A marker group for nasopharyngeal carcinoma screening and diagnosis and use thereof

Abstract

The application discloses a marker group for nasopharyngeal carcinoma screening and diagnosis and application thereof, and belongs to the field of molecular biological technology. The marker group comprises at least four polypeptides shown in sequences SEQ ID NO. 1-36. The marker group constructed by the application has more excellent sensitivity and specificity when used for the auxiliary diagnosis and early screening of nasopharyngeal carcinoma. The auxiliary diagnosis results of the marker group selected by the application are all better than common MSCT (the sensitivity and specificity are 84.90% and 72.22% respectively). The early screening results are all better than the most widely used and most mature nasopharyngeal carcinoma diagnosis marker, EBV related marker.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of molecular biology technology, and particularly relates to a marker group for nasopharyngeal carcinoma screening and diagnosis and use thereof. BACKGROUND

[0002] Nasopharyngeal carcinoma (NPC) is one of the local malignant tumors prevalent in East Asia and Southeast Asia. At present, the diagnosis and follow-up of conventional nasopharyngeal carcinoma mainly relies on pathological tissue, serum tumor markers and medical imaging, etc. Among them, pathological examination as the "gold standard" for diagnosing nasopharyngeal carcinoma is difficult to operate and is a painful invasive detection method for patients, and it is difficult to carry out in the early screening of nasopharyngeal carcinoma. Nasopharyngeal imaging examination is very important for the diagnosis and clinical staging of nasopharyngeal carcinoma. CT is helpful to understand the NPC tumor involvement of the surrounding bone structure, such as invasion of the skull base, orbital cavity, etc., but the sensitivity of CT for the diagnosis of early nasopharyngeal carcinoma is not as good as that of MR. For early occult nasopharyngeal carcinoma, both endoscopy and CT examination may have missed diagnosis.

[0003] Nonkeratinizing nasopharyngeal carcinoma is the main type of nasopharyngeal carcinoma in China, and Epstein-Barr virus (EBV) infection is the main pathogenic factor of nonkeratinizing nasopharyngeal carcinoma. Almost all nonkeratinizing nasopharyngeal carcinoma patients have EBV infection. Therefore, EBV-related markers are the main component of nasopharyngeal carcinoma markers and the most widely used and mature diagnostic and prognostic markers in clinical application. In recent years, it has been found that through quantitative detection of multiple indicators such as EBV nuclear antigen 1 (EBNA1) IgA antibody, EBV capsid antigen (VCA) IgA antibody, EB virus antigen titer and EB virus DNA in the serum of nasopharyngeal carcinoma patients, and comprehensive analysis of the above results, the specificity is high, which has important predictive value for evaluating the risk of NPC in high-risk population. However, although EB virus detection has high sensitivity and specificity, its diagnosis rate is not 100%, that is, the EB virus antibody of nasopharyngeal carcinoma patients is positive, but the patient with EB virus positive is not necessarily nasopharyngeal carcinoma, but only indicates that he has experienced EB virus infection. Generally, the accuracy rate of combined antibody detection is higher, and at present, there is a lack of a unified standard for EB virus antibody detection, so a consensus on the detection method is needed. In early (stage I+II) nasopharyngeal carcinoma, the sensitivity and specificity of EBNA1 / IgA are 77.8% and 96.9%, respectively, while EBV-DNA has high sensitivity in advanced nasopharyngeal carcinoma, but has a high false negative rate in stage I nasopharyngeal carcinoma, so it is more recommended for the auxiliary diagnosis of patients in the late stage.

[0004] Liquid biopsy, as a new diagnostic technology, can overcome the influence of tumor heterogeneity, provide more comprehensive tumor molecular information, and provide strong support for guiding clinical treatment and evaluating prognosis. Studies have shown that peripheral blood EBV-DNA is a reliable early diagnosis indicator of nasopharyngeal carcinoma, and is expected to be applied to population screening in nasopharyngeal carcinoma high-incidence areas. Because the nasopharynx is relatively superficial, other researchers have attempted to directly sample nasopharyngeal exfoliated cells. For example, Octavia Rama-yanti et al. collected nasopharyngeal swabs, peripheral blood and pathological tissue specimens from subjects, and detected the EBV-DNA copy number, whole genome methylation state and RNA expression profile of the three samples, and the results showed that the nasal exfoliated cells can more truly reflect the status of the primary cancer, and is a potential minimally invasive sampling method for early screening of nasopharyngeal carcinoma. Gourzones et al. found that miR-BART17 was significantly increased in the plasma samples of NPC patients, and the significant increase of miR-BART17 was accompanied by an increase in the tumor mass of a patient, with a sensitivity of 77% and a specificity of 90%. This suggests that the concentration of miR-BART17 in the plasma may be related to the progression of NPC. Compared with EBV DNA, plasma EBV miRNAs can more directly reflect the activity of the tumor, which may be because EBV-miR-BARTs play a key role in host cell survival, immune escape, cell proliferation, apoptosis and tumor metabolism as viral oncogenes, and promote the occurrence of NPC. However, EBV miRNAs are currently in the experimental stage and have not been verified on a large scale, and there are problems such as the existence of detection quality influencing factors, the lack of evidence from large prospective studies, and the need for standardization of the method. The method needs to be solved urgently. SUMMARY

[0005] In view of the above problems in the prior art, the present application provides a marker group for nasopharyngeal carcinoma screening and diagnosis and its use. In the auxiliary diagnosis and early screening of nasopharyngeal carcinoma, it has excellent sensitivity and specificity, and is expected to be applied to the diagnosis and treatment of nasopharyngeal carcinoma.

[0006] To achieve the above-mentioned purpose, the technical scheme adopted by the present application to solve its technical problems is:

[0007] A marker group for nasopharyngeal carcinoma screening and diagnosis, comprising at least four of the polypeptides shown in SEQ ID NO. 1-36, and the specific sequence is shown in Table 1.

[0008] Table 1 Polypeptide sequence

[0009]

[0010] Among them, the fourth amino acid G in polypeptide 5 has Phospho modification; the third amino acid G in polypeptide 10 has Phospho modification; the second amino acid K in polypeptide 20 has Acetyl modification; the fourth amino acid G in polypeptide 28 has Dehydrated modification; the first amino acid Q in polypeptide 32 has Gln->pyro-Glu modification, and the sixth amino acid N has Dehydrated modification.

[0011] Further, the marker group comprises the polypeptides shown in sequence 1 and sequence 2, and the following polypeptide combinations:

[0012] The polypeptide combination is one of sequence 3 and sequence 4; sequence 5 and sequence 6; sequence 11 and sequence 12; sequence 30 and sequence 35; sequence 3, sequence 4 and sequence 5; sequence 5, sequence 6 and sequence 7; sequence 11, sequence 12 and sequence 20; sequence 9, sequence 30 and sequence 35.

[0013] Further, the marker group comprises the polypeptides shown in sequence 7 and sequence 8, and the following polypeptide combinations:

[0014] The polypeptide combination is one of sequence 3 and sequence 4; sequence 9 and sequence 10; sequence 3, sequence 4 and sequence 9; sequence 9, sequence 10 and sequence 11.

[0015] Further, the marker group comprises the polypeptides shown in sequence 1, sequence 10 and sequence 30, and the polypeptides shown in sequence 35; sequence 20; sequence 35 and sequence 9 or sequence 20 and sequence 22.

[0016] Further, the marker group comprises the polypeptides shown in sequence 5 and sequence 15, and the following polypeptide combinations:

[0017] The polypeptide combination is one of sequence 25 and sequence 35; sequence 2 and sequence 16; sequence 9, sequence 25 and sequence 35; sequence 2, sequence 16 and 20.

[0018] Further, the marker group comprises the polypeptides shown in sequence 4, sequence 8, sequence 19, sequence 20 and / or sequence 22.

[0019] Further, the marker group comprises the polypeptides shown in sequence 7, sequence 10, sequence 17, sequence 18 and / or sequence 22.

[0020] Further, the marker group comprises the polypeptides shown in sequence 2, sequence 4, sequence 9 and sequence 11.

[0021] Further, the marker group comprises the polypeptides as shown in sequence SEQ ID NO. 1~36.

[0022] The use of the marker group in preparation of a preparation for screening and diagnosis of nasopharyngeal carcinoma.

[0023] The use of the marker group in medical basic research for non-diagnostic / treatment purposes.

[0024] Further, the medical basic research is Western Blot, immunohistochemistry or flow cytometry, etc.

[0025] Advantages of the present application:

[0026] The marker combination constructed in the present application has more excellent sensitivity and specificity when used for auxiliary diagnosis and early screening of nasopharyngeal carcinoma, and is expected to be applied to the diagnosis and treatment of nasopharyngeal carcinoma. The auxiliary diagnosis results of the marker combination selected in the present application are all better than ordinary MSCT (the sensitivity and specificity are 84.90% and 72.22%, respectively). The early screening results are all better than the most widely used and maturest nasopharyngeal carcinoma diagnosis marker: EBV-related marker. In early-stage (stage I+II) nasopharyngeal carcinoma, the sensitivity and specificity of EBNA1 / IgA are 77.8% and 96.9%, respectively. The positive rate of EBV-DNA in serum (EBV-DNA≥500 copies / mL) is 89.1% (95% CI: 87.0%~90.9%) and the specificity is 85.0% (95% CI: 83.0%~86.9%). BRIEF DESCRIPTION OF DRAWINGS

[0027] Figure 1 The ROC curve diagram of the marker combination 1;

[0028] Figure 2 The ROC curve diagram of the marker combination 2;

[0029] Figure 3 The ROC curve diagram of the marker combination 3;

[0030] Figure 4 The ROC curve diagram of the marker combination 4;

[0031] Figure 5 The ROC curve diagram of the marker combination 5;

[0032] Figure 6 The ROC curve diagram of the marker combination 6;

[0033] Figure 7 The ROC curve diagram of the marker combination 7;

[0034] Figure 8 The ROC curve diagram of the marker combination 8;

[0035] Figure 9ROC curve plot for marker combination 9;

[0036] Figure 10 ROC curve plot for marker combination 10;

[0037] Figure 11 ROC curve plot for marker combination 11;

[0038] Figure 12 ROC curve plot for marker combination 12;

[0039] Figure 13 ROC curve plot for marker combination 13;

[0040] Figure 14 ROC curve plot for marker combination 14;

[0041] Figure 15 ROC curve plot for marker combination 15;

[0042] Figure 16 ROC curve plot for marker combination 16;

[0043] Figure 17 ROC curve plot for marker combination 17;

[0044] Figure 18 ROC curve plot for marker combination 18;

[0045] Figure 19 ROC curve plot for marker combination 19;

[0046] Figure 20 ROC curve plot for marker combination 20;

[0047] Figure 21 ROC curve plot for marker combination 21;

[0048] Figure 22 ROC curve plot for marker combination 22;

[0049] Figure 23 ROC curve plot for marker combination 23;

[0050] Figure 24 ROC curve plot for marker combination 24;

[0051] Figure 25 ROC curve plot for marker combination 25;

[0052] Figure 26 ROC curve plot for marker combination 26. DETAILED DESCRIPTION

[0053] The specific embodiments of the present application are described below to facilitate the understanding of the present application for those skilled in the art, but it should be clear that the present application is not limited to the scope of the specific embodiments, and for those skilled in the art, it is obvious that various changes are within the spirit and scope of the present application defined and determined by the appended claims, and all the inventions utilizing the concept of the present application are within the scope of protection.

[0054] The patient samples used in the present application are from Zhongshan Hospital Affiliated to Fudan University, and have passed the ethical review.

[0055] The experimental methods used in the present application are as follows:

[0056] I. Serum sample collection

[0057] 1) Sample type: serum.

[0058] 2) Collection requirements: fasting collection, 5 mL of venous blood is drawn using a coagulation tube, and after standing for 30 min, 3000 rpm centrifugation for 15 min, about 1 mL of serum is taken out and placed in a cryopreservation tube.

[0059] 3) Sample storage:

[0060] Used on the same day, the storage condition is 2-8℃;

[0061] If it cannot be used on the same day, it is stored at -20℃, and can be stored for 30 days;

[0062] If stored for a long time (more than one month), it needs to be stored at -80℃.

[0063] Repeated freezing and thawing should not exceed 3 times.

[0064] II. Extraction of the analyte in serum

[0065] 1) After calibrating the mass spectrometer, open the solid-phase biosystem automatic sample analysis system instrument SPS1000 / SPS4000, put in consumables, matched reagent kit and samples to be tested;

[0066] Select the program method "solid-phase sample addition";

[0067] Run the program:

[0068] a. Open the hole;

[0069] b. Take not less than 10µL of serum sample and activated reagent, mix them in a ratio of 1:1, and then place them in the G row reserved hole for use;

[0070] c. Wash the customized suction head in cleaning reagent 1 and cleaning reagent 2 in order. When washing, take not less than 10µL of liquid each time, and repeat the suction and beating not less than 3 times;

[0071] d. Use the cleaned custom tip to process the serum mixture in the G row well. Each time the solution is sucked up no less than 10 µL, and the suction is repeated no less than 3 times;

[0072] e. Use cleaning reagent 3 to clean the custom tip after adsorbing the serum mixture. Each time the solution is sucked up no less than 10 µL, and the suction is repeated no less than 3 times;

[0073] f. Transfer no less than 10 µL of buffer reagent to the H row reserved well, and place the custom tip after using cleaning reagent 3 into the liquid to suck up no less than 10 µL of liquid, and repeat the suction no less than 3 times;

[0074] g. Transfer no less than 10 µL of sample matrix solution to the H row reserved well to complete the sample processing;

[0075] h. Suck up the solution in the H well and spot 2.0 µL on the hydrophobic coated biochip (Wuxi Pimcore Technology Co., Ltd.);

[0076] i. Vacuum dry for 240 s.

[0077] The main components of each reagent are shown in Table 2.

[0078] Table 2. Reagent composition

[0079]

[0080] III. Mass spectrometry data acquisition and uploading

[0081] Place the vacuum-dried hydrophobic coated biochip (Wuxi Pimcore Technology Co., Ltd.) into the mass spectrometer;

[0082] Use the set SP1 voltage (target high voltage), SP2 voltage (pulse high voltage), focusing voltage (lens high voltage), detector voltage (MCP voltage), pulse delay time, acquisition card range, target point diameter, laser frequency, calibration method, and laser intensity to perform data acquisition.

[0083] IV. Quality control

[0084] 1) After data acquisition, upload the data to the mass spectrometry data analysis software;

[0085] 2) The software reads the sample information and signal spectrum, and determines whether the sample and sample pretreatment are qualified according to the quality control model. Unqualified quality control may include various possibilities, including non-disease samples, and signal spectrum intensity does not meet the standard.

[0086] 3) If the quality control is unqualified, according to the quality control results, correct the corresponding parameters and reprocess the serum sample extraction process;

[0087] 4) If the quality control is qualified, enter the next process.

[0088] V. Establishment of positive judgment value and result analysis

[0089] The research of positive judgment value uses nasopharyngeal carcinoma samples with clear diagnostic information and normal samples, covering benign lesion patients such as nasopharyngeal inflammation, adenoid hyperplasia, tuberculosis or lymphoma. The core algorithm is supervised learning based on known nasopharyngeal carcinoma sample atlas. Through a series of processes such as smoothing denoising baseline, screening characteristic peaks, constructing classification model, and calculating the similarity of hormone signal atlas and the known hormone signal atlas stored in the software (Cannataro M, Guzzi P H, Mazza T, et al. Preprocessing, Management, and Analysis of Mass Spectrometry Proteomics Data [J]. 2005.), the maximum Youden index method is used to determine the similarity score of the kit as the positive judgment value. When the similarity score < positive judgment value, the detection result of the sample is negative; when the similarity score ≥ positive judgment value, the detection result of the sample is positive. Taking the maximum similarity score as the positive judgment value, the sensitivity = true positive number / (true positive number + false negative number) × 100%, and the specificity = true negative number / (true negative number + false positive number) × 100% when assisting in the diagnosis of nasopharyngeal carcinoma or early screening of nasopharyngeal carcinoma.

[0090] Example 1. Screening and identification of markers

[0091] The application obtains 36 polypeptide substances in blood with nasopharyngeal carcinoma diagnostic ability by time-of-flight mass spectrometry on 350 normal human samples (178 males (50.9%), 172 females (49.1%), age distribution range: 25-68 years old, average age: 45.8±11.6 years old. Specific age distribution: 58 cases of 25-34 years old, 87 cases of 35-44 years old, 108 cases of 45-54 years old, 97 cases of 55-68 years old), 350 nasopharyngeal carcinoma samples (246 males (70.3%), 104 females (29.7%), age distribution range: 30-65 years old, average age: 50.3±9.8 years old. Specific age distribution: 63 cases of 30-39 years old, 128 cases of 40-49 years old, 126 cases of 50-59 years old, 33 cases of 60-65 years old), first-order mass spectrometry, comprehensive consideration of the relative abundance difference of the characteristic peak data in normal people and nasopharyngeal carcinoma patients, statistical difference of data (p<0.05, t test), influence factor sorting of feature screening by machine learning algorithm (random forest) and matching degree of data in database, 36 polypeptide substances in blood with nasopharyngeal carcinoma diagnostic ability are found. The mass-to-charge ratio (m / z) of these polypeptides, relative abundance and influence factor are shown in Table 1. The relative abundance is normalized based on the normal sample, that is, ln (average signal intensity of cancer patient sample / average signal intensity of normal sample). The screening of characteristic peaks in machine learning is based on the feature importance evaluation of ensemble learning. By constructing multiple decision trees, the contribution of each mass-to-charge ratio (m / z) peak in classification / prediction is quantified. The feature importance score, that is, the influence factor, is obtained by calculating the average reduction of impurity brought by the feature at all tree node splits (Biau, G., Scornet, E. A random forest guided tour. TEST 25, 197-227 (2016). https: / / doi.org / 10.1007 / s11749-016-0481-7). The specific first-order mass spectrometry parameters are as follows:

[0092] Ionization method: matrix-assisted laser desorption ionization (MALDI), and the matrix is alpha-cyano-4-hydroxycinnamic acid (CHCA).

[0093] Mass range: 100-4000 Da.

[0094] Resolution: 20000 (full mass range).

[0095] Laser energy: 30-40%.

[0096] Acquisition mode: positive ion mode.

[0097] Calibration: external mass calibration is performed using peptide standard (Bruker Peptide Calibration Standard).

[0098] Subsequently, the sequences of the 36 substances in the clinical serum were confirmed by secondary mass spectrometry (secondary mass spectrometry (MS / MS or TOF / TOF) is the recommended method for polypeptide identification by the China Food and Drug Administration and the U.S. Food and Drug Administration (FDA) guidelines). The secondary mass spectrometry data analysis process is as follows:

[0099] Data analysis method: Database search was performed using Mascot software (version 2.8), and the database was the UniProt human proteome database (released in 2023). Search parameters: enzyme setting "no enzyme digestion", parent ion mass error allowed ±0.5 Da, fragment ion mass error allowed ±0.3 Da, fixed modification cysteine urea methylation, variable modification methionine oxidation.

[0100] Sequence confirmation standard: The confirmation of polypeptide sequence is based on the matching of fragment ion spectrum (b- and y-ions) with theoretical spectrum, and Mascot score higher than 30 (p<0.05) is considered significant.

[0101] False positive exclusion: Specificity is verified by reverse database search, and the false positive rate is controlled below 1%.

[0102] Secondary mass spectrometry parameters:

[0103] Collision-induced dissociation (CID).

[0104] Collision energy: 30 eV.

[0105] Fragment ion mass range: 100-3500 Da.

[0106] Data acquisition: At least 1000 laser scans per sample were collected to improve the signal-to-noise ratio.

[0107] The sequences and specificities of the 36 markers were confirmed by secondary mass spectrometry, and the false positive problem was excluded. The specific sequences are shown in Table 1.

[0108] Example 2 Verification of marker combination

[0109] According to the 35 polypeptide markers identified and confirmed in Example 1 (see Table 1 for sequences and mass-to-charge ratios), different marker combinations shown in Table 3 were formed, and the sensitivity and specificity of the marker combinations in the auxiliary diagnosis and early screening of nasopharyngeal carcinoma were verified. The analysis process is as follows: for all the samples in the verification cohort, the same MALDI-TOF MS platform and parameters as in Example 1 were used for detection to obtain the mass spectrometric peak intensity data of all markers in Table 1. The entire detection process was carried out in a blind manner, i.e., the experimental operator was unaware of the grouping information of the samples. The raw mass spectrometric data were processed by baseline correction, smoothing and normalization (using the internal standard peak intensity as the reference), and then the peak area or intensity value of each marker was extracted. Statistical analysis was performed using R software (version 4.0.2). The preprocessed marker intensity data were input into the logistic regression (Logistic Regression) model. For auxiliary diagnosis verification, ten-fold cross-validation was performed using all samples in the cohort (Sun T, Liu J, Yuan H, Li X, Yan H. Construction of a risk prediction model for lung infection after chemotherapy in lung cancer patients based on the machine learning algorithm. Front Oncol. 2024 Aug 9;14:1403392. doi: 10.3389 / fonc.2024.1403392. PMID: 39184040; PMCID: PMC11341396.). For early screening verification, due to the uneven sample size, the SMOTE (Synthetic Minority Over-sampling Technique) technique was used to process the data before model training and testing (van den Goorbergh R, van Smeden M, Timmerman D, Van Calster B. The harm of class imbalance corrections for risk prediction models: illustration and simulation using logistic regression. J Am Med Inform Assoc. 2022 Aug 16;29(9):1525-1534. doi: 10.1093 / jamia / ocac093. PMID: 35686364; PMCID: PMC9382395.).The data analysis procedure referred to general guidelines for clinical prediction model construction (Zweig MH, Campbell G. Receiver-operating characteristic (ROC) plots: a fundamental evaluation tool in clinical medicine. Clin Chem. 1993 Apr;39(4):561-77. Erratum in: Clin Chem 1993 Aug;39(8):1589. PMID: 8472349.). Sensitivity, Specificity and Area under the Receiver-Operating Characteristic Curve (AUC) were calculated for each marker combination. The detailed calculation results of performance indicators are shown in Table 3, and the corresponding ROC curves are shown in Figures 1-26 , where AUC is the area under the curve, sens is the sensitivity value, and spec is the specificity value.

[0110] 1) 350 cases of nasopharyngeal carcinoma patients (246 cases of male (70.3%), 104 cases of female (29.7%), age range 32-64 years old, average age 49.8±9.5 years old. The specific distribution is: 58 cases of 32-39 years old, 128 cases of 40-49 years old, 126 cases of 50-59 years old, 38 cases of 60-64 years old) and 350 cases of healthy people samples (178 cases of male (50.9%), 172 cases of female (49.1%), age range 25-65 years old, average age 44.3±11.2 years old. The specific distribution is: 63 cases of 25-34 years old, 98 cases of 35-44 years old, 108 cases of 45-54 years old, 81 cases of 55-65 years old) were used to verify the auxiliary diagnosis of nasopharyngeal carcinoma by each marker combination.

[0111] 2) 400 cases of nasopharyngeal carcinoma patients (282 cases of male (70.5%), 118 cases of female (29.5%), age range 30-63 years old, average age 48.6±9.7 years old. The specific distribution is: 63 cases of 30-38 years old, 128 cases of 39-47 years old, 143 cases of 48-56 years old, 66 cases of 57-63 years old) and 4000 cases of healthy people samples (2036 cases of male (50.9%), 1964 cases of female (49.1%), age range 22-68 years old, average age 43.8±12.4 years old. The specific distribution is: 523 cases of 22-31 years old, 812 cases of 32-41 years old, 1087 cases of 42-51 years old, 987 cases of 52-61 years old, 591 cases of 62-68 years old) were used to verify the early screening of nasopharyngeal carcinoma by each marker combination.

[0112] Table 3 Sensitivity and specificity of marker combination for auxiliary diagnosis and early screening

[0113]

[0114] Table 1

[0115]

[0116] Table 1

[0117]

[0118] According to the detection results of Table 3 and Figures 1-26 The auxiliary diagnosis results of the marker combination selected in the application are better than those of ordinary MSCT (the sensitivity and specificity are 84.90% and 72.22%, respectively). The early screening results are better than the most widely used and maturest diagnostic marker of nasopharyngeal carcinoma: EBV-related marker. In early (stage I+II) nasopharyngeal carcinoma, the sensitivity and specificity of EBNA1 / IgA are 77.8% and 96.9%, respectively. The sensitivity and specificity of EBV-DNA positive (EBV-DNA≥500 copies / mL) in serum for the diagnosis of nasopharyngeal carcinoma are 89.1% (95%CI: 87.0%-90.9%) and 85.0% (95%CI: 83.0%-86.9%), respectively.

[0119] Finally, it should be noted that the above specific embodiments are only used to illustrate the technical solutions of the application and are not limiting. Although the application has been described in detail with reference to examples, those skilled in the art should understand that the technical solutions of the application can be modified or replaced by equivalents without departing from the spirit and scope of the technical solutions of the application, and they should be covered in the scope of the claims of the application.

Claims

1. A marker group for nasopharyngeal carcinoma screening, diagnosis, characterized in that, The marker group is selected from one of the following polypeptide combinations: A) Sequence 1, Sequence 2, Sequence 3 and Sequence 4; B) Sequence 1, Sequence 2, Sequence 5 and Sequence 6; C) Sequence 1, Sequence 2, Sequence 11 and Sequence 12; D) Sequence 1, Sequence 2, Sequence 30 and Sequence 35; E) Sequence 1, Sequence 2, Sequence 3, Sequence 4 and Sequence 5; F) Sequence 1, Sequence 2, Sequence 5, Sequence 6 and Sequence 7; G) Sequence 1, Sequence 2, Sequence 11, Sequence 12 and Sequence 20; H) Sequence 1, Sequence 2, Sequence 9, Sequence 30 and Sequence 35; I) Sequence 2, Sequence 16, Sequence 5 and Sequence 15; J) Sequence 2, Sequence 16, Sequence 20, Sequence 5 and Sequence 15; K) Sequence 2, Sequence 4, Sequence 9 and Sequence 11; L) Sequence 1~Sequence 36; The amino acid sequences of the Sequence 1~Sequence 36 are shown in SEQ ID NO. 1~36; Wherein, the fourth amino acid G in Sequence 5 has Phospho modification; the third amino acid G in Sequence 10 has Phospho modification; the second amino acid K in Sequence 20 has Acetyl modification; the fourth amino acid G in Sequence 28 has Dehydrated modification; the first amino acid Q in Sequence 32 has Gln->pyro-Glu modification, and the sixth amino acid N has Dehydrated modification.

2. Use of the reagent for detecting the marker group of claim 1 in the preparation of a preparation for screening and diagnosing nasopharyngeal carcinoma.

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