WO2014190451A1

WO2014190451A1 - Reagent testing kit for aided diagnosis of pulmonary adenocarcinoma patient

Info

Publication number: WO2014190451A1
Application number: PCT/CN2013/000631
Authority: WO
Inventors: 赫捷; 谭锋维; 陈照丽; 孙楠
Original assignee: 中国医学科学院肿瘤医院
Priority date: 2013-05-28
Filing date: 2013-05-28
Publication date: 2014-12-04

Abstract

Disclosed is a reagent testing kit for aided diagnosis of a pulmonary adenocarcinoma patient. The reagent testing kit provided in the present invention comprises a product used for detecting protein marker IDH1, a product used for detecting protein marker CEF, a product used for detecting protein marker CA125, a product used for detecting protein marker CYFRA21-1, and a carrier describing the following function: formula I, where x1 represents the concentration of IDH1, x2 represents the concentration of CEA, x3 represents the concentration of CA125, and x4 represents the concentration of CYFRA21-1. Aided diagnosis of the pulmonary adenocarcinoma patient by employing the reagent testing kit provided in the present invention and following a corresponding diagnostic criterion provides the characteristics of high sensitivity and strong specificity, allows the reliability of a diagnosis result to be far greater than that of employing each of the individual protein markers for diagnosis, and has a significant value and application prospect in diagnosis and treatment of pulmonary adenocarcinoma.

Description

Kit for assisting diagnosis of patients with lung adenocarcinoma

The invention relates to a kit for assisting diagnosis of a patient with lung adenocarcinoma.

Background technique

Lung cancer is a major disease that seriously threatens human life and health. According to the 2008 global statistics published by the World Health Organization (WHO) International Agency for Research on Cancer (IAHO) in 2008, lung cancer is the leading malignant tumor with the highest morbidity and mortality. The number of new cases worldwide is 1,600,000, and the number of deaths is 1,377,000, accounting for 12.7% and 18.2% of all malignant tumors. According to statistics published by the American Cancer Society, it is estimated that in 2012, the number of new cases of lung cancer in the United States will reach 26,000, and the number of deaths due to lung cancer will reach 160,000. In China, the results of the third national death cause review conducted by the Ministry of Health showed that: lung cancer is the leading cause of death in malignant tumors in the sample area, and the crude death rate is 30.83/100,000, of which 41.34/100,000 are male. Women 19.84/100,000; the first cause of cancer death among men and women. With the acceleration of global aging and the increase of environmental pollution, the incidence of lung cancer will continue to rise. Therefore, the demand for the development of diagnosis and treatment of lung cancer must be more and more intense.

Pathologically, lung cancer can be divided into non-small cell lung cancer (NSCLC, about 85%) and small cell lung cancer (SCLC, about 15%). Among them, NSCLC is mainly composed of squamous cell carcinoma (also known as squamous cell carcinoma or lung squamous cell carcinoma), adenocarcinoma (also known as lung adenocarcinoma) and large cell carcinoma. Squamous and adenocarcinomas each account for 40% of NSCLC.

The incidence of lung cancer is relatively hidden. More than 70% of patients have progressed to the advanced stage at the time of treatment, and surgery has been lost. This is also one of the main reasons for the poor efficacy of lung cancer and high mortality. Therefore, early screening and diagnosis have become the top priority in the prevention and treatment of lung cancer. At present, clinically used lung cancer screening and diagnosis methods are mainly imaging techniques such as chest X-ray examination and CT scanning. Although these imaging data play an important role in diagnosis, there are also many limitations, such as high false positive rate, inability to detect recessive lesions, subclinical lesions, and micrometastatic lesions. In addition, the diagnosis of lung cancer also includes invasive procedures such as bronchoscopy and perforating biopsy. These methods take a long time and increase patient suffering. Therefore, the search for non-invasive, non-radiative, low-cost, rapid diagnosis, sensitive and specific screening and diagnosis methods is an urgent need for early diagnosis of cancer.

A tumor marker refers to a biologically active substance produced by a tumor cell or tissue due to abnormal expression of an oncogene or other tumor-related genes and products thereof, or is itself detached from a cancerous tissue, and is positive There is a certain degree of expression or little yield in often organized or benign diseases. It reflects the occurrence and development of cancer, can be detected in tumor tissue, body fluids and excretion, widely used in the diagnosis of tumors, monitoring recurrence, metastasis, prognosis, predictive efficacy and so on. Several major markers that are widely recognized in clinical practice are: Tumor antigen 125 ( CA125 ) for ovarian cancer, antigen 19-9 (CA19-9) for pancreatic cancer, for colorectal cancer. Carcinoembryonic antigen (CEA) and prostate specific antigen (PSA) for prostate cancer. Other cancers, such as breast and lung cancer, still lack credible tumor markers for screening.

The markers that have been used in the clinical diagnosis of lung cancer are only protein markers, which are the following:

(1) CEA: Gold and Freeman first discovered CEA in colorectal cancer in 1965. It is a glycoprotein with a molecular weight of 180KD and a half-life of 3-4 days. It is an antigen produced during embryonic development and drops significantly after birth. Its specificity is not strong, but it can be used for monitoring tumor development, efficacy judgment and prognosis estimation. It is not sensitive to early diagnosis. Although lung cancer patients have elevated, there are high false negatives and false positives, which are not suitable for lung cancer screening. Used alone in the diagnosis;

(2) CA125: Bast first discovered CA125, a macromolecular glycoprotein with a half-life of about 4. 8 days, mainly used in ovarian cancer, and also in lung cancer.

(3) CYFRA21-1 (CYFRA211): CYFRA211 is a water-soluble fragment of cytokeratin 19 with a half-life of about 4 days;

Although the above protein markers have been used for the diagnosis of lung cancer, their diagnostic efficiency is low and they do not meet clinical needs. Therefore, the development of new diagnostic markers and diagnostic methods can not wait.

IDH1 is a NADP ⁺ -dependent isocitrate dehydrogenase located in the cytoplasm and peroxisomes that catalyzes the oxidative decarboxylation of isocitrate to a-ketoglutarate. The inventors of the present invention have found in previous studies that the levels of IDH1 in patients with lung squamous cell carcinoma and lung adenocarcinoma are significantly higher than those in the normal population and benign lung tumors, and can be used as a new diagnostic marker for lung cancer.

Invention disclosure

It is an object of the present invention to provide a kit for assisting in the diagnosis of a patient with lung adenocarcinoma.

The kit for assisting diagnosis of lung adenocarcinoma of the present invention comprises a product for detecting protein marker IDH1, a product for detecting protein marker CEA, a product for detecting protein marker CA125, and for detecting The product of the protein marker CYFRA21-1 and the carrier having the following functional formula are described: P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι χΐ represents the concentration of the protein marker IDH1, χ2 represents the concentration of the protein marker CEA, and χ3 represents the concentration of the protein marker CA125, Χ4 represents the concentration of the protein marker CYFRA21-1.

The protein marker IDH1 may specifically be a protein represented by GENBAM ACCESSION NO. CAG46496. 1 (sequence 1 of the Sequence Listing; GENBA ACCESSION NO. CAG46496. 1; 01: 49456351; linear PRI 16-OCT-2008). The protein marker CEA may specifically be a protein represented by GENBANK ACCESSION NO. CAE75559. 1 (sequence 3 of the sequence listing; GENBANK ACCESSION NO. CAE75559. 1; 01: 38522495; linear PAT 25-NOV-2003). The protein marker CA125 may specifically be a protein represented by GENBANK ACCESSION NO. AAL65133. 2 (GENBANK ACCESSION NO. AAL65133. 2; 01: 24419041; linear PRI 29- OCT-2002). The protein marker CYFRA21-1 may specifically be a protein represented by GENBANK ACCESSION NO. NP-002267. 2 (sequence of sequence listing 2; 01: 24234699; linear PRI 06- MAY-2012).

The unit of the concentration of the protein marker IDH1 is specifically U/L, the unit of the concentration of the protein marker CEA is specifically ng/mL, and the unit of the concentration of the protein marker CA125 is specifically U/mL, The unit of the concentration of the protein marker C YFRA21 - 1 is specifically ng/mL.

The following diagnostic criteria can also be recorded on the carrier: When P 0. 689, the patient to be tested is a candidate lung adenocarcinoma patient, when? <0. At 689, the patient to be tested was a candidate for non-lung adenocarcinoma.

The present invention also protects a product for detecting the protein marker IDH1, a product for detecting the protein marker CEA, a product for detecting the protein marker CA125, a product for detecting the protein marker CYFRA21-1, and a functional formula. And diagnostic criteria vectors for the preparation of kits for the diagnosis of patients with lung adenocarcinoma:

The function is as follows:

P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1, the unit is U/L, and x2 represents the concentration of the protein marker CEA in ng /mL, x3 represents the concentration of the protein marker CA125, the unit is U / mL, x4 represents the concentration of the protein marker CYFRA21-1, the unit is ng / mL;

The diagnostic criteria are as follows: When P^O.689, the patient to be tested is a candidate lung adenocarcinoma patient, when? <0. At 689, the patient to be tested was a candidate for non-lung adenocarcinoma.

The protein marker IDH1 may specifically be a protein represented by GENBAM ACCESSION NO. CAG46496. 1 (sequence 1 of the Sequence Listing; GENBANK ACCESSION NO. CAG46496. 1; 01: 49456351; linear PRI 16-0CT-2008). The protein marker CEA may specifically be a protein represented by GENBANK ACCESSION NO. CAE75559. 1 (sequence 3 of the sequence listing; GENBANK ACCESSION NO. CAE75559. 1; 01: 38522495; linear PAT 25-NOV-2003). The protein marker CA125 may specifically be a protein represented by GENBANK ACCESSION NO. AAL65133. 2 (GENBANK ACCESSION NO. AAL65133. 2; 01: 24419041; linear PRI 29- OCT-2002). The protein marker CYFRA21-1 may specifically be a protein represented by GENBAM ACCESSION NO. NP-002267. 2 (sequence 2 of the Sequence Listing; 01: 24234699; linear PRI 06- MAY-2012).

The product for detecting the protein marker IDH1 according to any of the above may specifically be an IDH1 enzyme-linked immunosorbent assay kit (ELISA Kit; including IDH1 standard) purchased from Wuhan Yousheng Technology Co., Ltd.

The product for detecting the protein marker CEA according to any of the above may specifically be a CEA quantitative assay kit purchased from Roche Diagnostics (Shanghai) Co., Ltd.

The product for detecting the protein marker CYFRA21-1 as described above may specifically be a CYFRA21-1 quantitative assay kit purchased from Roche Diagnostics (Shanghai) Co., Ltd.

The product for detecting the protein marker CA125 according to any of the above may specifically be a CA125 quantitative assay kit of Roche Diagnostics (Shanghai) Co., Ltd.

The invention also protects a method for assisting diagnosis of a patient with lung adenocarcinoma, comprising the following steps: (1) separately detecting the concentration of the protein marker IDH1 in the peripheral venous blood of the patient to be tested, the concentration of the protein marker CEA, and the protein marker CA125. Concentration and concentration of the protein marker CYFRA21-1;

(2) Use the following functional formula for auxiliary diagnosis:

P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1, the unit is U/L, and x2 represents the concentration of the protein marker CEA in ng /mL, x3 represents the concentration of the protein marker CA125, the unit is U / mL, x4 represents the concentration of the protein marker CYFRA21-1, the unit is ng / mL; when P 0. 689, the patient to be tested is a candidate lung gland Cancer patients, when? <0. At 689, the patient to be tested was a candidate for non-lung adenocarcinoma. The protein marker IDH1 may specifically be a protein represented by GENBANK ACCESSION NO. CAG46496. 1 (sequence 1 of the Sequence Listing; GENBANK ACCESSION NO. CAG46496. 1; GI: 49456351; l inear PRI 16-OCT-2008). The protein marker CEA may specifically be a protein represented by GENBANK ACCESSION NO. CAE75559. 1 (sequence 3 of the sequence listing; GENBANK ACCESSION NO. CAE75559. 1; 01: 38522495; l inear PAT 25-NOV-2003) The protein CA125 may specifically be a protein represented by GENBANK ACCESSION NO. AAL65133. 2 (GENBANK ACCESSION Ν0· AAL65133. 2; 01: 24419041; l inear PRI 29- OCT-2002). The protein marker CYFRA21-1 may specifically be GENBANK

ACCESSION NO. NP- 002267. 2 protein (sequence of sequence listing 2; 01: 24234699; l inear PRI 06- MAY-2012).

The product for detecting the concentration of the protein marker IDH1 may specifically be an IDH1 enzyme-linked immunosorbent assay kit (ELISA Kit; including IDH1 standard) purchased from Wuhan Yousheng Technology Co., Ltd.

The product for detecting the concentration of the protein marker CEA may specifically be a CEA quantitative assay kit purchased from Roche Diagnostics (Shanghai) Co., Ltd.

The product for detecting the concentration of the protein marker CYFRA21-1 may specifically be a CYFRA21-1 quantitative assay kit purchased from Roche Diagnostics (Shanghai) Co., Ltd.

The product for detecting the concentration of the protein marker CA125 may specifically be a CA125 quantitative assay kit of Roche Diagnostics (Shanghai) Co., Ltd.

DRAWINGS

Figure 1 shows the R0C curve of the lung adenocarcinoma diagnostic model. The ordinate is sensitivity and the abscissa is specific.

The best way to implement the invention

The following examples are provided to facilitate a better understanding of the invention but are not intended to limit the invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The test materials used in the following examples, unless otherwise specified, were purchased from conventional biochemical reagent stores. In the quantitative tests in the following examples, three replicate experiments were set, and the results were averaged. Sensitivity: The number of cancer patients diagnosed as positive as a percentage of the total number of cancer patients. Specificity: The number of healthy people diagnosed as negative as a percentage of the total healthy population. Example 1. Establishment of the method of the present invention

First, the establishment of the model

A total of 454 plasma samples were obtained from patients with lung adenocarcinoma, and 479 healthy human (normal control) plasma samples were obtained from patients with lung cancer who were diagnosed by pathology in the Department of Thoracic Surgery, Chinese Academy of Medical Sciences, and the normal population who participated in the cancer screening. The experiment was approved by the ethics committee, and the subjects were informed of the informed consent to sign the informed consent form. Among the patients with lung adenocarcinoma, there were 227 males and 227 females, aged 26-86 years, with an average age of 60 years. In the normal population, 241 males and 238 females, aged 41-70 years, mean age It is 56 years old. Plasma samples were obtained before surgery, and the patients did not receive chemotherapy, radiotherapy, intervention, etc. before.

Data processing uses curve expert 1.4 to process standard curves; GraphPad Prism 5.01 to process Column charts; SPSS 13.0 for data fitting, mathematical model; SAS 9.2 for mathematical model checking; MedCalc 9.6.2.0 for calculating model ROC curves and cutoff values; Adobe Illustrator CS3 for image processing.

R0C curve (receiver operating characteristic curve) with sensitivity as the ordinate (Y-axis) and specificity as the abscissa (X-axis). When different thresholds (cutoff) are given, it can be generated. The corresponding coordinate point, the curve generated by the continuous line is the R0C curve. The R0C curve can compare the diagnostic efficiency of different tumor markers, and the larger the area under the curve (AUC), the higher the diagnostic efficiency.

The data was analyzed by Logistic binomial regression of SPSS software, and the model was tested using SAS, and a model was finally determined for the diagnosis of lung adenocarcinoma. The calculated results of the model were plotted on the R0C curve (see Figure 1). The area under the curve was 0.890 (95% confidence interval: 0.858-0.917, /?<0.0001), and the marker with the highest diagnostic efficiency for lung adenocarcinoma - IDH1 (AUC was 0.858) compared to a significant difference (/ 0.001).

Second, the specific description of the model

In step 1, by performing a large amount of data simulation and model establishment on a large number of clinical samples for detection of various protein markers, the functional formula for assisting diagnosis of lung adenocarcinoma is as follows:

p = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1 (unit: U/L), and x2 represents the concentration of the protein marker CEA (in units of Ng/mL), x3 represents the concentration of the protein marker CA125 (unit: U/mL), and x4 represents the concentration of the protein marker CYFRA21-1 (unit: ng/mL). The method for assisting diagnosis of lung adenocarcinoma based on the above functional formula is as follows:

(1) Take the peripheral venous blood in the fasting state of the patient to be tested, let stand at room temperature for lh, then centrifuge at 3000 r/min for 15 min, take the supernatant, dispense 1.5 mL EP tube, and store in -80 °C refrigerator.

(2) detecting the protein marker IDH1 concentration, the protein marker CEA concentration, the protein marker CA125 concentration, and the protein marker CYFRA211 concentration in the sample obtained in the step (1), and then calculating the P value by the above functional formula, when P 0. The patient to be tested at 689 was a candidate lung adenocarcinoma patient. When P < 689, the patient to be tested was a candidate for non-lung adenocarcinoma. Example 2, verification of the method of the invention

The patients used in this example were 150 patients with clinically diagnosed lung adenocarcinoma (peripheral venous blood was obtained before surgery, patients had not received chemotherapy, radiotherapy, intervention, etc.) and 50 healthy people (non-lung adenocarcinoma) Patients), all volunteers with informed consent.

1. Take 5 ml peripheral venous blood in the fasting state of the patient to be tested, let stand for 1 h at room temperature, then centrifuge at 3000 r/min for 15 min, take the supernatant, and dispense in two 1.5 mL EP tubes, and store at -80 °C in the refrigerator.

2. Dissolve the sample from step 1 for 3 h at room temperature, then centrifuge at 3000 r/min for 5 min.

3. The concentration of the protein marker IDH1 (unit: U/L), the protein marker CA125 concentration (unit: U/mL), and the protein marker CYFRA211 concentration (in ng/) in the sample (supernatant) obtained in step 2 mL) and protein marker CEA concentration (in ng/mL).

The protein marker IDH1 may specifically be a protein represented by GENBANK ACCESSION NO. CAG46496. 1 (sequence 1 of the sequence listing; GENBANK ACCESSION NO. CAG46496. 1; 01: 49456351; linear PRI 16-OCT-2008). The protein marker CA125 may specifically be a protein represented by GENBANK ACCESSION NO. AAL65133. 2 (GENBANK ACCESSION NO.

AAL65133. 2; 01 : 24419041; linear PRI 29- OCT- 2002 ). The protein marker CYFRA21-1 may specifically be a protein represented by GENBANK ACCESSION NO. NP-002267. 2 (sequence 2 of the list; 01: 24234699; linear PRI 06- MAY-2012). The protein marker CEA can specifically be a protein represented by GENBANK ACCESSION NO. CAE75559. 1 (sequence 3 of the sequence listing; GENBANK ACCESSION NO. CAE75559. 1; 01 : 38522495; linear PAT 25-NOV- 2003).

IDH1 enzyme-linked immunosorbent assay kit purchased from Wuhan Youersheng Technology Co., Ltd. (ELISA Kit; which includes the IDH1 standard; catalog number E97839Hu 96T) and the concentration of the protein marker IDH1 was determined as described in the kit.

Use the CYFRA21-1 Quantitative Assay Kit (catalog number 11820966122) purchased from Roche Diagnostics (Shanghai) Co., Ltd. and test the protein markers according to the kit instructions.

Concentration of CYFRA 21-1.

The concentration of the protein marker CEA was determined using the CEA Quantitative Assay Kit from Roche Diagnostics (Shanghai) Co., Ltd. and following the instructions in the kit (Cat. No. 11731629322).

The CA125 Quantitative Assay Kit (Cat. No. 11776223822) from Roche Diagnostics (Shanghai) Co., Ltd. was used and the concentration of the protein marker CA125 was measured according to the kit instructions.

4. Substituting the protein marker IDH1 concentration, protein marker CEA concentration, protein marker CA125 concentration and protein marker CYFRA21-1 concentration obtained in step 3 into the functional formula of Example 1, to obtain a P value according to the standard of Example 1. Determine if the patient is a patient with lung adenocarcinoma.

The results of steps 3 and 4 are shown in Tables 1 and 2 (in gender, 1 for men and 2 for women). In Tables 1 and 2, the diagnosis results of the individual protein markers were used as a control for the results of the diagnosis using the functional formula of Example 1. When the protein marker IDH1 was used alone, according to the conventional threshold, when the concentration was 2.19, the patient to be tested was a candidate lung adenocarcinoma patient, and when the concentration was < 2.19, the patient to be tested was a candidate for non-lung adenocarcinoma. When the protein marker CA125 is used alone, according to the conventional threshold, when the concentration is 35, the patient to be tested is a candidate lung adenocarcinoma patient, and when the concentration is <35, the patient to be tested is a candidate for non-lung adenocarcinoma. When the protein marker CYFRA21-1 was used alone, according to the conventional threshold, the patient to be tested was a candidate lung adenocarcinoma when the concentration was 3. 3, and the patient to be tested was a candidate for non-lung adenocarcinoma when the concentration was < 3. 3. When the protein marker CEA is used alone, according to the conventional threshold, when the concentration is 5, the patient to be tested is a candidate lung adenocarcinoma patient, and when the concentration is <5, the patient to be tested is a candidate for a non-lung adenocarcinoma patient.

Table 1 Confirmed lung adenocarcinoma patients (+represents positive diagnosis, - represents negative diagnosis) Lung adenocarcinoma Gender Age CEA IDH1 CA125 CYFRA21- 1 Functional patient concentration Diagnostic concentration Diagnostic concentration Diagnostic concentration Diagnostic P value Diagnosis knot No. Result Result Result Result

151 2 55 1. 49 4. 087686 + 13. 46 1. 1 0. 94878 +

152 1 78 4. 25 0. 24847 40. 61 + 4. 36 + 0. 59965

153 1 78 0. 254 6. 874914 + 8. 8 1. 82 0. 99807 +

154 1 67 14. 96 + 0. 155375 125. 3 + 5. 3 + 0. 99862 +

155 1 69 7. 4 + 2. 027362 9. 29 4. 55 + 0. 67455

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00/ClOZN3/X3d 1 57 11. 09 + - 0. 04643 52. 31 + 2. 05 0. 67962

1 73 16. 08 + 2. 572167 + 59. 91 + 6. 68 + 0. 99599 +

1 54 1. 22 2. 931319 + 34. 38 1. 16 0. 94502 +

2 56 1. 44 2. 065663 7. 68 1. 78 0. 50585

2 75 111. 9 + 2. 600898 + 126. 8 + 6. 5 + 1. 00000 +

2 51 1. 53 5. 034998 + 18. 75 5. 29 + 0. 99402 +

1 56 1. 79 4. 292845 + 78. 57 + 14. 44 + 0. 99993 +

1 62 1. 04 -0. 46873 9. 1 1. 02 0. 03662

1 38 1. 8 2. 98965 + 8. 72 1. 28 0. 77278 +

1 70 1. 88 2. 66188 + 15. 26 7. 47 + 0. 89622 +

2 48 1. 88 2. 116749 16. 93 1. 29 0. 65944

2 63 36. 33 + 3. 509149 + 18. 7 11. 2 + 0. 99555 +

2 75 2. 07 3. 016237 + 31. 9 1. 68 0. 94766 +

2 69 2. 16 1. 971694 10. 94 4. 32 + 0. 62821

1 47 5. 55 + 1. 942698 8. 39 3. 09 0. 56721

2 55 8. 84 + 1. 431389 8. 72 2. 96 0. 43730

1 58 1. 9 2. 092678 3. 67 1. 11 0. 42728

2 66 2. 17 2. 988934 + 18. 3 2. 13 0. 88193 +

2 55 2. 59 3. 720436 + 8. 72 3. 62 + 0. 92687 +

1 51 2. 84 1. 742143 13. 29 0. 963 0. 48076

1 54 2. 62 4. 336663 + 21. 53 2. 16 0. 98177 +

2 67 4. 84 3. 207599 + 42. 4 + 6. 72 + 0. 99102 +

1 76 1. 04 1. 7542 8. 62 1. 66 0. 41398

2 64 4. 87 2. 825291 + 16. 7 3. 18 0. 87594 +

1 77 1. 12 2. 610851 + 4. 18 2. 28 0. 63384

2 74 1. 19 1. 998746 3. 81 1. 27 0. 39905

2 50 4. 87 1. 277136 25. 95 2. 97 0. 63577

1 65 2. 06 4. 644938 + 7. 23 1. 51 0. 96435 +

2 60 1. 24 0. 930916 7. 87 2. 93 0. 22036

2 65 5. 92 + 7. 68965 + 5. 16 2. 27 0. 99935 +

1 76 5. 94 + 3. 907407 + 40. 38 + 9. 06 + 0. 99714 +

1 70 1. 48 2. 087728 7. 71 3. 81 + 0. 58740

2 34 6. 74 + 1. 574456 22. 3 2. 44 0. 66600

2 46 1. 89 1. 971694 7. 87 3. 09 0. 53135

1 66 8. 01 + 2. 058712 44. 3 + 1. 7 0. 93959 +

1 61 8. 1 + 2. 349091 + 19. 35 2. 5 0. 82505 +

Table 2 Confirmed healthy person (+represents positive diagnosis, - represents negative diagnosis) Health Year CEA IDH1 CA125 CYFRA21-1 Functional Human Other Age Concentration Diagnostic Concentration Diagnostic Concentration Diagnostic Concentration Diagnostic P Value Diagnosis No. Result Result Result Result Result

301 1 62 2. 28 0. 553185 7. 66 3. 18 0. 15638

302 1 64 0. 578 0. 011111 5. 47 3. 91 + 0. 07638

303 2 58 3. 23 1. 103938 9. 95 3. 14 0. 31231

304 2 63 5. 72 + 0. 787059 2. 91 5. 06 + 0. 21555

305 2 58 1. 94 1. 115052 15. 44 1. 35 0. 32783

306 1 55 1. 41 1. 580032 13. 52 6. 35 + 0. 61365

307 1 58 17. 89 + 0. 728569 2. 67 0. 648 0. 17631

308 2 58 0. 896 0. 2028 13. 15 2. 75 0. 13197

309 1 55 3. 02 2. 084666 27. 42 2. 78 0. 83112 +

310 1 60 2. 54 -0. 6469 5 2. 4 0. 02892

311 2 56 10. 43 + 2. 46001 + 2. 74 0. 815 0. 60261

312 2 57 1. 39 1. 710904 9. 99 1. 69 0. 42749

313 1 54 3. 68 -0. 05965 11. 07 1. 61 0. 08185

314 1 50 3. 47 1. 021162 6. 35 2. 77 0. 23432

315 2 51 16. 15 + 2. 298696 + 1. 98 1. 21 0. 60948

316 1 55 3. 84 0. 377928 13. 75 1. 91 0. 16485

317 1 65 4. 89 0. 407129 6. 13 3. 57 + 0. 14005

318 1 57 0. 766 1. 30602 10. 71 2. 34 0. 33310

319 1 65 3. 4 0. 298011 7. 97 2. 25 0. 11063

320 1 57 7. 11 + 1. 046244 10 1. 7 0. 28586

321 2 58 9. 28 + -0. 1983 1. 6 2. 14 0. 05050

322 1 61 4. 35 -0. 00135 4. 1 4. 4 + 0. 08488

323 2 54 1. 41 1. 402064 6. 99 2. 15 0. 30485

324 2 56 9. 35 + 0. 377928 1. 32 1. 17 0. 08701

325 1 60 12. 91 + 1. 578068 2. 55 1. 36 0. 36541 326 1 63 1. 66 1. 363802 6. 16 2. 39 0. 29226

327 2 63 3. 5 1. 147969 7. 94 1. 96 0. 26290

328 2 57 8. 4 + 1. 161651 1. 85 2. 58 0. 24277

329 1 62 2. 04 0. 144452 7. 57 4. 22 + 0. 11158

330 2 65 2. 72 0. 774517 7. 39 3. 92 + 0. 21541

331 2 59 2. 69 6. 771228 + 13. 7 2. 77 0. 99875 +

332 1 51 13. 63 + 2. 111876 11. 78 3. 79 + 0. 75859 +

333 2 70 1. 55 0. 430524 8. 56 1. 86 0. 11864

334 1 57 0. 974 0. 69933 10. 21 1. 78 0. 17042

335 1 57 8. 57 + -0. 58369 11. 82 2. 96 0. 06619

336 1 57 6. 97 + 0. 231979 1. 47 0. 902 0. 06517

337 1 63 5. 55 + 1. 075089 19. 07 1. 65 0. 41707

338 1 58 12. 65 + 2. 407226 + 5. 1 1. 95 0. 68218

339 2 70 3. 33 0. 681601 38. 16 + 1. 72 0. 59181

340 1 67 3. 02 1. 161651 11. 85 3. 59 + 0. 37076

341 1 66 1. 76 0. 642938 9. 49 1. 41 0. 15102

342 2 58 1. 03 0. 364447 9. 66 3. 04 0. 13426

343 2 68 1. 98 1. 414274 10. 21 2. 55 0. 37540

344 2 54 1. 15 0. 463013 7. 6 1. 61 0. 11093

345 2 43 0. 817 1. 344979 18. 12 2. 07 0. 45505

346 2 65 4. 78 0. 745316 23. 11 2. 27 0. 39511

347 2 49 0. 957 1. 207906 9. 65 1. 45 0. 26598

348 2 57 1. 19 1. 695267 7. 42 1. 44 0. 37032

349 2 53 1. 35 -0. 28141 11. 56 1. 72 0. 06024

350 2 55 2. 26 0. 37326 12. 4 1. 74 0. 14084 The results were determined according to the functional formula and the diagnostic method of Example 1. The sensitivity was 59.33% and the specificity was 94%. The protein marker IDH1 and its threshold were used for the determination of the results, and the sensitivity was 54.67% and the specificity was 92%. The protein marker CA125 and its threshold value were used for the determination of the results, and the sensitivity was 12.67% and the specificity was 98%. The protein marker CYFRA21-1 and its threshold were used alone for the determination of the results, with a sensitivity of 29.33% and a specificity of 82%. Protein marker alone The CEA and its threshold were judged to have a sensitivity of 36.67% and a specificity of 7%. Taking into account the sensitivity and specificity, the functional formula and diagnostic method of Example 1 is significantly superior to the diagnosis using several separate protein markers.

Industrial application

Lung cancer ranks first in the world for cancer deaths and is the number one cancer killer that harms the health of our people. Therefore, it is a hot spot in recent years to find tumor markers that can be used for early diagnosis, individual treatment, prognosis and monitoring of recurrence of lung cancer. Using the kit provided by the present invention and assisting diagnosis of lung adenocarcinoma patients according to the corresponding diagnostic criteria, the sensitivity and the specificity are high, and the reliability of the diagnosis is much higher than that of the diagnosis using the individual protein markers. It has great value and application prospect for the diagnosis and treatment of lung adenocarcinoma.

Claims

Rights request

1. Kits used to assist in the diagnosis of lung adenocarcinoma patients, including products used to detect protein marker IDH1, products used to detect protein marker CEA, products used to detect protein marker CA125, products used to detect protein markers The products and carriers of CYFRA21-1 have the following functional formula:

P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1, x2 represents the concentration of the protein marker CEA, x3 represents the concentration of the protein marker CA125, x4 represents the concentration of protein marker CYFRA21-1.

2. The kit according to claim 1, characterized in that: the protein marker IDH1 is the protein represented by GENBANK ACCESSION NO. CAG46496.1; the protein marker CEA is the protein represented by GENBANK ACCESSION NO. CAE75559.1 Protein; The protein marker CA125 is the protein shown in GENBANK ACCESSION NO. AAL65133.2; The protein marker CYFRA21-1 is the protein shown in GENBANK ACCESSION NO. NP-002267.2.

3. The kit according to claim 1 or 2, characterized in that: the concentration unit of the protein marker IDH1 is U/L, the concentration unit of the protein marker CEA is ng/mL, and The concentration unit of the protein marker CA125 is U/mL, and the concentration unit of the protein marker CYFRA21-1 is ng/mL.

4. The kit according to claim 3, characterized in that: the carrier also records the following diagnostic criteria: When P 0.689, the patient to be tested is a candidate lung adenocarcinoma patient, when? When < 0.689, the patient to be tested is a candidate non-lung adenocarcinoma patient.

5. Products used to detect the protein marker IDH1, products used to detect the protein marker CEA, products used to detect the protein marker CA125, products used to detect the protein marker CYFRA21-1 and products with documented functional formulas and diagnoses Application of standard vectors in the preparation of kits to assist in the diagnosis of lung adenocarcinoma patients:

The functional formula is as follows:

P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1 in U/L, x2 represents the concentration of the protein marker CEA in ng /mL, x3 represents the concentration of the protein marker CA125 in U/mL, x4 represents the concentration of the protein marker CYFRA21-1 in ng/mL; the diagnostic criteria are as follows: When P

0. When 689, the patient to be tested is a candidate lung adenocarcinoma patient, when? When < 0.689, the patient to be tested is a candidate non-lung adenocarcinoma patient.

6. The application according to claim 5, characterized in that: the protein marker IDH1 is the protein represented by GENBANK ACCESSION NO. CAG46496.1; the protein marker CEA is represented by GENBANK ACCESSION NO. CAE75559.1 The protein; the protein marker CA125 is the protein shown in GENBANK ACCESSION NO. AAL65133.2; the protein marker CYFRA21-1 is the protein shown in GENBANK ACCESSION NO. NP-002267.2.

7. A method for auxiliary diagnosis of patients with lung adenocarcinoma, including the following steps:

(1) Detect the concentration of the protein marker IDH1, the concentration of the protein marker CEA, the concentration of the protein marker CA125 and the concentration of the protein marker CYFRA21-1 in the peripheral venous blood of the patient to be tested;

(2) Use the following functional formula for auxiliary diagnosis:

P = -

1 + (0.031 * 3.622 ^χ ι * 1.041 ^χ 2 * 1.070 ^χ 3 * 1.158 ^χ 4)-ι xl represents the concentration of the protein marker IDH1, the unit is U/L, x2 represents the concentration of the protein marker CEA, the unit is ng /mL, x3 represents the concentration of the protein marker CA125, the unit is U/mL, x4 represents the concentration of the protein marker CYFRA21-1, the unit is ng/mL; when P 0.689, the patient to be tested is a candidate lung gland Cancer patients, when? When < 0.689, the patient to be tested is a candidate non-lung adenocarcinoma patient.

8. The method of claim 7, characterized in that: the protein marker IDH1 is the protein represented by GENBANK ACCESSION NO. CAG46496.1; the protein marker CEA is represented by GENBANK ACCESSION NO. CAE75559.1 The protein; the protein marker CA125 is the protein shown in GENBANK ACCESSION NO. AAL65133.2; the protein marker CYFRA21-1 is the protein shown in GENBANK ACCESSION NO. NP-002267.2.