CN113943803A - Application of HTR6 in diagnosis and prognosis of breast cancer - Google Patents
Application of HTR6 in diagnosis and prognosis of breast cancer Download PDFInfo
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Abstract
The invention discloses an application of HTR6 in diagnosis and prognosis of breast cancer. In a first aspect of the invention, there is provided the use of a reagent for the quantitative detection of HTR6 in the manufacture of a kit for the diagnosis and/or prognosis of breast cancer. According to the application of the embodiment of the application, at least the following beneficial effects are achieved: the invention screens the differentially expressed genes or the proteins thereof in the breast cancer and the normal breast tissue by a series of bioinformatics analysis methods, firstly provides the HTR6 gene or the HTR6 protein as the biomarker for diagnosing the breast cancer or continuously judging the progression or prognosis of the breast cancer, makes up the deficiency of the existing breast cancer diagnosis index, and has good clinical diagnosis value.
Description
Technical Field
The application relates to the technical field of breast cancer diagnosis, in particular to application of HTR6 in diagnosis and prognosis of breast cancer.
Background
Breast cancer is a common malignant tumor and one of the tumors with high lethality. The main cause of death of breast cancer is not the growth of carcinoma in situ, and currently, the diagnosis of breast cancer includes clinical symptom diagnosis, pathological diagnosis and imaging diagnosis, but indexes used in the diagnosis method have obvious indication effect only when the disease develops to a certain stage. The pathological diagnosis is mainly tissue biopsy, the method is tedious and time-consuming, the sampling is not easy, and the sampling process can cause damage to a subject. Imaging is by mammography, but this approach has low early accuracy. In addition to the above methods, diagnosis using biomarkers is currently under rapid development. Currently, diagnostic markers of breast cancer are mainly serum markers, including carcinoembryonic antigen (CEA), sugar chain antigen 153(CA153), and tumor abnormal sugar chain glycoprotein (TAP). However, the accuracy of these serum markers for breast cancer diagnosis remains to be improved. Therefore, there is a need to find more diagnostic biomarkers for breast cancer.
Disclosure of Invention
The present application is directed to solving at least one of the problems in the prior art. Therefore, the application provides a breast cancer marker with good diagnostic value, and the reagent for quantitatively detecting the markers can be used for breast cancer diagnosis or prognosis judgment.
In a first aspect of the application, there is provided the use of a reagent for the quantitative detection of HTR6 in the manufacture of a kit for the diagnosis and/or prognosis of breast cancer.
According to the application of the embodiment of the application, at least the following beneficial effects are achieved:
the application screens genes or proteins thereof differentially expressed in breast cancer and normal breast tissues by a series of bioinformatics analysis methods, and provides the HTR6 gene or the HTR6 protein as a biomarker for breast cancer diagnosis or continuous judgment on the progression or prognosis of the breast cancer for the first time, so that the defect of the existing breast cancer diagnosis index is overcome, and the clinical diagnosis value is good.
Among them, HTR6(5-hydroxytryptamine receptor 6, also known as 5-HT6, 5-HT6R, Gene ID: 3362) is a 5-hydroxytryptamine (serotonin) receptor 6, and the protein encoded by the Gene belongs to the family of seven-transmembrane G protein-coupled receptor proteins, and can bind to the Gs alpha subunit and stimulate adenylate cyclase to activate cyclic AMP-dependent signaling pathway. Diagnosis of breast cancer refers to the determination of whether a subject has breast cancer, while prognosis of breast cancer refers to the prediction of the likely future course and outcome of a subject, including but not limited to the risk of different courses or outcomes of breast cancer, such as recurrence, metastasis, death, etc.
In some embodiments of the present application, the reagents quantitatively detect HTR6 at the gene level or protein level. The reagent for quantitatively detecting nucleic acid at the gene level is carried out by methods including, but not limited to, Polymerase Chain Reaction (PCR), isothermal amplification reaction (such as loop-mediated isothermal amplification (LAMP), Recombinase Polymerase Amplification (RPA) and the like), probe hybridization technique, northern blotting and the like. Reagents for quantitative detection of HTR6 protein at the protein level were performed by methods including, but not limited to, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (IRA), immunohistochemical staining, western blotting, electrophoresis, liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS), and the like.
In some embodiments of the present application, the reagents for quantitative detection of HTR6 at the gene level are selected from the group consisting of primers, probes, and gene chips. The primer is a primer capable of specifically amplifying an HTR6 gene, the probe is a probe capable of specifically recognizing an HTR6 gene or a transcript of the gene, and the gene chip is a composite structure formed by an array in which the probes are immobilized on a substrate material (specifically, but not limited to, a polymer such as a nylon membrane, a nitrocellulose membrane, glass, and the like).
In some embodiments of the present application, the reagent that quantitatively detects HTR6 at the protein level is an antibody. The antibody is an antibody capable of specifically recognizing the HTR6 protein, and specifically includes at least one of a monoclonal antibody and a polyclonal antibody.
In some embodiments of the present application, the breast cancer is Luminal a or Luminal B breast cancer. Currently, the clinical classification of breast cancer based on immunophenotypic profiles of Estrogen (ER), Progestin (PR) and human epidermal growth factor receptor-2 (Her-2) can be roughly divided into four categories: luminal type A, Luminal type B, Her-2 overexpression, and Basal-like. Wherein, Luminal A type is ER positive and/or PR positive and Her-2 negative, Luminal B type is ER positive and/or PR positive and Her-2 positive, Her-2 overexpression type is ER negative, PR negative and Her-2 positive, and Basal-like type is ER negative, PR negative and Her-2 negative. In the experiments of the present application, it was found that in breast cancer, the expression of HTR6 had a significant correlation with recurrence-free survival in patients of lumineal a and lumineal B type, but not in patients of Basal-Like type.
In some embodiments of the present application, prognosis of breast cancer includes predicting the risk of developing invasion, metastasis, or the like of breast cancer. In the embodiment of the application, the expression of HTR6 of invasive breast cancer or breast cancer metastasis is obviously reduced compared with that of in-situ breast cancer, so that the risk of breast cancer metastasis or invasion can be judged by judging whether the expression level of HTR6 of a subject is lower than a set threshold value; if the risk is lower than the threshold value, judging that the risk of metastasis or invasion is lower; and if the wind speed is higher than the threshold value, the wind with the metastasis or invasion is judged to be higher. The set threshold value can be obtained by comprehensive calculation according to expression data of HTR6 in a specific breast cancer patient database, or by comprehensive calculation after detection of a specific clinical experiment sample.
In some embodiments of the present application, the metastasis is lymphatic metastasis and distant metastasis. The lymph metastasis and the number of metastases of breast cancer are important indicators affecting the survival, recurrence and therapeutic effect of breast cancer patients, and in the examples of the present application, it is found that the expression of HTR6 in breast cancer metastases, particularly lymph node metastases, is significantly reduced compared to that in situ tumors, so that the lymph metastasis status of breast cancer can be predicted by the expression level of HTR 6.
In some embodiments of the present application, prognosis comprises predicting survival or survival of the subject. Wherein, the survival period is a judgment on how long the survivor of the testee can be kept according to the illness condition, physical condition and the like of the testee. And survival rate is the probability of survival under a specific time node. In general, survival can be divided into overall survival, median survival, progression-free survival, recurrence-free survival, and the like. In the examples of the present application, applicants found that HTR6 expression in breast cancer patients correlates well with their survival, and thus the survival of subjects can be predicted from the expression level of HTR 6.
In some embodiments of the present application, the survival is at least one of overall survival, relapse-free survival, progression-free survival. Overall survival (OA) refers to the time from the confirmation of breast cancer to death due to any cause. Recurrence-free survival (RFS) refers to the time from initial surgery to the earliest evidence of recurrence. Progression-free survival (PFS) refers to the time from randomization to the first onset of disease progression, or death of any cause, wherein disease progression refers to tumor growth, metastasis of primary tumor lesions, discovery of new lesions, etc. In the examples of the present application, applicants found that the expression level of HTR6 in a subject had a significant correlation with the overall survival, progression-free survival, recurrence-free survival of the subject, and thus the survival of the subject could be predicted by the expression level of HTR 6.
In some embodiments of the present application, the survival rate comprises at least one of an annual survival rate, a two-year survival rate, a three-year survival rate, a five-year survival rate, a ten-year survival rate.
Additional aspects and advantages of the present application will be set forth in part in the description which follows and, in part, will be obvious from the description, or may be learned by practice of the present application.
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FIG. 1 is a graph showing the expression profile of mRNA of HTR6 in example 1 of the present application in various tumor cell lines.
FIG. 2 is the result of the differential expression of HTR6 between breast cancer patients and normal persons in example 1 of the present application.
FIG. 3 shows the results of immunohistochemistry experiment of HTR6 protein of breast cancer patient in example 1 of the present application.
Fig. 4 is a survival curve for breast cancer patients with different levels of HTR6 expression in example 1 of the present application.
Fig. 5 is a survival curve for different types of breast cancer patients with different levels of HTR6 expression in example 1 of the present application.
Detailed Description
The conception and the resulting technical effects of the present application will be clearly and completely described below in conjunction with the embodiments to fully understand the objects, features and effects of the present application. Obviously, the described embodiments are only a part of the embodiments of the present application, and not all embodiments, and other embodiments obtained by those skilled in the art without inventive efforts based on the embodiments of the present application belong to the protection scope of the present application.
The following detailed description of embodiments of the present application is provided for the purpose of illustration only and is not intended to be construed as a limitation of the application.
In the description of the present application, the meaning of a plurality is one or more, the meaning of a plurality is two or more, and the above, below, exceeding, etc. are understood as excluding the present number, and the above, below, within, etc. are understood as including the present number. If the first and second are described for the purpose of distinguishing technical features, they are not to be understood as indicating or implying relative importance or implicitly indicating the number of technical features indicated or implicitly indicating the precedence of the technical features indicated.
In the description of the present application, reference to the description of the terms "one embodiment," "some embodiments," "an illustrative embodiment," "an example," "a specific example," or "some examples," etc., means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the present application. In this specification, the schematic representations of the terms used above do not necessarily refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
Example 1
The CCLE (Cancer Cell Line Encyclopedia, https:// portals. broadassociating. org/CCLE) database contains multiple sets of maps for thousands of Cancer Cell lines, and RNA-seq data for HTR6 were retrieved from the CCLE database, and as a result, as shown in FIG. 1, it can be seen that the gene is expressed in a variety of different tumor Cell lines.
GEPIA (Gene Expression Profiling Interactive Analysis, http:// GEPIA. cancer-pku. cn /) is a visual website based on data integration of GTEx and TCGA, differential Expression data of HTR6 in breast cancer and normal breast tissue are searched in GEPIA and analyzed, and the result is shown in FIG. 2, wherein the abscissa T is 1083 breast cancer patients, N is 291 normal persons, and the ordinate is Transcripts Per Million (TPM). As can be seen in the figure, HTR6 expression is reduced in breast cancer compared to normal or para-cancerous tissue.
The results of detecting the expression of HTR6 in 44 breast cancer patients (including 3 patients with in situ breast cancer, 24 patients with invasive breast cancer, 11 patients with distant metastatic breast cancer, and 6 patients with lymphatic metastatic breast cancer) by immunohistochemical method are shown in fig. 3, where a is the result of staining and b is the result of staining score, and it can be seen from the figure that the expression of HTR6 protein is significantly reduced in the breast cancer tissues in the invasive and metastatic stages compared to in situ breast cancer.
The prognostic value of HTR6 for breast cancer patients was further analyzed using the Kaplan-Meier plotter database, survival curves were calculated using the Kaplan-Meier method, logrank test statistical significance, and a risk ratio (hazard ratio, HR) was calculated with 95% confidence intervals. The results are shown in fig. 4, where the abscissa of the curve is time, the ordinate is survival rate without recurrence, the upper red curve is high expression, the lower black curve is low expression, and the number of risk persons (number at risk) at different time points is below the curve. As can be seen from the figure, the HR 95% confidence interval is 0.78(0.7-0.87), and logrank P is 6.8 × 10-6. This result indicates that the higher the expression level of HTR6, the higher the recurrence-free survival rate of breast cancer patients.
The prognosis value of HTR6 on different types of breast cancer patients is analyzed by continuously using a Kaplan-Meier plotter database, and the result is shown in FIG. 5, wherein A to C are survival curves of Luminal A, Luminal B and Basal-Like types respectively, HR in A is 0.76(0.65-0.9), and logrank P is 0.0011; HR in B is 0.79(0.66-0.94), logrank P is 0.0077; in C, HR is 0.93(0.74-1.16), and logrank P is 0.51. The abscissa of the curve is time, the ordinate is recurrence-free survival rate, the upper red curve is high expression, the lower black curve is low expression, and the number of risk people (number at risk) at different time points is below the curve. This result indicates that, in breast cancer, expression of HTR6 correlates closely with recurrence-free survival in Luminal a and Luminal B patients, while there is no significant correlation with recurrence-free survival in Basal-Like patients. Therefore, the prognosis of Luminal A and Luminal B patients can be judged by the expression of HTR 6.
The HTR6 can be used as an independent diagnosis or prognosis factor of the breast cancer patient and has good diagnosis or prognosis judgment value. It will be appreciated that HTR6 may also be used in combination with other markers to further improve the accuracy or sensitivity of diagnosis or prognosis of breast cancer.
Example 2
Selecting a tissue section of a Luminal B-type breast cancer patient, detecting the relative expression amount of HTR6 protein by an immunohistochemical method, and obtaining an immunohistochemical score of the tissue according to the relative expression amount, wherein when the relative expression amount is lower than a set threshold value, the patient has higher risk of poor prognosis.
The present application has been described in detail with reference to the embodiments, but the present application is not limited to the embodiments described above, and various changes can be made within the knowledge of those skilled in the art without departing from the gist of the present application. Furthermore, the embodiments and features of the embodiments of the present application may be combined with each other without conflict.
Claims (10)
1. Use of a reagent for the quantitative detection of HTR6 in the manufacture of a kit for the diagnosis and/or prognosis of breast cancer.
2. The use according to claim 1, wherein the reagent quantitatively detects HTR6 at the gene level or protein level.
3. The use according to claim 2, wherein said reagents for quantitative detection of HTR6 at the gene level are selected from the group consisting of primers, probes, and gene chips.
4. The use according to claim 2, characterized in that the reagent for the quantitative detection of HTR6 at the protein level is an antibody.
5. The use of claim 1, wherein the breast cancer is a Luminal a or Luminal B breast cancer.
6. The use of claim 1, wherein the prognosis comprises predicting the risk of invasion, metastasis of breast cancer.
7. Use according to claim 6, wherein the metastasis is lymphatic metastasis.
8. The use of claim 1, wherein the prognosis comprises predicting survival or survival of the subject.
9. The use of claim 8, wherein the survival is at least one of overall survival, recurrence-free survival, progression-free survival.
10. The use of claim 8, wherein the survival rate comprises at least one of an annual survival rate, a two-year survival rate, a three-year survival rate, a five-year survival rate, and a ten-year survival rate.
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