CN110687283B - Application of autoantibodies in diagnosis and/or treatment of tumors - Google Patents

Abstract

The present invention provides the application of a reagent for detecting autoantibodies in the preparation of products for diagnosis, treatment and/or prognosis evaluation of tumors and autoimmune diseases, and the diagnosis, treatment and/or prognosis evaluation of tumors or autoimmune diseases Markers, said markers are autoantibodies. The expression level of markers is detected by rapid double-antibody sandwich enzyme-linked immunosorbent technique (rapid-ELISA) to judge or assist in judging the patient's response to tumor immunotherapy and long-term benefits.

Description

Application of autoantibodies in diagnosis and/or treatment of tumors

technical field

The present invention relates to the field of biomedical detection, in particular to the application of autoantibodies in the diagnosis, treatment and prognosis assessment of tumors or autoimmune diseases, wherein the autoantibodies are selected from CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, One or a combination of two or more of LPCAT4 or VMAC.

Background technique

The World Health Organization's Global Burden of Disease Study shows that cancer is the second highest mortality disease in the world and in China after cardiovascular disease. Immunotherapy has become an emerging approach to cancer treatment by boosting a patient's immune system to fight the disease. In many immunotherapy strategies, immune checkpoint inhibitors (Immune Checkpoint Blockade, ICB) by blocking the intrinsic down-regulation programs of immunity, such as cytotoxic T lymphocyte antigen 4 (CTLA-4) and programmed cell death 1 (PD- 1) or its ligand (PD-L1), which increases anti-tumor immunity, has shown significant benefits in the treatment of various cancers. The advent of ICB has increased the overall survival rate of patients with various cancers. At present, a variety of inhibitor drugs have been approved by the US Food and Drug Administration and the China Food and Drug Administration for the treatment of malignant tumors, but the clinical results show that they are in absolute danger. The objective effective rate of patients receiving ICB monotherapy in most and unselected solid tumors is only 10% to 30%. Most patients cannot benefit from it, that is, primary drug resistance. Disease progression, that is, secondary drug resistance, will also occur later. With the advent of more ICB drugs and the gradual increase in the number of drug users, it is very important to find effective predictive markers and establish relevant predictive models, which can not only improve the tumor response rate, but also save treatment costs and time for patients Cost, provide personalized diagnosis and treatment strategies for the treatment of cancer patients, improve the survival period and quality of life of patients.

At present, it is known that early tumor cells can express and produce some abnormal proteins, which can be recognized by the body's immune system and produce specific antibodies. These proteins are called tumor-associated antigens, and their antibodies are called autoantibodies, which are studied in autoimmune diseases In recent years, there have been some researches on tumor screening, early diagnosis, prognosis judgment and curative effect monitoring, and have shown great potential. Compared with other markers, serum autoantibodies are obtained in a simple way, and can be continuously sampled and monitored during treatment. Therefore, using serum autoantibodies to predict the curative effect of ICB has certain advantages. Early studies used methods such as SEREX (serological screening of recombinant cDNA expression library), phage peptide library panning, and SERPA (serum proteomics) to screen tumor autoantibodies. At present, there are a number of published tumor autoantibodies Antibodies are used in the diagnosis of tumors. Among them, NY-SEO-1, p53, Annexin I, 14-3-3θ, LAMR1, PGP9.5, c-myc, HER2, CAGE, GBU-4-5, SOX2 and other autoantibodies are used for lung cancer diagnosis; p53, HSP70 , HCC-22-5, peroxiredoxin VI, KM-HN-1, p90 and other autoantibodies are used for the diagnosis of gastric cancer; p62, HCC1 and other autoantibodies are used for the diagnosis of liver cancer; interleukin-29 (IL29), survivin (SUR), growth hormone (GRH), osteoprotegerin (OPG), and resistin (RES) and other autoantibodies are used for the diagnosis of breast cancer. Regarding the relationship between autoantibodies and the therapeutic effect of ICB, studies have reported the relationship between the production of autoantibodies and the toxic and side effects of ICB treatment, and there is a certain relationship between the level of autoantibodies and the therapeutic effect of ICB.

Eukaryotic translation initiation factor 4E (Eukaryotic translation initiation factor 4E, EIF4E) is a cap-binding protein that can specifically recognize the cap structure at the 5' end of mRNA and plays an important role in the initiation of eukaryotic translation. EIF4E is one of the factors closely related to malignant tumors. It is highly expressed in a variety of human malignant tumors and is closely related to the occurrence, invasion and metastasis of tumors. For example: Literature: EIF4E Expression and Significance in Liver Cancer Tissue, Shandong Medicine, discloses that the positive expression rate of EIF4E in liver cancer tissue is significantly higher than that in paracancerous liver tissue and normal liver tissue, and its expression is related to the patient's own condition, tumor size and The degree of completion of the capsule has nothing to do with the degree of differentiation, clinical stage, and presence or absence of portal vein tumor thrombus. Because the expression of EIF4E protein increases with the degree of differentiation of liver cancer, EIF4E can be used for the diagnosis and treatment of liver cancer. Patent CN106460064A discloses a method for determining the prognosis of patients suffering from high-grade serous ovarian cancer (HG-SOC), the method comprising determining mutations in genes such as ANKHD1-eIF4EBP3. Literature: Research Progress of EIF4E in Cervical Cancer, Modern Oncology, It is disclosed that the expression of EIF4E in cervical tumors is increased and can promote the expression of HPVE7 protein. With the increase of cervical tissue malignancy, the expression of EIF4E is also increased. , so EIF4E is a potential marker for the diagnosis and prognosis of cervical cancer. However, there are still many tumor diseases with low expression of EIF4E. At the same time, EIF4E or EIF4E autoantibodies have not been disclosed as markers for ICB treatment of tumors or autoimmune diseases.

Literature: Expression of CCDC8 Gene in Breast Cancer, Li Yunfen, Tumor Research and Clinic, using quantitative reverse transcription polymerase chain reaction to analyze the expression of CCDC8 in 40 cases of breast cancer combination and 22 cases of benign breast tumor tissues, the results showed , the expression of CCDC8 gene in benign breast tumors was significantly higher than that in breast cancer tissues, and the expression was statistically significant. Since CCDC8 is expressed in breast tumor tissue, and the expression level of malignant tissue is lower than that of benign tissue, the expression of CCDC8 is correlated with age, tumor size, and nm23. Therefore, CCDC8 may be a tumor suppressor gene of breast cancer that affects the development of breast cancer. happen, develop. Literature: Functional research on the new molecule CCDC134, Huang Jing et al. have confirmed through experiments that CCDC134, as a potential cytokine, plays an important role in tumor immunity and autoimmune diseases, and has potential application value. Document: The expression and clinical significance of CCDC67 gene in papillary thyroid carcinoma, Lei Mengyuan, master's degree thesis of Zhengzhou University, the expression of CCDC67 mRNA and protein in differentiated thyroid cancer cell lines by real-time fluorescent quantitative PCR technology and cellular immunofluorescence technology The situation was analyzed to determine the role of CCDC67 gene in thyroid tumors. Literature: Effect of Silencing DDX49 Gene on Osteosarcoma Cell Proliferation, Lanzhou University, Postgraduate Dissertation, Publicly Detecting DDX49 Gene Expression to Determine Its Role in Osteosarcoma Cell Proliferation and Apoptosis, Provide New Diagnosis and Treatment for Osteosarcoma molecular targets. However, the above-mentioned existing technologies all use a single autoantibody as a target as a potential marker for diagnosis or prognosis, and the sensitivity and specificity are limited; and the combination of EIF4E2, CCDC130, UBALD1, LPCAT4 and/or VMAC autoantibodies has not been disclosed. The application of the combination of FATE1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies and the combination of CPLX2, DDX49, PHACTR1, EIF4E2 and VMAC autoantibodies in the preparation of products for therapeutic effect prediction or prognosis evaluation of tumor or autoimmune diseases.

Contents of the invention

The inventors successfully detected the levels of CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4, and VMAC autoantibodies in the blood of patients with tumors or autoimmune diseases by rapid double-antibody sandwich enzyme-linked immunosorbent assay (rapid-ELISA) , and the levels of these types of autoantibodies in the same serum sample are different. Among patients with high autoantibodies, there are more patients with good curative effect on immunotherapy, indicating that autoantibodies can be used as potential tumor diagnosis, immunotherapy efficacy prediction and prognosis evaluation of markers.

An object of the present invention is to provide the use of reagents for detecting autoantibodies in the preparation of products for diagnosis, treatment and/or prognosis evaluation of tumors and autoimmune diseases, and the use of autoantibodies in the preparation of tumors for diagnosis, treatment and/or prognosis evaluation , Application in products for autoimmune diseases.

Another object of the present invention is to provide a marker for the diagnosis, treatment and/or prognosis assessment of tumors and autoimmune diseases.

Another object of the present invention is to provide a product for diagnosis, treatment and/or prognosis assessment of tumors and autoimmune diseases.

Another object of the present invention is to provide a method for detecting autoantibodies.

Another object of the present invention is to provide a method for diagnosis, treatment and/or prognosis assessment of tumors and autoimmune diseases.

The first aspect of the present invention provides the use of reagents for detecting autoantibodies in the preparation of products for diagnosing and/or treating tumors and autoimmune diseases, wherein the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

Preferably, the detection of autoantibodies is to detect the presence or expression level of autoantibodies.

Preferably, the method for detecting autoantibodies by the reagent for detecting autoantibodies is selected from one or more of ELISA, rapid-ELISA, western blot, indirect immunofluorescence, enzyme immunospot method or immunoluminescent method. combination.

In a specific embodiment of the present invention, the reagent for detecting autoantibodies and the method for detecting autoantibodies are rapid-ELISA.

The second aspect of the present invention provides the use of reagents for detecting autoantibodies in the preparation of products for evaluating the prognosis of tumors or autoimmune diseases, wherein the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the prognosis assessment of tumor treatment is the prognosis assessment after treatment with immune checkpoint inhibitors, or the combination of immune checkpoint inhibitors and other drugs, radiotherapy, chemotherapy and other treatment methods.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

Preferably, the detection of autoantibodies is to detect the presence or expression level of autoantibodies.

Preferably, the method for detecting autoantibodies by the reagent for detecting autoantibodies is selected from one or more of ELISA, rapid-ELISA, western blot, indirect immunofluorescence, enzyme immunospot method or immunoluminescent method. combination.

In a specific embodiment of the present invention, the reagent for detecting autoantibodies and the method for detecting autoantibodies are rapid-ELISA.

The third aspect of the present invention provides the use of autoantibodies in the preparation of products for diagnosing and/or treating tumors and autoimmune diseases, wherein the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

The fourth aspect of the present invention provides the use of autoantibodies in the preparation of products for evaluating the prognosis of tumors or autoimmune diseases, wherein the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the prognosis assessment of the tumor is treated with immune checkpoint inhibitors, or the prognosis assessment after the combination of immune checkpoint inhibitors and other drugs, radiotherapy, chemotherapy and other treatment methods.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

The fifth aspect of the present invention provides a marker for diagnosis and/or treatment of tumors and autoimmune diseases, said marker comprising autoantibodies, said autoantibodies being selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

In a specific embodiment of the present invention, the markers may also include other immune checkpoint proteins other than CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies or themselves Antibodies are used as accompanying markers to detect the therapeutic effect of tumor or autoimmune diseases in combination with CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies. Wherein, the accompanying marker is selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160.

The sixth aspect of the present invention provides a marker for evaluating the prognosis of a tumor or an autoimmune disease, said marker comprising an autoantibody, and said autoantibody is selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the prognosis assessment of the tumor is treated with immune checkpoint inhibitors, or the prognosis assessment after the combination of immune checkpoint inhibitors and other drugs, radiotherapy, chemotherapy and other treatment methods.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

In a specific embodiment of the present invention, the markers may also include other immune checkpoint proteins other than CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies or themselves As a companion marker, the antibody can be used in combination with CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies to evaluate the prognosis of tumor or autoimmune diseases. Wherein, the accompanying marker is selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160.

The seventh aspect of the present invention provides a product for diagnosing and/or treating tumors and autoimmune diseases, said product comprising reagents for detecting autoantibodies, said autoantibodies being selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

Preferably, the detection of autoantibodies is to detect the presence or expression level of autoantibodies.

Preferably, the method for detecting autoantibodies by the reagent for detecting autoantibodies is selected from one or more of ELISA, rapid-ELISA, western blot, indirect immunofluorescence, enzyme immunospot method or immunoluminescent method. combination.

In a specific embodiment of the present invention, the reagent for detecting autoantibodies and the method for detecting autoantibodies are rapid-ELISA.

Preferably, the products for diagnosing and/or treating tumors and autoimmune diseases may also contain other immune checkpoint proteins except CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies Or its autoantibodies as accompanying markers, combined with CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies to detect the therapeutic effect of tumors and autoimmune diseases. Wherein, the accompanying marker is selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160.

The eighth aspect of the present invention provides a product for evaluating the prognosis of a tumor or an autoimmune disease. The product includes a reagent for detecting autoantibodies, and the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

Preferably, the detection of autoantibodies is to detect the presence or expression level of autoantibodies.

Preferably, the method for detecting autoantibodies by the reagent for detecting autoantibodies is selected from one or more of ELISA, rapid-ELISA, western blot, indirect immunofluorescence, enzyme immunospot method or immunoluminescent method. combination.

In a specific embodiment of the present invention, the reagent for detecting autoantibodies and the method for detecting autoantibodies are rapid-ELISA.

Preferably, the product for evaluating the prognosis of tumor or autoimmune disease may also contain other immune checkpoint proteins or Its autoantibodies are used as accompanying markers, combined with CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies to detect the prognosis of tumors and autoimmune diseases. Wherein, the accompanying marker is selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160.

The ninth aspect of the present invention provides a method for diagnosing tumors or autoimmune diseases. The method includes detecting the presence or expression level of autoantibodies in the body, and the autoantibodies are selected from one of the following groups: :

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

The tenth aspect of the present invention provides a method for evaluating the prognosis of a tumor or autoimmune disease, the method comprising detecting the presence or expression level of autoantibodies in the organism, and the autoantibodies are selected from the following group A sort of:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the autoantibodies are autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the autoantibodies are autoantibodies in serum and/or plasma.

The eleventh aspect of the present invention provides a method for treating tumors or autoimmune diseases, the method comprising administering an effective dose of an immune checkpoint inhibitor to a patient with a tumor, or an immune checkpoint inhibitor and Combination of other drugs and treatment methods such as radiotherapy and chemotherapy, wherein the expression of autoantibodies is detected in the patient, and the autoantibodies are selected from one of the following groups:

(1) EIF4E2;

(2) EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC;

(3) EIF4E2, CCDC130, FATE1, LPCAT4, VMAC;

(4) CPLX2, DDX49, PHACTR1, EIF4E2, VMAC;

(5) EIF4E2, CCDC130, LPCAT4, VMAC;

(6) UBALD1;

(7) FATE1;

(8) EIF4E2, UBALD1;

(9) EIF4E2, FATE1;

(10) CPLX2, DDX49, PHACTR1;

(11) EIF4E2, VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma.

In a specific embodiment of the present invention, the tumor is lymphoma.

Preferably, the immune checkpoint inhibitor is selected from PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160 inhibitors.

Preferably, detecting the expression of autoantibodies in the patient is detecting the expression of autoantibodies in serum, plasma, interstitial fluid, cerebrospinal fluid or urine of the patient. Wherein, the higher the expression level of the autoantibody, the better the effect of the immune checkpoint inhibitor, or the combination of immune checkpoint inhibitor and other drugs, chemotherapy, radiotherapy and other treatment methods.

In a specific embodiment of the present invention, detecting the expression of autoantibodies in the patient is detecting the expression of autoantibodies in serum and/or plasma of the patient.

The twelfth aspect of the present invention provides a method for detecting autoantibodies, comprising coating the corresponding protein on the surface of a carrier, adding a sample to be detected, adding enzymes and substrates, and measuring the concentration.

Preferably, the autoantibody is selected from CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC.

Preferably, the autoantibodies include one or a combination of two or more of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD.

Preferably, the sample to be detected is biological serum, plasma, interstitial fluid, cerebrospinal fluid or urine.

In a specific embodiment of the present invention, the sample to be detected is biological serum.

Preferably, the sample to be detected is diluted with a dilution buffer before being added, and the dilution concentration is 1:100-1000.

More preferably, the dilution concentration is 1:150-500.

Preferably, the dilution concentration is 1:200-400.

In a specific embodiment of the present invention, the dilution buffer is milk; the milk is milk diluted with PBST.

Preferably, the enzyme is an enzyme-labeled antibody. More preferably, the enzyme-labeled antibody is IgG.

In a specific embodiment of the present invention, the substrate is TMB.

Preferably, the method for measuring the concentration is to measure the absorbance value at 450 nm.

In a specific embodiment of the present invention, described method comprises:

1) Coat the capture antibody in a 96-well plate, overnight at 4°C, wash; dilute with milk and block for 1-3h; preferably, dilute with milk and block for 2h;

At the same time, add the corresponding plasmid to the in vitro expression system, react in the dark for 1-3 hours, preferably 2 hours, add it to a 96-well plate after dilution with milk, wash at room temperature for 0.5-2 hours, preferably after 1 hour, preferably, the in vitro expression system is IVTT; preferably Yes, the serum sample is diluted with milk to 1:200-400;

2) adding the diluted serum sample to a 96-well plate, incubating, and washing; the preferred incubation time is 0.5-2 hours;

3) Add freshly diluted anti-human IgG HRP enzyme-labeled antibody, incubate and wash; the preferred incubation time is 0.5-2 hours; the more preferred incubation time is 0.5-1 hour;

4) Add temporarily prepared TMB substrate to develop color in the dark; add sulfuric acid to terminate the reaction; preferably the color development time in the dark is 10-30 minutes;

5) Measure the absorbance value at 450nm to determine the autoantibody expression level in the sample.

The reagents for detecting the expression level of autoantibodies in the present invention are selected from test strips, protein chips, magnetic beads, fluorescent reagents and the like. The detection principle adopts antigen-antibody combination, wherein the detection antigen is protein, polypeptide or antibody.

The product according to the invention comprises the reagent for detecting the level of autoantibodies according to the invention. Preferably, the product is selected from kits and mass spectrometers.

A kit for detecting autoantibodies, comprising reagents for detecting autoantibody levels.

A chip for detecting autoantibodies, comprising reagents for detecting the level of autoantibodies.

A kit for diagnosing and/or treating tumors, comprising reagents for detecting CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibody levels and reagents for detecting other immune checkpoints. The other immune checkpoints are selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160. Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma. In a specific embodiment of the present invention, the tumor is lymphoma.

A kit for evaluating the prognosis of tumor treatment, comprising reagents for detecting CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibody levels and reagents for detecting other immune checkpoints. The other immune checkpoints are selected from one or a combination of two or more of PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1, 2B4 or CD160. Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma. In a specific embodiment of the present invention, the tumor is lymphoma.

Diagnosing tumors in the present invention refers to diagnosing whether there is a tumor, or assessing the prognosis of a tumor patient, or assessing the degree of benefit of a tumor patient treated with an immune checkpoint inhibitor.

The treatment of tumors in the present invention refers to detecting the expression levels of CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4 and/or VMAC autoantibodies to determine whether to be treated with immune checkpoint inhibitors.

The tumor of the present invention is selected from lymphoma, non-small cell lung cancer, leukemia, ovarian cancer, breast cancer, endometrial cancer, colon cancer, rectal cancer, gastric cancer, bladder cancer, lung cancer, bronchial cancer, bone cancer, prostate cancer , pancreatic cancer, liver and bile duct cancer, esophageal cancer, kidney cancer, thyroid cancer, head and neck cancer, testicular cancer, glioblastoma, astrocytoma, melanoma, myelodysplastic syndrome, and sarcoma. Wherein, the leukemia is selected from acute lymphoblastic (lymphoblastic) leukemia, acute myelogenous leukemia, myelogenous leukemia, chronic lymphocytic leukemia, multiple myeloma, plasma cell leukemia, and chronic myelogenous leukemia; The lymphoma is selected from Hodgkin's lymphoma and non-Hodgkin's lymphoma, including B-cell lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, marginal zone B-cell lymphoma , T-cell lymphoma, and Waldenstrom macroglobulinemia; said sarcoma is selected from the group consisting of osteosarcoma, Ewing sarcoma, leiomyosarcoma, synovial sarcoma, soft tissue sarcoma, angiosarcoma, liposarcoma, fibrosarcoma, rhabdomyosarcoma , and chondrosarcoma. Preferably, the tumor is selected from lymphoma, non-small cell lung cancer or soft tissue sarcoma. In a specific embodiment of the present invention, the tumor is lymphoma.

The autoimmune disease described in the present invention is selected from organ-specific autoimmune disease and systemic autoimmune disease. Among them, the organ-specific autoimmune disease is selected from chronic lymphocytic thyroiditis, hyperthyroidism, insulin-dependent diabetes mellitus, myasthenia gravis, ulcerative colitis, pernicious anemia with chronic atrophic gastritis, pulmonary hemorrhage nephritic syndrome, common Pemphigus, pemphigoid, primary biliary cirrhosis, multiple sclerosis, acute idiopathic polyneuritis, etc. The systemic autoimmune disease is selected from systemic lupus erythematosus, rheumatoid arthritis, cutaneous rheumatoid nodules, arteritis, pericarditis, scleritis, lymphadenitis, hepatosplenomegaly, neuropathy, systemic vasculitis , scleroderma, pemphigus, dermatomyositis, mixed connective tissue disease, autoimmune hemolytic anemia, thyroid autoimmune disease, or ulcerative colitis.

Description of drawings

Hereinafter, embodiments of the present invention will be described in detail in conjunction with the accompanying drawings, wherein:

Figure 1: The rapid-ELISA sandwich method detected the distribution of EIF4E2 autoantibodies in the serum of 117 tumor patients. The specific tumor patients were lymphoma, lung cancer and soft tissue sarcoma.

Figure 2: The distribution of EIF4E2 autoantibodies in the immunotherapy effective group and ineffective group, wherein the effective group is the tumor immunotherapy effective group, that is, the response group, and the ineffective group is the tumor immunotherapy ineffective group, that is, the non-response group.

Figure 3: ROC curve for the predictive effect of EIF4E2 autoantibodies on the efficacy of immunotherapy.

Figure 4: Comparison of the relative levels of EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC autoantibodies in the serum of lymphoma patients detected by ELISA sandwich method.

Figure 5: ROC curve for predicting the curative effect of immunotherapy for 57 lymphoma training set patients combined with five autoantibodies including EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC.

Figure 6: EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC five autoantibodies combined to predict the long-term survival of patients receiving immunotherapy in 57 lymphoma training set patients.

Figure 7: Validation ROC curve of the combination of five autoantibodies EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC on the efficacy prediction of immunotherapy in 32 patients with lymphoma test set.

Figure 8: Comparison of the relative levels of EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC autoantibodies in serum of lymphoma patients detected by ELISA sandwich method.

Figure 9: ROC curve for predicting the curative effect of immunotherapy for 57 lymphoma training set patients combined with five autoantibodies including EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC.

Figure 10: EIF4E2, CCDC130, FATE1, LPCAT4, VMAC five autoantibodies combined to predict the long-term survival of patients receiving immunotherapy in 57 lymphoma training set patients.

Figure 11: Validation ROC curve of the combination of five autoantibodies EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC on the efficacy prediction of immunotherapy in 32 patients with lymphoma test set.

Figure 12: Comparison of the relative levels of CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC autoantibodies in serum of lymphoma patients detected by ELISA sandwich method.

Figure 13: ROC curve for predicting the efficacy of five autoantibodies CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC combined with immunotherapy for 57 lymphoma training set patients.

Figure 14: The prediction of the long-term survival of 57 lymphoma training set patients receiving immunotherapy with five autoantibodies, CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC.

Figure 15: Validation ROC curve of five autoantibodies CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC combined to predict the efficacy of immunotherapy in 32 lymphoma test set patients.

Detailed ways

The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.

Example 1

1. Sample collection

Serum samples were collected from the Cancer Hospital of the Chinese Academy of Medical Sciences. The median age of cancer patients was 34 (18-74) years old, and the male to female ratio was 71:46, including 89 patients with lymphoma, 16 patients with non-small cell lung cancer, and 12 patients with soft tissue sarcoma , all patients obtained informed consent. The tumor diagnosis results were confirmed by pathological results, all patients received PD-1 antibody immunotherapy, and curative effect evaluation information was obtained.

Among them, the median age of 89 lymphoma patients was 34 (18-69) years old, and the ratio of male to female was 52:37. The grouping information is as follows:

1) According to the Response Evaluation Criteria for Solid Tumors (RECIST 1.1), the curative effect of lymphoma treatment is divided into four grades, CR (Complete Remission), PR (Partial Response), SD (Stable Disease), and PD (Progressive Disease). reduce. In order to evaluate the association between autoantibodies and the efficacy of immunotherapy in patients with lymphoma, combined with previous literature reports and actual efficacy, 3 (CPLX2, EIF4E2, DDX49, PHACTR1, VMAC), 4.5 (EIF4E2, CCDC130, FATE1, LPCAT4, VMAC), 6 (EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC) within 6 (EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC) patients who sustained CR, PR, and SD were classified into the effective group, while patients with PD within 3 months were classified into the ineffective group.

2) The serum of 89 patients was randomly divided into training set (57 cases) and test set (32 cases).

2. Detection method

Proteins detected: CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4, and/or VMAC

Procedure: A rapid enzyme-linked immunosorbent assay (rapid-ELISA) was performed to assess the concentration of serum autoantibodies. Coat capture antibody IgG, anti-GST antibody (10ng/μL*50μL) in a 96-well plate, wash overnight at 4°C, and block with 50μL of PBST-diluted milk buffer (milk 5%, Tween 0.2%) 2 At the same time, the corresponding plasmid (1ng/μL*50μL) was coated in a 96-well plate, washed overnight at 4°C, added to the in vitro expression system IVTT, protected from light at 37°C for 1.5 hours, diluted with milk, and added to the 96-well plate. After reacting for 1 hour at room temperature, wash. Serum samples were diluted in dilution buffer (1:300 dilution), and 50 μL diluted samples/well were added to a 96-well microtiter plate, incubated at 37° C. for 1 hour, and then washed. Add 50 μL of freshly diluted anti-human IgG HRP enzyme-labeled antibody (1:8000 dilution) to each reaction well, incubate at 37°C for 1 hour, and wash. Then, 0.1 mL of temporarily prepared TMB substrate solution was added to each reaction well, and the color was developed at 37°C in the dark for 25 minutes. 50 μL of 0.05 M sulfuric acid was added to each well to terminate the reaction, and the signal was determined by measuring the absorbance at 450 nm.

3. Statistical analysis

The Mann–Whitney U Test was used to compare the differences in variables between groups, and P<0.05 was considered statistically significant.

4. Experimental results

1) This method can successfully detect the level of autoantibodies in the serum of tumor patients

Using the above enzyme-linked immunosorbent assay conditions, the expression levels of EIF4E2 autoantibodies in serum samples from 89 cases of lymphoma, 16 cases of non-small cell lung cancer, and 12 cases of soft tissue sarcoma tumors were detected, and the serum levels of EIF4E2, CCDC130, UBALD1, LPCAT4, VMAC five autoantibodies levels, EIF4E2, CCDC130, FATE1, LPCAT4, VMAC five autoantibody levels and CPLX2, EIF4E2, DDX49, PHACTR1, VMAC five autoantibody levels. The results showed that the above enzyme-linked immunosorbent assay method could successfully detect the relative levels of autoantibodies in the serum of tumor patients. Antibody (see Figure 4), five autoantibodies of EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC (see Figure 8), and five autoantibodies of CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC (see Figure 12) were different, and the patients' serum Different autoantibody levels vary.

2) Relationship between serum autoantibody levels and immunotherapy efficacy in tumor patients

How to more effectively distinguish effective and ineffective patients in tumor immunotherapy is a major clinical problem that needs to be solved urgently. Therefore, the OD value was first standardized, and serum CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2 were detected in lymphoma patients , CCDC130, LPCAT4 and/or VMAC autoantibody levels in relation to PD1 immunotherapy.

Analysis of the single index EIF4E2: 89 cases of lymphoma patients were analyzed, and the results are shown in Figure 2. The results showed that the level of EIF4E2 autoantibody in patients with good treatment effect was higher than that in the non-responding group.

Analysis of five autoantibodies EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC: The training set of 57 patients was analyzed, and the results are shown in Figure 5. The ability, the area under the curve can reach 0.75. Furthermore, as shown in Figure 6, the survival status of effective and ineffective patients predicted by the combination of indicators in 57 patients was found to be longer in effective patients, indicating that the combination of the five autoantibodies has a certain long-term prognostic ability.

Analysis of five autoantibodies EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC: The training set of 57 patients was analyzed, and the results are shown in Figure 9. The ability, the area under the curve can reach 0.73. Furthermore, as shown in Figure 10, the combination of indicators predicted the survival status of effective and ineffective patients in 57 patients, and it was found that effective patients had a longer survival period, indicating that the combination of the five autoantibodies has certain long-term prognostic capabilities.

Analysis of five autoantibodies CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC: The training set of 57 patients was analyzed, and the results are shown in Figure 13. The combination of serum autoantibody levels in lymphoma patients can predict PD-1 immunotherapy The ability, the area under the curve can reach 0.77. Furthermore, as shown in Figure 14, the combination of indicators predicted the survival status of effective and ineffective patients in 57 patients, and it was found that effective patients had a longer survival period, indicating that the combination of the five autoantibodies has a certain long-term prognostic ability.

3) The ROC curve of the predictive effect of autoantibodies on the curative effect of immunotherapy.

The area under the curve can reach 0.73 when the EIF4E2 index alone is used to distinguish patients with different curative effects (see Figure 3). Using five autoantibodies EIF4E2, CCDC130, UBALD1, LPCAT4, and VMAC to verify 32 patients in the test set, the results are shown in Figure 7, which confirms the effectiveness of the combined detection of these five autoantibodies in predicting the efficacy of lymphoma immunotherapy effect, the area under the curve can reach 0.70. Using five autoantibodies EIF4E2, CCDC130, FATE1, LPCAT4, and VMAC to jointly verify 32 patients in the test set, the results are shown in Figure 11, which confirms the role of combined detection of these five autoantibodies in predicting the efficacy of lymphoma immunotherapy. The area under the curve can reach 0.62. The five autoantibodies CPLX2, EIF4E2, DDX49, PHACTR1, and VMAC were used to verify the combination of 32 patients in the test set. The results are shown in Figure 15, which confirmed the role of the combined detection of these five autoantibodies in predicting the efficacy of lymphoma immunotherapy. The area under the curve can reach 0.69. It can be seen that the levels of serum CPLX2, DDX49, PHACTR1, FATE1, UBALD1, EIF4E2, CCDC130, LPCAT4, and VMAC autoantibodies are potential markers for tumor immunotherapy.

The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention. These simple modifications All belong to the protection scope of the present invention.

In addition, it should be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable way if there is no contradiction. The combination method will not be described separately.

Claims (8)

1. The application of autoantibodies in the preparation of products for diagnosing tumors or evaluating the prognosis of tumor treatments, characterized in that, the tumor is selected from lymphoma, and the autoantibodies are selected from one of the following groups: (1) EIF4E2; (2) EIF4E2, CCDC130, UBALD1, LPCAT4 and VMAC; (3) EIF4E2, CCDC130, FATE1, LPCAT4 and VMAC; (4) CPLX2, DDX49, PHACTR1, EIF4E2 and VMAC; (5) EIF4E2, CCDC130, LPCAT4 and VMAC; (6) EIF4E2 and UBALD1; (7) EIF4E2 and FATE1; (8) EIF4E2 and VMAC. 2. The application according to claim 1, wherein the autoantibody comprises one or both of autoantibody subtypes IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgM, IgE or IgD A combination of the above; the autoantibody is an autoantibody in serum, plasma, interstitial fluid, cerebrospinal fluid or urine. 3. The application according to claim 1, characterized in that the prognosis assessment of the tumor treatment is the use of immune checkpoint inhibitors, or the combined treatment of immune checkpoint inhibitors and other drugs, radiotherapy and chemotherapy. post-prognosis assessment. 4. The application according to claim 3, wherein the immune checkpoint inhibitor is selected from PD-1, PD-L1, CTLA-4, BTLA, TIM-3, LAG-3, TIGIT, LAIR1 , 2B4 or CD160 inhibitors. 5. The application according to claim 1, characterized in that said product includes a reagent for detecting autoantibodies. 6. application according to claim 5, is characterized in that, the method for detecting autoantibody of the reagent of described detection autoantibody is selected from ELISA, immunoblotting, indirect immunofluorescence, enzyme immunospot method or immunoluminescent method one or a combination of two or more. 7. The application according to claim 6, wherein the ELISA is rapid-ELISA. 8. The application according to any one of claims 5-7, wherein the detection of autoantibodies is the detection of the presence or expression level of autoantibodies.

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