primer (see Table 2) 2 tubes (A1, A2) 10uM concentration, 200uL / tube; Real-Time PCR amplification downstream primer (see Table 2) 1 tube 10uM concentration 2mL / tube;

[0057] C. 1 tube of standard containing quantitative microRNAs;

[0058]This group of microRNAs-specific reverse transcription primers is based on the mature microRNAs sequence to add a fixed Loop (loop structure) fragment, and then synthesized by the American Invitrogen Company, the purity is PAGE grade, and the synthesized primers are DEPC H 2 O was dissolved to a final concentration of 10 μM.

[0059] Specific microRNAs upstream and downstream primers: add empirical sequences based on the specific sequences of mature microRNAs and adjust the GC con...

Embodiment 3

[0061] Embodiment 3: the detection method of this kit

[0062] (1), collecting plasma samples, extracting microRNAs and reverse transcription:

[0063] Collecting peripheral blood for detection has the advantages of non-invasive and convenient collection. Therefore, early screening of lung cancer by collecting and detecting a series of microRNAs molecular markers in peripheral blood is not only easy to popularize, but also has the advantages of high accuracy. Timely detection of early Brachyury-positive lung cancer can achieve the goal of early detection and early treatment, and improve the survival rate of patients.

[0064] Plasma samples for verification were collected from the inpatient department and outpatient department of Shanghai Changzheng Hospital, 30 specimens of primary lung cancer or lung cancer bone metastases (3 cases were selected as Brachyury-positive patients verified by immunohistochemistry), 30 healthy volunteer specimens (randomly Select 3 of them) and 3...

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Abstract

The invention relates to the technical field of medical and biological detection. According to the invention, a plurality of significantly increased microRNAs regulated and controlled by Brachyury are screened by a microRNA chip, and Real-Time PCR validation is carried out on the series of microRNAs. The invention provides a molecular marker of Brachyury positive tumors, and the molecular marker Has-miR-4286, hsa-miR-449c occurs significant high expression in lung cancer patients and is free of obvious abnormal expression level in other lung solid lesion patients or normal people. The invention further provides an application of microRNAs in preparation of a reagent or a kit for early screening or diagnosing Brachyury positive tumors. The microRNAs provided by the invention can be used for screening or diagnosing tumors at the early stage and has important significance on the early treatment of tumors and saving the medical cost.

Description

technical field [0001] The invention relates to the technical field of medical biological detection, in particular to the application of microRNAs in the preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumors. Background technique [0002] With the increasing incidence of tumors, human health and life are facing great challenges, and the prevention and treatment of tumors has become an urgent responsibility of the medical profession. Early detection and timely treatment become the most effective way to reduce tumor mortality. With the continuous deepening of research on tumor molecules and cells and the serious side effects of existing treatments such as radiotherapy and chemotherapy, the concept of "tumor target" has been widely recognized and a lot of research has been carried out. The core of this concept is to find Molecules specifically expressed in tumors but not expressed or limitedly expressed in normal tissues directly hit tum...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6886C12Q2600/158C12Q2600/178
Inventor 陈素矫健姜东杰周振华肖建如
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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