KR20240063787A - Biomarker composition for diagnosing biliary tract cancer and method of providing information for diagnosing biliary tract cancer using the same - Google Patents

Abstract

The present invention relates to a biomarker composition for diagnosing biliary tract cancer and a method of providing information for diagnosing biliary tract cancer using the same. More specifically, one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A and PON1, or this It relates to a biomarker composition for diagnosing biliary tract cancer containing a coding gene and a method of providing information for diagnosing biliary tract cancer using the same.
Using the biomarker composition according to the present invention, biliary tract cancer can be diagnosed more effectively, and early diagnosis can help treat biliary tract cancer.

Description

Biomarker composition for biliary tract cancer diagnosis and method of providing information for biliary tract cancer diagnosis using the same {BIOMARKER COMPOSITION FOR DIAGNOSING BILIARY TRACT CANCER AND METHOD OF PROVIDING INFORMATION FOR DIAGNOSING BILIARY TRACT CANCER USING THE SAME}

The present invention relates to a biomarker composition for diagnosing biliary tract cancer and a method of providing information for diagnosing biliary tract cancer using the same. More specifically, one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A and PON1, or this It relates to a biomarker composition for diagnosing biliary tract cancer containing a coding gene and a method of providing information for diagnosing biliary tract cancer using the same.

The bile duct is a tube that carries bile produced in the liver to the duodenum. When it leaves the liver, the left and right bile ducts come together and merge into one stem. Bile ducts are divided into intrahepatic bile ducts, which pass through the liver, and extrahepatic bile ducts, which leave the liver and extend to the duodenum. Among the extrahepatic bile ducts, the sac that temporarily stores and concentrates bile is called the gallbladder. These intrahepatic and extrahepatic bile ducts and the gallbladder are collectively called the biliary tract, but usually the words bile duct and bile duct are used interchangeably.

Cholangiocarcinoma, also called cholangiocarcinoma, is a malignant tumor that occurs in the epithelium of the bile duct. Depending on the site of occurrence, it is classified into intrahepatic biliary cancer and extrahepatic biliary cancer. Since biliary tract cancer often spreads as if seeping into surrounding tissues and does not form a clear tumor mass, it is not easy to accurately diagnose biliary tract cancer. Recently, with the development of diagnostic imaging technology, abdominal ultrasonography, computed tomography (CT), magnetic resonance imaging (MRI), percutaneous transhepatic cholangiography (PTC), percutaneous transhepatic biliary drainage (PTBD), and endoscopic retrograde cholangiopancreatography ( Biliary tract cancer can be diagnosed using techniques such as ERCP or angiography, but it has the disadvantages of being expensive and complicated, and making early diagnosis difficult.

Therefore, there has been a demand for the development of a biomarker that can easily diagnose biliary tract cancer at an early stage.

Republic of Korea Patent Publication No. 10-2017-0124683 (published on November 13, 2017)

The purpose of the present invention is to provide a biomarker composition for diagnosing biliary tract cancer that is excellent in diagnosing biliary tract cancer.

Another object of the present invention is to provide a method of providing information for diagnosing biliary tract cancer using the biomarker for diagnosing biliary tract cancer.

The technical problems to be achieved by the invention are not limited to the technical problems mentioned above, and other technical problems not mentioned can be clearly understood by those skilled in the art from the description of the present invention.

In order to achieve the above object, the present invention provides a biomarker composition for diagnosing biliary tract cancer, comprising one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1 or a gene encoding the same.

In the present invention, the protein expression levels of ERC1 and CA11 are increased in individuals with biliary tract cancer compared to healthy individuals or individuals with benign biliary diseases.

In the present invention, the protein expression levels of HTRA1, KDM1A, and PON1 are reduced in individuals with biliary tract cancer compared to healthy individuals or individuals with benign biliary diseases.

In addition, the present invention provides a diagnosis of biliary tract cancer, including the step of measuring the expression level of one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1, or genes encoding the same, in biological samples isolated from the test subject. Provides a method of providing information for

In the present invention, the method of providing information for diagnosing biliary tract cancer is performed in a biological sample isolated from the test subject in which the expression level of one or more proteins selected from the group consisting of ERC1 and CA11 or the gene encoding them is determined by healthy individuals or The expression level of one or more proteins selected from the group consisting of HTRA1, KDM1A, and PON1, or the gene encoding them, is upregulated compared to the expression level in an individual with a benign biliary disease If it is downregulated compared to the expression level, the test subject can be judged to have biliary tract cancer.

The present invention can provide a biomarker composition for diagnosing biliary tract cancer that is excellent in diagnosing biliary tract cancer, and can help treat biliary tract cancer by effectively diagnosing biliary tract cancer at an early stage using the biomarker composition.

Additionally, the present invention can provide a method of providing information for diagnosing biliary tract cancer using the biomarker for diagnosing biliary tract cancer.

The effects of the present invention are not limited to the effects mentioned above, and other effects not mentioned will be clearly understood by those skilled in the art from the description of the claims.

Figure 1 schematically shows the experimental process for selecting biliary tract cancer-related biomarkers according to an embodiment of the present invention.
Figure 2 shows peptides and proteins identified through LC-MS/MS data analysis.
Figure 3 verifies the protein identified according to Figure 2 by comparing it with genes registered in Vesiclepedia and ExoCarta.
Figure 4 is data comparing the proteins identified according to Figure 2 with data reported for various carcinomas.
Figure 5 shows in 3D the principal component analysis of samples collected in one embodiment of the present invention.
Figure 6 shows the results of clustering analysis.
Figure 7 shows the results of analyzing differentially expressed proteins for each group.
Figure 8 shows the results of analyzing differentially expressed proteins in the benign group and cancer group.
Figure 9 is a diagram showing a scoring system that can identify biliary tract cancer risk groups.
Figure 10 is a graph showing the results of statistical analysis of biomarkers for diagnosing biliary tract cancer according to an embodiment of the present invention.

The terms used in this specification are general terms that are currently widely used as much as possible while considering the function in the present invention, but this may vary depending on the intention or precedent of a person working in the art, the emergence of new technology, etc. In addition, in certain cases, there are terms arbitrarily selected by the applicant, and in this case, the meaning will be described in detail in the description of the relevant invention. Therefore, the terms used in the present invention should be defined based on the meaning of the term and the overall content of the present invention, rather than simply the name of the term.

Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as generally understood by a person of ordinary skill in the technical field to which the present invention pertains. Terms defined in commonly used dictionaries should be interpreted as having meanings consistent with the meanings they have in the context of the related technology, and unless clearly defined in the present application, should not be interpreted in an ideal or excessively formal sense. No.

The numerical range includes the values defined in the range above. Every maximum numerical limit given throughout this specification includes all lower numerical limits as if the lower numerical limit were explicitly written out. Every minimum numerical limit given throughout this specification includes every higher numerical limit as if such higher numerical limit was clearly written. All numerical limits given throughout this specification will include all better numerical ranges within the broader numerical range, as if the narrower numerical limits were clearly written.

Hereinafter, the present invention will be described in detail.

As used herein, “diagnosis” means confirming the presence or characteristics of a pathological condition. For the purpose of the present invention, diagnosis is to confirm the occurrence of biliary tract cancer. Specifically, in a sample isolated from an individual suspected of having biliary tract cancer, the protein or the gene encoding it is compared to a healthy individual or an individual with a benign biliary tract disease. Biliary tract cancer can be confirmed by checking whether its expression or activity is increased. More specifically, the method is to confirm biliary tract cancer by checking whether the expression or activity of the protein or the gene encoding it is increased in a sample isolated from an individual suspected of having biliary tract cancer compared to a healthy individual or an individual with a benign biliary tract disease. You can.

As used herein, “benign biliary disease” refers to a wide range of non-malignant congenital and acquired diseases that can affect the intrahepatic and/or extrahepatic bile ducts, regardless of the presence or absence of liver parenchymal invasion, and are diseases that can manifest in an acute or chronic clinical course. .

In this specification, “biliary cancer” is an epithelial cell malignant tumor that occurs in various locations in the biliary tract, which is the path through which bile flows from the liver to the duodenum.

In this specification, "biomarker" is an indicator that can detect changes in the body, and is a substance that can confirm the normal or pathological state of a living organism and whether there is a change in this, and includes polypeptides, nucleic acids, lipids, glycolipids, and glycoproteins. , organic biomolecules such as sugars (monosaccharides, disaccharides, oligosaccharides, etc.), and can be used to diagnose biliary tract cancer as in the present invention.

The biomarker according to the present invention shows the same results even in repeated experiments and changes in its expression level show significant results, so it can be viewed as a highly reliable marker, and thus the predicted results can be reasonably trusted.

Biomarker composition for diagnosing biliary tract cancer

The present invention can provide a biomarker composition for diagnosing biliary tract cancer, comprising one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1, or a gene encoding the same, preferably ERC1, CA11, and HTRA1. , it is possible to provide a biomarker composition for diagnosing biliary tract cancer, including KDM1A and PON1 proteins or genes encoding them.

The protein or its gene may be derived from exosomes.

The expression level of the ERC1 and CA11 proteins or genes encoding them may be increased in individuals with biliary tract cancer compared to healthy individuals or individuals with benign biliary diseases.

The expression levels of HTRA1, KDM1A, and PON1 proteins or genes encoding them may be reduced in individuals with biliary tract cancer compared to healthy individuals or individuals with benign biliary diseases.

The present invention can provide a composition for diagnosing biliary tract cancer, which includes as an active ingredient an agent for measuring the expression level of one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1 or the genes encoding them. Preferably, a composition for diagnosing biliary tract cancer may be provided, which includes as an active ingredient an agent for measuring the expression level of the ERC1, CA11, HTRA1, KDM1A, and PON1 proteins or the genes encoding them.

The agent may be a primer, probe, protein or antibody that specifically binds to one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A and PON1 or the gene encoding them.

The present invention can provide a kit for diagnosing biliary tract cancer, including the composition for diagnosing biliary tract cancer.

How to provide information for biliary tract cancer diagnosis

The present invention provides information for diagnosing biliary tract cancer, including the step of measuring the expression level of one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1, or genes encoding the same, in a biological sample isolated from a test subject. Provides a method to provide .

The method of providing information for diagnosing biliary tract cancer includes measuring the expression level of one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1, or the genes encoding them, in biological samples isolated from test subjects. It may further include comparing with an individual having a benign disease.

Preferably, the present invention provides a method for providing information for diagnosing biliary tract cancer, comprising the step of measuring the expression level of ERC1, CA11, HTRA1, KDM1A and PON1 proteins or genes encoding them in biological samples isolated from the test subject. can be provided.

More preferably, the present invention includes the steps of measuring the expression level of ERC1, CA11, HTRA1, KDM1A and PON1 proteins or genes encoding them in a biological sample isolated from a test subject; and comparing the measured expression level with that of a healthy individual or an individual with a benign biliary disease.

The method of providing information for diagnosing biliary tract cancer includes the expression level of one or more proteins selected from the group consisting of ERC1 and CA11 or the genes encoding them in a biological sample isolated from the subject being tested in a healthy individual or a person with a benign biliary tract disease. The expression level of one or more proteins selected from the group consisting of HTRA1, KDM1A, and PON1 or the gene encoding them is upregulated compared to the expression level in the subject, and the expression level of the gene encoding the same is downregulated compared to the expression level in a healthy subject or an subject with a benign biliary disease. If so, it can be determined that the test subject has biliary tract cancer.

The biological sample may be derived from a specific tissue or organ of the test subject. Non-limiting examples of the tissues include connective tissue, skin, muscle, or nervous tissue, and representative examples of organs include the eyes, brain, lungs, liver, spleen, bone marrow, thymus, heart, lymph, blood, bone, cartilage, These include the pancreas, kidneys, gallbladder, stomach, small intestine, testes, ovaries, uterus, rectum, nervous system, glands, and internal blood vessels. More specifically, the biological sample includes body fluid samples, including blood, serum, plasma, lymph, breast milk, urine, feces, ocular fluid, saliva, semen, brain extract (e.g., brain pulverized material), spinal fluid, appendix, These include, but are not limited to, spleen and tonsil tissue extracts and exosomes. Preferably, the biological sample may be exosomes isolated from plasma.

Hereinafter, the present invention will be described in detail through examples to aid understanding. However, the following examples only illustrate the content of the present invention and the scope of the present invention is not limited to the following examples. Examples of the present invention are provided to more completely explain the present invention to those skilled in the art.

<Example 1> Confirmation of biomarkers related to biliary tract cancer

As schematically shown in Figure 1, biliary tract cancer-related biomarkers were discovered following the sample preparation, analysis, and validation steps.

1. Blood collection

After obtaining informed consent from patients with biliary tract cancer and patients with biliary tract diseases diagnosed as common bile duct stones and intrahepatic cholangiolithiasis, 9 ml of blood was provided in EDTA tubes, and the protocol was approved by the Medical Research Ethics Review Committee of Pusan National University Hospital. (IRB no. 2011-014-097)

As shown in Table 1 below, a total of 45 exosome samples (5 of healthy control (HC), 10 of benign, 30 of cancer) were used in the experiment. For the HC group, three men and two women aged between 32 and 82 years; for the Benign group, five men and five women aged between 34 and 85 years; and for the Cancer group, 20 men aged between 45 and 84 years. It was provided to 10 people and 10 women.

Table 1 below shows the characteristics of patients who provided samples.

2. Blood separation

Blood obtained from patients was stored on a shaker at room temperature and separated within 4 hours after blood collection. The blood was centrifuged at 800g for 10 minutes, and the upper layer of plasma was separated, divided into 1.5 ml low-protein adhesion tubes, and stored in an ultra-low temperature freezer at -80°C until use.

3. Removal of albumin and IgG from plasma (pretreatment)

Most of the proteins contained in plasma consist of albumin and IgG, and in order to analyze the significantly different proteomes of cancer patients and diseased patients, albumin and IgG must be removed. Therefore, in this example, plasma was pretreated to be suitable for proteomic analysis using a resin column capable of adsorbing albumin and IgG.

The obtained plasma was quickly dissolved at room temperature within 3 minutes, mixed well with 300 μl of plasma and 9700 μl of resin column attachment buffer, then centrifuged at 300 g for 10 minutes, excluding the remaining cells that had settled. The supernatant was transferred to a new tube, and then centrifuged at 2000 g for 10 minutes. After centrifugation for 30 minutes, the supernatant was transferred to a new tube, excluding dead cells that had settled. After centrifugation at 10000 g for 30 minutes, the supernatant was transferred to a new tube, excluding cell debris that had settled.

4. Isolation of exosomes from plasma

The pretreated plasma was centrifuged at 100,000g at 4°C for 18 hours using an ultra-high-speed centrifuge to remove the supernatant. Add 5 ml of clean physiological saline (PBS) to the sediment below and centrifuge again at 120,000g at 4°C using an ultra-high-speed centrifuge. After centrifugation for 70 minutes to remove the supernatant, the sediment below was obtained and resuspended by adding 150 μl of physiological saline (PBS) to obtain exosomes.

5. Exosome pretreatment

Exosome samples were stored at -20°C using acetone and 1: 5 of healthy control, 10 of benign, 30 of cancer 5 of healthy control, 10 of benign, 30 of cancer 5 of healthy control, 10 of benign, 30 of cancer was mixed at a ratio of 4. Exosome samples were placed at -70°C for 2 hours to precipitate exosomes. After centrifugation at 16,000 g for 5 minutes using Minispin 5452 (Eppendorf), the supernatant was removed. After adding 8 M urea to the precipitated exosomes, the sample was dissolved through an ultrasonic disruptor. Through BCA quantification, dithiothreitol (DTT) at a final concentration of 10 mM was added to 160 μg of exosomes and then subjected to a reduction reaction at 37°C and 450 rpm for 30 minutes in a Thermomixer 5382 (Eppendorf). After the reduction reaction was completed, a final concentration of 25 mM iodoacetamide (IAA) was added and alkylation reaction was performed for 30 minutes at room temperature in the dark. After the alkylation reaction, 100 mM ammonium bicarbonate (ABC) was used to lower the urea concentration below 1M. Trypsin (1:25 = protein:enzyme) was added and reacted at 37°C for 16 hours. The reaction was terminated by adding TFA to a final concentration of 1%. The peptided sample was vacuum dried using a SpeedVac after removing the salt contained in the sample using a SOLA HRP C18 cartridge.

6. TMT labeling & LC fractionation

After dividing 45 exosome samples into 3 sets for analysis using TMT 16-plex, in order to normalize each set, the 134N reagent of TMT 16-plex was applied to each set to equalize the total exosome samples. included. The TMT reagent was prepared by dissolving 25 μL acetonitrile (ACN) in a TMT 16-plex tube. 50 μg of the dried peptide sample was dissolved by adding 100 μL of 100 mM triethylammonium bicarbonate (TEAB), then transferred to a TMT reagent tube and reacted at room temperature for 1 hour. After the reaction, 8 μL of 5% hydroxylamine was added to terminate the reaction. The TMT labeled samples were combined into a new tube and vacuum dried using SpeedVac until ~600 μL remained. The remaining sample was desalted using a SOLA HRP C18 cartridge and then vacuum dried using SpeedVac. The dried peptide sample was dissolved in 10mM ABC and then divided into 24 fractions using LC fractionation. After drying each fractionation sample using SpeedVac, peptide samples were dissolved using 0.1% TFA and MS analysis was performed.

7. LC-MS analysis

Peptide-ylated exosome samples were analyzed using a Thermo Dionex Ultimate 3000 UPLC system and a Thermo Orbitrap Exploris 480 MS. For the LC mobile phase conditions, 0.1% formic acid and 5% DMSO were added to H ₂ O for mobile phase A, and 0.1% formic acid and 5% DMSO were added to 80% ACN for mobile phase B. The flow rate of the mobile phase was 250 nL/min, and the analysis column used was PepMapTM RSLC C18, 75 um x 50 cm to separate peptides. The conditions for ESI spray were spray voltage of 2.5 kV and ion transfer tube temperature of 275°C. MS conditions were MS1 scan range from 350 to 1800, MS1 resolution was 60,000, and HCD collision energy was 32.

8. LC-MS/MS data analysis

3,315 proteins were identified from the human protein sequence (UP000005640) provided by the UniProt database and mass spectral data using Comet. When creating the database, semi-tryptic peptides with a length of 7 to 50 were used, and up to one missed cleavage was allowed. Cysteine alkylation (+57.0214 Da) and methionine oxidation (+15.9949 Da) were considered as peptide modifications. False discovery rate (FDR) was applied at 1% each at the spectrum, peptide, and protein levels. Only spectra with precursor isolation purity of 0.7 or higher were used.

As shown in Figure 2, exosomes were obtained by separating blood plasma, and during quantitative analysis, 25,382 peptides and 3,315 proteins were extracted.

9. Comparison of public extracellular vesicle (EV) proteome data

To determine how much of the proteins identified according to this example were included in the previously known extracellular vesicle proteome, they were verified by comparison with the most widely used EV databases, Vesicle pedia (13,396 genes) and ExoCarta (5,402 genes).

As a result, referring to FIG. 3, out of about 3,300 proteins, about 2,600 are previously reported EV-related proteins, and about 730 were identified only in this example. The reason why the 3,349 proteins in this example is more than the 3,315 proteins in the previous analysis is that during qualitative analysis, peptides of the same type were offset and overlapping proteins were removed, and when analyzing other data, overlapping proteins with different details were removed. The numbers are different because they are maintained.

As a result of gene ontology analysis with proteins identified only in this example, GOBP was enriched with vesicle-related GO terms (e.g., vesicle-mediated transport, endocytosis), and as well known in exosome biology, cell migration / It can be seen that GO terms related to ECM (e.g., cell migration, ECM organization) and GO terms related to immunity (e.g., neutrophil immunity mediated, complement activation) are highly enriched, and in GOCC, the terms seen in GOBP are similarly enriched. You can confirm that it appears.

In addition, we conducted a comparison with the latest papers that studied the cancer-associated vesicle proteome for various cancer types. The above paper analyzed the extracellular vesicle proteome in various bodily fluids such as tissue explants, plasma, serum, and bile duct. The results compared approximately 8,800 proteins identified in all types of samples with 3,300 proteins identified in this example. , As shown in Figure 4, approximately 2,300 proteins were confirmed to overlap, of which approximately 1,100 proteins were confirmed to overlap when compared to the biliary tract cancer bile duct EV proteome. In other words, there were about 2,300 vesicle proteins related to cancer, which matched about 70% of the data in this example.

Referring to Figure 5, when the principal component analysis of each sample was shown in 3D, it can be seen that there was no significant difference between each group, and the difference according to detailed diagnosis was also not significant. In addition, as shown in Figure 6, when checking the cluster, it can be confirmed that no significant separation is achieved even when compared with factors such as each cancer type, AST ALT, age, gender CA19-9, etc.

10. Search for differential expression proteins (DEP)

The difference in expression levels between the experimental and control groups was calculated for the protein quantification values identified in the raw data. The experimental group and control group were defined as cancer patient group and healthy control group, benign tumor patient group and healthy control group, and cancer patient group and benign tumor control group. The difference in expression level was obtained by taking the median value of protein expression level in each patient group. The p-value for the difference in expression level was calculated by comparing it with the null distribution generated through a permutation test. Through this, proteins with an expression level difference of more than 1.3 times and a p-value of less than 0.1 among all proteins were defined as differentially expressed proteins.

Referring to Figure 7, in the comparison between Cancer and HC, there were 102 up genes and 31 down proteins, and in the comparison between Benign and cancer, there were 12 up genes and 3 down proteins.

More specifically, as a result of analysis paying more attention to the difference between benign and cancer, as shown in Figure 8, among the 12 upstream proteomes, two yellow-marked proteomes that were not known to be biomarkers (not in the database) were identified. It is possible that these proteomes have undergone exocytosis. Additionally, the three upstream proteomes were confirmed to be known proteomes in each database. These 15 up- or down-regulated proteomes were significantly different between benign and cancer.

11. Gene Set Enrichment Analysis (GSEA)

GSEA was performed to confirm the biological processes significantly enriched by the identified proteins and differentially expressed proteins.

<Example 2> Comparison of biliary tract cancer diagnostic effects of biomarkers for biliary tract cancer

The biliary tract cancer diagnostic effects of the biliary tract cancer-related biomarkers identified in Example 1 were compared. More specifically, it was verified whether the differentially expressed proteins identified in Example 1 had clinical significance.

1. Development of scoring system

First, subjects with biliary tract cancer and patients with biliary tract diseases were recruited from five university hospitals located in Gyeongsangnam-do, and risk factors for biliary tract cancer were collected from these subjects through a multicenter survey. At this time, the distribution number of the subjects by institution is shown in Table 2, and the verification distribution number by biliary tract cancer type of the subjects is shown in Table 3.

Through correlation analysis between the multi-center survey and the occurrence of biliary tract cancer, a scoring system was developed to identify risk groups by scoring risk factors, and the scoring system is shown in Figure 9.

2. Statistical analysis

In order to verify the clinical significance of the differentially expressed proteins of the biliary tract cancer-related biomarkers identified in Example 1, risk factors were collected from multiple institutions, a scoring system was developed, and statistical analysis was performed to verify the differentially expressed proteins.

Statistical data were expressed as mean ± standard deviation or median and range for continuous variables and as frequencies and percentages for categorical variables. To test independence for the above categorical variables, the chi-square test or Fisher's exact test was used. To test the independence of the continuous variables, the two groups were compared using the independent samples t-test if they followed a normal distribution, and the Mann-Whitney test if they did not. The Shapiro-Walk test was used to test normality.

Univariate and multivariate logistic regression analyzes were performed to analyze prognostic factors affecting the diagnosis of biliary tract cancer. Significant prognostic factors were selected using the backward elimination method, a nomogram was created as the final model, and a scoring system was developed (Figure 9). In order to determine the prediction accuracy of the developed scoring system as a diagnostic tool, the area under the receiver operating characteristic curve was calculated and evaluated as shown in FIG. 10. At this time, the optimal cut-off value was selected to maximize the Youden index. Negative predictive value, sensitivity, positive predictive value, and specificity were displayed together, and 95% CI was calculated using Delong's method. All statistical analyzes were performed using SPSS 26 software and MedCalc statistical software version 20.115. A p value of less than 0.05 was considered statistically significant. The statistical analysis results are shown in Table 4 and Figure 10.

As a result, among the 15 biliary tract cancer-related biomarkers (ANGPTL3, C1S, EML3, GLB1, GPNMB, IGHA2, PIGR, SLC2A3, ATM, ERC1, CA11, ST8SIA3, HTRA1, KDM1A and PON1) identified in Example 1, KDM1A, It was confirmed that CA11, PON1, ERC1, and HTRA1 were effective in diagnosing biliary tract cancer. For reference, CA19-9 is currently most commonly used as a diagnostic marker in biliary tract cancer, so it was used as a positive control. When compared to CA19-9, a conventional biliary tract cancer biomarker, KDM1A, CA11, PON1, ERC1, and HTRA1 were confirmed to have high AUC, diagnostic sensitivity, and specificity as biliary tract cancer biomarkers.

So far, we have looked at specific embodiments of the present invention. A person skilled in the art to which the present invention pertains will understand that the present invention may be implemented in a modified form without departing from the essential characteristics of the present invention. Therefore, the disclosed embodiments should be considered from an illustrative rather than a restrictive perspective. The scope of the present invention is indicated in the claims rather than the foregoing description, and all differences within the equivalent scope should be construed as being included in the present invention.

Claims (5)

A biomarker composition for diagnosing biliary tract cancer, comprising one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A, and PON1, or a gene encoding the same.
According to paragraph 1,
The ERC1 and CA11 are
A biomarker composition for diagnosing biliary tract cancer, characterized in that the protein expression level is increased in subjects with biliary tract cancer compared to healthy subjects or subjects with benign biliary diseases.
According to paragraph 1,
The HTRA1, KDM1A and PON1 are
A biomarker composition for diagnosing biliary tract cancer, characterized in that the protein expression level is decreased in subjects with biliary tract cancer compared to healthy subjects or subjects with benign biliary diseases.
Measuring the expression level of one or more proteins selected from the group consisting of ERC1, CA11, HTRA1, KDM1A and PON1 or genes encoding them in biological samples isolated from the test subject; providing information for diagnosing biliary tract cancer, including; method.
According to paragraph 4,
The method of providing information for diagnosing biliary tract cancer is
In the biological sample isolated from the test subject, the expression level of one or more proteins selected from the group consisting of ERC1 and CA11 or the gene encoding them is upregulated compared to the expression level in a healthy individual or an individual with a benign biliary disease,
When the expression level of one or more proteins selected from the group consisting of HTRA1, KDM1A, and PON1 or the gene encoding them is downregulated compared to the expression level in a healthy individual or an individual with a benign biliary disease,
A method of providing information for diagnosing biliary tract cancer by determining that the test subject has biliary tract cancer.