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Circulating tumor cell metabolic typing marker and application thereof

A tumor cell and typing technology, applied in the determination/testing of microorganisms, biochemical equipment and methods, etc., can solve the problems of increasing the phenotypic heterogeneity of CTCs, limiting the clinical application of CTCs molecular biology detection, etc., and achieve good value Effect

Active Publication Date: 2018-07-31
NANFANG HOSPITAL OF SOUTHERN MEDICAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, EMT is a dynamic transformation process, and CTCs in the blood can undergo mesenchymal-epithelial transition (MET) when they reach a suitable colonization site, thus increasing the phenotypic heterogeneity of CTCs, that is, identification according to EMT markers The same subtype of CTCs may have different trends and outcomes
This outcome uncertainty poses challenges to the interpretation of CTCs subtype analysis, thus limiting the clinical application of CTCs molecular biology detection

Method used

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  • Circulating tumor cell metabolic typing marker and application thereof
  • Circulating tumor cell metabolic typing marker and application thereof
  • Circulating tumor cell metabolic typing marker and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Screening and Identification of Carbohydrate Metabolism Markers Related to Tumor Metastasis

[0038] 1. Materials and methods:

[0039] (1) Cell lines: PC-3M 2B4, PC-3M 1E8, LNCAP, PC-3, DU145 cells. Among them, PC-3M 2B4 and PC-3M 1E8 are a pair of low-metastatic and high-metastatic prostate cancer cell lines with homologous gene background, purchased from the Cell Center of the Chinese Academy of Medical Sciences; LNCAP, PC-3 and DU145 are three commonly used prostate cancer cell lines The cell lines, whose transfer ability was enhanced in turn, were purchased from the Chinese Academy of Sciences Collection Cell Bank.

[0040] (2) Culture medium and fetal bovine serum: purchased from Guangzhou Xinbang Biotechnology Co., Ltd. (manufactured by Gibco). Among them, DMEM medium is used to cultivate prostate cancer PC-3M 2B4, PC-3M 1E8 and PC-3 cells; RPMI 1640 medium is used to cultivate prostate cancer LNCAP and DU145 cells; 10% fetal bovine serum is added dur...

Embodiment 2

[0058] Example 2 CTCs glucose metabolism typing

[0059] 1. Materials and Methods

[0060] (1) CTCs isolation and identification: CanPatrol CTCs isolation kit was purchased from Guangzhou Yishan Biotechnology Company. 5mL of anticoagulated venous blood was collected from tumor patients, and the red blood cells were lysed. After that, the microporous membrane filtration technology was used to separate and enrich CTCs according to the size difference between white blood cells and CTCs. The CTCs enriched on the filter membrane were fluorescently labeled with DAPI dye nuclear staining and CD45 detection probe, and CTCs were identified by combining the atypia characteristics of the nucleus and the negative expression of CD45.

[0061] (2) CTCs glucose metabolism typing:

[0062] 1) Probes for detecting PGK1 and G6PD were synthesized. The detection probes targeting PGK1 and G6PD were designed and synthesized by Invitrogen, and the sequences were as follows:

[0063] Probes for PG...

Embodiment 3

[0086] Example 3 Correlation between glucose metabolism typing of CTCs and tumor metastasis

[0087] 1. Materials and Methods

[0088] (1) 48 patients with pathological diagnosis of prostate cancer were recruited, including 26 patients with metastasis and 22 patients without metastasis. With the informed consent of the patients, 5 mL of peripheral blood was collected to detect the number of CTCs and their glucose metabolism subtypes by the above method.

[0089] (2) Simultaneously detect the EMT phenotype of CTCs, and use the classic EMT typing markers (E marker EpCAM / CKs and M marker Vimentin / Twist) to divide CTCs into epithelial type (E-CTCs), mixed type ( H-CTCs) and mesenchymal type (M-CTCs), the detection kit was purchased from Guangzhou Yishan Biotechnology Company.

[0090] (3) To compare the characteristics of CTCs glucose metabolism and EMT subtypes between metastatic and non-metastasis patients, and to evaluate the clinical value of CTCs typing in distinguishing pr...

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Abstract

The invention discloses a circulating tumor cell metabolic typing marker and application thereof, and the circulating tumor cell metabolic typing marker includes phosphoglycerate kinase 1PGK1 and 6-phosphate glucose dehydrogenase G6PD. Functional typing of circulating tumor cells (CTCs) is performed from the perspective of metabolic activity, and change trends and biological behaviors of the CTCscan be better reflected. Validation results in prostate cancer specimens indicate that GM+CTCs subtype is more correlated with tumor metastasis compared with EMT subtype, and can be used as an auxiliary diagnostic marker for tumor metastasis. Therefore, CTCs glucose metabolism typing is an important supplement to CTCs counting and EMT phenotype typing, can provide useful information for clinical disease assessment, and has a good application prospect in tumor metastasis diagnosis and monitoring.

Description

technical field [0001] The invention belongs to the field of biological detection, in particular to the application of a metabolic typing marker of circulating tumor cells. Background technique [0002] The molecular biological analysis of circulating tumor cells (CTCs) is mainly the detection and analysis of the expression and mutation of tumor-specific markers, including cell morphology phenotype markers (such as EpCAM / CKs / Vimentin / Twist), tumor stem cell markers ( Such as CD44 / CD133 / ALDH1), tissue type-specific markers (such as ER / PSA / ERCC1) and drug targets (such as EGFR / ALK / VEGF), etc. On the basis of the separation and counting of CTCs, the characteristic analysis of CTCs can further provide information related to the occurrence and development of the source tumor, which is of great significance to the clinical diagnosis and treatment of tumors. Recently, the detection of epithelial-mesenchymal transition (EMT) phenotype of CTCs has received the most attention, that i...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886
CPCC12Q1/6886C12Q2600/158
Inventor 王前蔡贞陈静郑磊裘宇容
Owner NANFANG HOSPITAL OF SOUTHERN MEDICAL UNIV
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