KR20160021531A - Maker for the diagnosis of gastric cancer - Google Patents

Abstract

The present invention relates to a single and/or composite marker of epithelial-derived neutrophil-activating peptide 78/chemokine (C-X-C motif) ligand 5 (ENA78/CXCL5) or stromal cell-derived factor-1/chemokine (C-X-C motif) ligand 12 (SDF-1/CXCL12) for diagnosing gastric cancer and analyzing the stage of gastric cancer. The marker for diagnosing gastric cancer or the marker for estimating the stage of gastric cancer according to the present invention can be used in diagnosing gastric cancer at an early stage as the marker has greatly improved diagnosis accuracy compared with traditional markers; and can be used in predicting the prognosis of gastric cancer as the marker can provide information on the stage of gastric cancer. Therefore, using the marker according to the present invention can contribute to increasing the survival chance of gastric cancer patients by providing vast information related to gastric cancer.

Description

[0001] The present invention relates to a marker for gastric cancer,

The present invention relates to an ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) or SDF-1 / CXCL12 (stromal cell dependent factor-1 / chemokine (CXC motif) ligand 12).

Gastric cancer is not only the second leading cause of cancer deaths worldwide, but is also well known as one of the major causes of death in Asian countries such as Korea and Japan (CA.Cancer. J. Clin. 2011) 61: 69-90). The primary treatment for gastric cancer is the removal of gastric cancer tissue through surgery, but the risk of recurrence is high enough to recur in more than 50% of patients who undergo resection, except early gastric cancer. Of gastric cancer is 30 to 70%, and in case of third gastric cancer, it is only 5 to 30%. Therefore, the development of a method for early diagnosis of gastric cancer is very important for increasing the survival rate of gastric cancer. In addition, if the stage of stomach cancer can be predicted, it can be very useful for determining the treatment method of the patient or predicting the progress before and after treatment. Therefore, in countries where the incidence of gastric cancer is high, such as Korea and Japan, X-ray and endoscopic diagnosis of stomach cancer are commonly performed. However, diagnosis through X-ray and endoscopy is usually performed when the patient feels abnormal symptoms and is examined. Most of the patients do not have symptoms at the beginning of gastric cancer. Therefore, when the patients feel abnormal symptoms, It is a universal fact that the stage of cancer has already proceeded a lot. Therefore, if biomarkers in the blood can be used to diagnose gastric cancer, it will be useful for early detection and treatment of patients with gastric cancer because it can be easily performed.

Accordingly, a number of studies on biomarkers for gastric cancer diagnosis are under way, and various biomarkers for the diagnosis of gastric cancer such as CEA and AFP have been actively developed using the latest technologies such as genomics and proteomics. However, the development of biomarkers with high diagnostic accuracy to diagnose early gastric cancer has yet to be developed. It is important to determine the treatment method of patients and to predict the stage of the disease before and after treatment. Development of markers is very scarce.

Thus, not only is it possible to diagnose early gastric cancer, but also the diagnostic accuracy of predicting the stage of stomach cancer is improved, and thus there is a demand for the development of commercially available biomarkers for stomach cancer diagnosis or stomach cancer staging.

DISCLOSURE Technical Problem The present invention has been made in order to solve the above problems of the prior art, and it is an object of the present invention to provide an ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 (CXC motif) ligand 5) or SDF-1 / CXCL12 (stromal cell-derived factor-1 / chemokine (CXC motif) ligand 12) and a kit for diagnosing gastric cancer or gastric cancer using the same For that purpose.

However, the technical problem to be solved by the present invention is not limited to the above-mentioned problems, and other matters not mentioned can be clearly understood by those skilled in the art from the following description.

Hereinafter, various embodiments described herein will be described with reference to the drawings. In the following description, for purposes of complete understanding of the present invention, various specific details are set forth, such as specific forms, compositions, and processes, and the like. However, certain embodiments may be practiced without one or more of these specific details, or with other known methods and forms. In other instances, well-known processes and techniques of manufacture are not described in any detail, in order not to unnecessarily obscure the present invention. Reference throughout this specification to "one embodiment" or "embodiment" means that a particular feature, form, composition, or characteristic described in connection with the embodiment is included in at least one embodiment of the invention. Accordingly, the appearances of the phrase " in one embodiment "or" an embodiment "in various places throughout this specification are not necessarily indicative of the same embodiment of the present invention. In addition, a particular feature, form, composition, or characteristic may be combined in any suitable manner in one or more embodiments.

As used herein, the term " level of chemokine "means the concentration, amount, activity, etc. of the chemokine in the sample, and is not limited thereto as long as it can quantitatively compare the chemokine in the sample. The method of measuring the level can be carried out by various immunoassay methods such as radioimmunoassay, radioimmunoprecipitation, immunoprecipitation, enzyme-linked immunosorbent assay (ELISA), and sandwich assay, and the level of chemokine There is no limitation to this.

As used herein, the term "sample " refers to all samples containing chemokines isolated from a patient so that the concentration, amount and activity of the chemokine of the patient can be measured, preferably blood, plasma, serum, Any type of measurable level of chemokine is not limited to this.

As used herein, the term "marker" refers to a molecule quantitatively or qualitatively related to the progress of gastric cancer, and preferably it may be chemokine, protein, nucleic acid, But are not limited to, molecules capable of providing information about diagnosis and / or staging of stomach cancer.

In the present specification, the term "stage" refers to a stage of a disease indicating the progress of gastric cancer, including a cancer size, a penetration depth of a gastric cancer (T-stage), a lymph node metastasis (N-stage ), Distant metastasis, and general cancer stage (stage I, II, III, IV), and may be the TMN stage , And is capable of providing information on the progress of the stomach cancer. Through the staging of the present specification, the progress of gastric cancer can be grasped, and the treatment method can be determined, the prognosis can be judged, and the therapeutic result can be used.

As used herein, the term " information providing method "is a method for acquiring information on factors that may indicate the degree of diagnosis of gastric cancer and / or the degree of progression of gastric cancer. Preferably, , The method of obtaining information on the progression stage (staging) of stomach cancer by estimating the information on whether or not stomach cancer has occurred and the information on the size, invasion depth, and metastasis status of stomach cancer But is not limited to, a method for obtaining information related to gastric cancer from a biological sample.

As used herein, the term "kit" refers to a device for screening that can be used to diagnose gastric cancer or to diagnose stomach cancer, and may be used to measure the level of a single or multiple marker of the present invention from a biological sample There is no restriction in the form. Preferably, it may be of a form capable of measuring the amount of substance bound to the chemokine, comprising a substance specifically binding to the chemokines ENA78 / CXCL5, SDF-1 / CXCL12, and / or CEA, The substance may preferably be an antibody, an aptamer, or the like, but is not limited thereto, as long as it is a substance that specifically binds to ENA78 / CXCL5, SDF-1 / CXCL12, and / or CEA.

The present invention relates to the use of ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) and / or SDF-1 / CXCL12 (stromal cell- CXC motif) ligand 12) and comparing the level of the normal control sample with that of the normal control sample to provide information about the diagnosis of gastric cancer or the stage of gastric cancer. In addition, the marker may further include a marker capable of providing information on stomach cancer diagnosis or stomach cancer staging.

In another embodiment of the present invention, the biological sample may be blood, plasma, serum, and the like. Any sample separated from the patient and containing chemokine is not limited.

The present invention also relates to a method of screening for an ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5), SDF-1 / CXCL12 (stromal cell- CXC motif ligand 12), and carcinoembryonic antigen (CEA), and provides information on the distant metastasis of gastric cancer by comparing the levels of one or more markers selected from the group consisting of CEA (carcinoembryonic antigen) and normal control samples . ≪ / RTI >

The present invention also provides a diagnostic kit for stomach cancer or a stomach cancer staging kit comprising an antibody or an aptamer that specifically binds to ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) do.

In one embodiment of the invention the kit comprises an antibody or aptamer specifically binding to SDF-1 / CXCL12 (stromal cell-derived factor-1 / chemokine (CXC motif) ligand 12) and / or carcinoembryonic antigen or an antibody or an aptamer that specifically binds to the antigen. In addition, it may further include an antibody or an aptamer that specifically binds to a marker capable of diagnosing gastric cancer or staging of gastric cancer.

Since the diagnostic marker for gastric cancer diagnosis according to the present invention or the marker for stomach cancer diagnosis is highly improved compared with the existing markers, it can be used not only for the diagnosis of early gastric cancer but also for predicting the prognosis of gastric cancer. Of gastric cancer patients can be used to provide various gastric cancer-related information, which is expected to increase the survival rate of gastric cancer patients.

1 is a graph showing an ROC curve and an AUC according to an embodiment of the present invention.
FIG. 2 is a graph showing an ROC curve and an AUC according to an embodiment of the present invention.
FIG. 3 is a graph showing an ROC curve and an AUC according to an embodiment of the present invention.

Hereinafter, the present invention will be described in more detail with reference to Examples. It will be apparent to those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Example

Example 1: Preparation of sample

To identify the new markers for gastric cancer diagnosis and stomach cancer diagnosis that can improve the accuracy of diagnosis, we performed gastrointestinal endoscopy and biopsy among patients who visited Yonsei University Health System. Patients with dysplasia or those with diagnosis of gastric cancer were classified as primary. The purpose of the study and the contents of the subjects were fully explained and the study was conducted after receiving the informed consent. Patients with chronic disease (chronic renal disease, liver cirrhosis, diabetes mellitus, etc.) and other cancer diagnoses should be screened for primary diagnosis to improve the accuracy of the test. Patients who have been diagnosed with other stomach tumors (gastrointestinal stromal tumor (GIST), mucosa-associated lymphoid tissue lymphoma, etc.), patients with previous gastric cancer diagnosed and treated Were excluded. Patients who were diagnosed with gastric cancer amongst the second-line patients were identified with staging of gastric cancer by image analysis such as X-ray, CT, and PET. Stomach cancer was classified according to the TMN stage of the 7th International Union Against Cancer, and a total of 300 patients were finally classified into four groups according to the stage of gastric cancer. Patients with intestinal metaplasia and dysplasia were diagnosed with high-risk group, submucosal layer of gastric cancer, early gastric cancer group (EGC), proper (proper) The gastric cancer group was classified as an advanced gastric cancer group (AGC) and normal control group (normal gastric mucosa) or simple gastritis (gastric cancer) Of the patients. Patients who were ultimately categorized were collected and stored as blood samples prior to medical treatment (treatment).

Blood samples from 300 patients were used as initial training dataset with 25 blood samples from each of the four groups, and blood samples from the remaining 200 patients were used as independent validation datasets The 200 patients included 70 controls, 30 high-risk patients, 50 early gastric cancer patients, and 50 advanced gastric cancer patients, respectively.

Some of the blood samples from patients diagnosed with gastric cancer were stained with a Giemsa solution and then examined for infection with Helicobacter pylori .

Example 2: Initial training data < RTI ID = 0.0 >

2.1. Identification of Serum Levels of ENA78 / CXCL5, SDF-1 / CXCL12 and CEA by Stage of Gastric Cancer

Serum concentrations of ENA78 / CXCL5, SDF-1 / CXCL12, and carcinoembryonic antigen (CEA) were measured to determine the changes in serum concentrations of each marker according to the progression of gastric cancer. CEA concentrations were measured using the Beckman Access CEA assay. ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) and SDF-1 / CXCL12 (stromal cell- 1 / chemokine (CXC motif) ligand 12) was measured using the MILLIPLEX MAP Human Cytokine / Chemokine Kit. In order to measure the concentration in the serum, 200 μL of assay buffer was added to each well of the filter plate included in the kit, followed by reaction at room temperature (RT) for 10 minutes 25uL of the standard solution provided in the kit was added to some wells of the prepared filter plate and 25uL of assay buffer was added to the well to which the experimental samples were added. Solution. After 25 uL of the matrix solution was added to both the background well, the standard reagent well and the experimental sample well, 25 uL of the sample prepared by the method of Example 1 was added to the experimental sample well. 25 uL of beads were added to each well followed by shaking at 4 [deg.] C, and after a reaction time of about 16 hours, all of the solution in each well was removed and washed twice. After adding 25 μL of detection antibody to each well and shaking for 1 hour at room temperature, 25 μL of a Streptavidin-phycoerythrin solution was further added to each well, followed by shaking at room temperature for 1 hour . After completion of the reaction, all of the solutions in the wells were removed and washed. 150uL of sheath fluid was added to the washed wells, mixed well, allowed to react for 5 minutes, and then fluorescence was measured using Luminex 100 ™ IS. Serum concentrations of each marker were measured using standard reagents. The results are shown in Tables 1 and 2.

Groups
(n) Normal
(25) High-risk
(25) EGC
(25) AGC
(25) p -value ^† Serum ENA78 (ng / ml) 1008.9 + - 536.7 * 989.6 ± 605.9 2234.3 ± 1103.9 3065.8 ± 1487.3 <0.001 Serum SDF-1 (ng / ml) 1655.5 ± 729.0 1774.9 + 712.7 2762.4 ± 1654.9 4474.2 + 2460.5 <0.001 Serum CEA (ng / ml) 2.2 ± 1.7 3.0 ± 2.6 3.0 ± 3.3 6.1 ± 8.6 0.028

As shown in Table 1, serum concentrations of ENA78 / CXCL5 and SDF-1 / CXCL12 did not show a significant increase until the high-risk group, but increased about 2-fold in the early gastric cancer group (EGC) And the post-hoc Bonferroni method was used to confirm that each p-value was a reliable value of 0.001 or less. On the other hand, CEA, known as a gastric cancer marker, was found to increase only in advanced gastric cancer, and it was found to be lower than ENA78 / CXCL5 and SDF-1 / CXCL12. From these results, it can be seen that ENA78 / CXCL5 and SDF-1 / CXCL12 do not change significantly in serum concentration until the onset of gastric cancer, but the two markers increase rapidly from early gastric cancer, , And it was confirmed that the concentration of serum was significantly increased according to the progression stage of gastric cancer, so that it can be used for the diagnosis of stomach cancer.

Groups
(n) Non-cancer groups
(50) Cancer groups
(50) p -value ^‡ Serum ENA78 (ng / ml) 999.3 + - 566.5 * 2650.0 占 1362.6 <0.001 Serum SDF-1 (ng / ml) 1715.2 + - 716.0 3618.3 ± 2248.1 <0.001 Serum CEA (ng / ml) 2.6 ± 2.2 4.6 ± 6.6 0.051

Furthermore, as shown in Table 2, serum concentrations of ENA78 / CXCL5 and SDF-1 / CXCL12 significantly increased more than 2-fold in gastric cancer group. On the other hand, CEA, which is a conventional marker for gastric cancer diagnosis, showed an increase of about twice the mean value, but it was confirmed that the significance was low. From the above results, it was confirmed that ENA78 / CXCL5 and SDF-1 / CXCL12 can be used as diagnostic markers for gastric cancer, which have improved accuracy compared to conventional gastric cancer diagnostic markers.

2.2. Correlation between Serum Concentrations of ENA78 / CXCL5, SDF-1 / CXCL12, and CEA and Clinicopathological Parameters

For statistical analysis of the measured values, IBM SPSS Statistics 20.0 was used. All measured values were expressed as a mean containing 25-75% standard deviation, and each group mean value was determined by one-way ANOVA test method using multiple comparisons of the post-hoc Bonferroni method Respectively. In order to confirm the correlation between serum concentration measured by the method of Example 2.1 and clinicopathological parameters, Pearson's correlation coefficient (g _p ) and Spearman's correlation coefficient ., was confirmed by using the coefficient, g _s) in the position of gastric cancer (tumor location) lower the antrum (antrum) - refers to the upper leg (angle) and, middle is the lower part of the above-refers to the middle part, upper means upper part - cardia above. The tumor size was classified as cancer less than 3 cm, cancer between 3 cm and 5 cm, and cancer greater than 5 cm. The TMN stage was classified according to the TMN classification of 7th International Union Against Cancer. The overall stage is divided into stages I, II, III, and IV. Stage I is the early stage of gastric cancer, and IV is the stage of late gastric cancer. Table 4 shows the results of confirming the serum concentration according to the T-stage, Table 3 shows the results of confirming the serum concentrations according to the N-stage, Table 4 shows the results of confirming the serum concentrations according to the M-stage, The results are shown in Table 6. The results are shown in Table 7.

Groups T1a
(34) T1b
(16) T2
(17) T3
(15) T4
(18) p -value ^† ENA78 (ng / mL) 1974.3 ± 1083.5 2356.9 + 1368.3 2824.3 ± 2012.0 2912.2 ± 1291.5 3002.6 + - 1451.7 0.063 SDF-1 (ng / mL) 2627.0 占 1626.7 2716.7 ± 1674.4 2645.3 ± 1883.7 3710.0 + - 2490.4 3558.2 ± 2329.1 0.246 CEA (ng / mL) 2.8 ± 3.1 1.9 ± 2.0 3.7 ± 5.6 4.0 ± 6.9 5.1 ± 5.9 0.346

Groups
(n) N0
(70) N1
(8) N2
(5) N3
(17) p -value ^† ENA78 (ng / mL) 2278.6 + 1389.4 2531.1 + 1484.7 2856.6 ± 1630.6 3326.3 占 1467.1 0.057 SDF-1 (ng / mL) 2592.9 ± 1624.2 2761.3 ± 1764.6 3786.9 ± 1798.1 4407.3 ± 2749.3 0.005 CEA (ng / mL) 3.1 ± 4.6 4.1 ± 6.1 1.5 ± 0.4 4.8 ± 5.2 0.467

Groups
(n) M0
(84) M1
(16) p -value ^‡ ENA78 (ng / mL) 2291.3 ± 1358.1 3632.0 占 1463.8 0.001 SDF-1 (ng / mL) 2548.8 + - 1641.1 5209.5 ± 2125.8 <0.001 CEA (ng / mL) 2.4 ± 2.4 9.0 ± 8.9 <0.001

Groups Ia
(50) Ib
(9) II
(13) III
(11) IV
(17) p -value ^† ENA78 (ng / mL) 2149.7 ± 1085.0 2904.0 ± 2103.3 2410.7 ± 1755.0 2108.4 + - 1231.8 3672.4 ± 1401.1 <0.001 SDF-1 (ng / mL) 2437.1 ± 1700.0 2768.8 ± 2075.7 2735.7 ± 1204.3 2915.3 ± 1515.8 4884.9 ± 2402.6 <0.001 CEA (ng / mL) 2.4 ± 2.5 2.1 ± 1.1 2.9 ± 3.2 1.8 ± 0.8 8.5 ± 8.8 0.018

Clinicopathological characteristics ENA78 / CXCL5 SDF-1 / CXCL12 CEA g _s ( p- value) g _s ( p- value) g _s ( p- value) Sex (Male: Female) -0.169 (0.098) -0.008 (0.938) -0.010 (0.921) Age (years) * 0.058 (0.575) 0.187 (0.066) 0.072 (0.486) H. pylori  infection (- / +) -0.014 (0.893) 0.041 (0.687) -0.133 (0.186) Histology (Well: Mod: Poorly: Signet-ring) -0.011 (0.922) 0.159 (0.269) 0.147 (0.309) Tumor location (Lower: Middle: Upper) ^† 0.117 (0.420) 0.222 (0.124) 0.064 (0.660) Size of tumor (<3 cm, 3-5 cm and> 5 cm) ^‡ 0.186 (0.238) 0.044 (0.783) 0.200 (0.204) T-stage (T1a: T1b: T2: T3: T4) ^§ 0.280 (0.049) 0.232 (0.105) 0.106 (0.462) N-stage (N0: N1: N2: N3) ^§ 0.271 (0.057) ^¶ 0.304 (0.032) 0.093 (0.520) Distant Metastasis (M0: M1) ^§ 0.461 (0.001) 0.532 (< 0.001) 0.303 (0.032) Overall stage (I: II: III: IV) 0.367 (0.009) 0.334 (0.018) 0.175 (0.224)

As shown in Tables 3 to 7, the concentrations of ENA78 / CXCL5, SDF-1 / CXCL12 and CEA in the serum were not related to sex, age, Helicobacter pylori infection, or the like. Serum concentrations of ENA78 / CXCL5 were found to be closely related to T stage showing the extent of tumor invasion around and M stage showing distant metastasis. Serum concentrations of SDF-1 / CXCL12 were correlated with N-stage lymph node metastasis, M stage with distant metastasis, and overall staging of gastric cancer. Using the ENA78 / CXCL5 as a diagnostic marker for gastric cancer, it is possible to identify the stage of the TM stage and the stomach cancer, so that the stage of cancer progression can be confirmed. Through the progress of the cancer, , Prediction of therapeutic effect, determination of treatment method, and so on. In addition, when ENA78 / CXCL5 and SDF-1 / CXCL12 are used together, it is possible to confirm the stage of gastric cancer (stage progression) including stage of TMN stage and stage of gastric cancer because additional stage confirmation of N stage is possible. In contrast, CEA, which is a conventional gastric cancer marker, can be used as a diagnostic marker to detect the occurrence of stomach cancer. However, CEA can be used as a diagnostic marker to detect the occurrence of gastric cancer. It can be confirmed that it is not suitable for the following.

2.3. Possibility of ENA78 / CXCL5 and SDF-1 / CXCL12 as diagnostic markers

In order to confirm that ENA78 / CXCL5 and SDF-1 / CXCL12 can be used as diagnostic markers in comparison with CEA, which is a conventional gastric cancer diagnostic marker, the values calculated in Example 2.1 are used to show the accuracy of diagnostic tests with IBM SPSS Statistics 20.0 Receiver operating characteristic (ROC) curves (Pattern Recognition Letters (2006) 27 (8): 861-874) indicating sensitivity and specificity are obtained from the recipient operating characteristic curves Area under the curves (AUC) were calculated and the diagnostic accuracy of each marker was compared. The cut-off point represents the probability of classifying a patient in a binary logistic regression that only determines the presence of gastric cancer. In addition, logistic regression was performed to measure the diagnostic accuracy of single marker and multiple marker. The results are shown in Fig. 1 and Table 8.

As shown in Fig. 1, the AUC value of ENA78 / CXCL5 was 0.90 (95% CI, 0.84-0.96), SDF-1 / CXCL12 was 0.75 (95% CI, 0.65-0.85) Was found to be 0.52 (95% CI, 0.41-0.64). From these results, it was confirmed that ENA78 / CXCL5, which has the highest sensitivity and specificity, is the marker with the highest diagnostic accuracy, and the diagnosis accuracy is improved even compared with the CEA, which is a conventional marker for gastric cancer diagnosis It is confirmed that the marker is a diagnostic marker that can be used.

Marker panel ^* Cut-off point ^† Sensitivity ^‡ Specificity ENA78 / CXCL5 0.45 78.0% 80.0% SDF-1 / CXCL12 0.40 70.0% 64.0% CEA 0.46 54.0% 40.0% ENA78 / CXCL5 + SDF-1 / CXCL12 0.58 78.0% 90.0% ENA78 / CXCL5 + CEA 0.45 78.0% 80.0% SDF-1 / CXCL12 + CEA 0.40 70.0% 64.0% ENA78 / CXCL5 + SDF-1 / CXCL12 + CEA 0.58 78.0% 90.0%

As shown in Table 8, ENA78 / CXCL5 showed sensitivity and specificity of 78.0% and 80.0%, respectively. ENA78 / CXCL5 and SDF-1 / CXCL5 , It was confirmed that the specificity can be increased up to 90.0%. In contrast, ENA78 / CXCL5 and SDF-1 / CXCL12 improved diagnostic accuracy compared to conventional gastric cancer markers, and the sensitivity and specificity of CEA were not significantly different It is confirmed that the accuracy of diagnosis can be further improved by using multiple markers.

Example 3: Independent validation dataset

3.1. Identification of Serum Levels of ENA78 / CXCL5, SDF-1 / CXCL12 and CEA by Stage of Gastric Cancer

In order to increase the accuracy of the results from the initial training data, independent verification data were performed. Serum concentrations of ENA78 / CXCL5, SDF-1 / CXCL12 and CEA were measured in the same manner as in Example 2.1. The results are shown in Table 9.

Groups
(n) Normal
(70) High-risk
(30) EGC
(50) AGC
(50) p -value ^† Serum ENA78 (ng / ml) 1125.5 + - 521.2 * 1092.3 ± 609.4 2133.6 + - 1146.9 2878.0 占 1635.5 <0.001 Serum SDF-1 (ng / ml) 1928.0 占 094.6 2169.6 + - 1144.6 2587.8 ± 1595.7 3361.3 ± 2244.0 <0.001 Serum CEA (ng / ml) 2.0 ± 1.5 3.3 ± 2.6 2.4 ± 2.7 5.1 ± 7.4 0.005 Groups
(n) Non-cancer groups
(100) Cancer groups
(100) p -value ^‡ Serum ENA78 (ng / ml) 1115.5 + 546.2 * 2505.8 ± 1454.3 <0.001 Serum SDF-1 (ng / ml) 2000.4 ± 1109.6 2974.5 ± 1975.8 <0.001 Serum CEA (ng / ml) 2.4 ± 1.9 3.8 ± 5.7 0.025

As shown in Table 9, the ENA78 / CXCL5 and SDF-1 / CXCL12 were both increased in gastric cancer patients more than twice as much as the initial training data. In addition, ENA78 / CXCL5 and SDF-1 / CXCL12 showed an increase in serum concentration as the stage of gastric cancer progressed. Especially, ENA78 / CXCL5 showed no increase in the high-risk group, And it was confirmed that these markers can be used for early diagnosis of gastric cancer. On the other hand, CEA, which is a marker for gastric cancer diagnosis, was found to increase in serum concentration only in advanced gastric cancer. Therefore, it was confirmed that ENA78 / CXCL5 or SDF-1 / CXCL12 can improve the accuracy of diagnosis compared with existing markers in early stomach cancer diagnosis.

3.2. Correlation between Serum Concentrations of ENA78 / CXCL5, SDF-1 / CXCL12, and CEA and Clinicopathological Parameters

The Pearson's correlation coefficient, g _p , and Spearman's correlation coefficient were used to determine the correlation between serum concentrations of ENA78 / CXCL5 or SDF-1 / CXCL12 and clinicopathologic factors ., was used as g _s) in the position of gastric cancer (tumor location) lower the antrum (antrum) - refers to the upper leg (angle) and, middle is the lower part of the above-refers to the middle part, upper is above Upper part - means cardia. The tumor size was classified as cancer less than 3 cm, cancer between 3 cm and 5 cm, and cancer greater than 5 cm. The TMN stage was classified according to the TMN classification of 7th International Union Against Cancer. The overall stage is divided into stages I, II, III, and IV. Stage I is the early stage of gastric cancer, and IV is the stage of late gastric cancer. The results are shown in Table 10.

Clinicopathological characteristics ENA78 / CXCL5 SDF-1 / CXCL12 CEA g _s ( p- value) g _s ( p- value) g _s ( p- value) Sex (Male: Female) -0.048 (0.501) -0.020 (0.776) -0.046 (0.522) Age (years) * 0.058 (0.419) 0.088 (0.218) 0.087 (0.224) H. pylori  infection (- / +) 0.045 (0.523) 0.049 (0.487) -0.014 (0.848) Histology (Well: Mod: Poorly: Signet-ring) 0.016 (0.876) 0.066 (0.518) 0.101 (0.156) Tumor location (Lower: Middle: Upper) ^† 0.015 (0.883) 0.090 (0.374) 0.157 (0.120) Size of tumor (<3 cm, 3-5 cm and> 5 cm) ^‡ 0.051 (0.647) -0.061 (0.584) 0.293 (0.007) T-stage (T1a: T1b: T2: T3: T4) ^§ 0.231 (0.021) 0.191 (0.058) ^¶ 0.197 (0.093) ^¶ N-stage (N0: N1: N2: N3) ^§ 0.181 (0.073) ^¶ 0.220 (0.029) 0.107 (0.292) Distant Metastasis (M0: M1) ^§ 0.357 (< 0.001) 0.425 (< 0.001) 0.264 (0.008) Overall stage (I: II: III: IV) 0.315 (0.001) 0.248 (0.013) 0.188 (0.062) ^¶

As shown in Table 10, it is confirmed that the results are similar to the initial training data. Serum concentrations of ENA78 / CXCL5 were highly correlated with T-stage, M stage and overall stage of cancer. Serum concentrations of SDF-1 / CXCL12 Concentrations were correlated with lymph node metastasis (N-stage), distant metastasis (M stage), and cancer stage (overall stage). Unlike CEA, ENA78 / CXCL5 and SDF-1 / CXCL12 can be used to identify cancer stages (stages of progression).

3.3. Possibility of ENA78 / CXCL5 and SDF-1 / CXCL12 as diagnostic markers

The ROC curve and AUC were calculated in the same manner as in Example 2.3 to confirm the possibility of ENA78 / CXCL5 and SDF-1 / CXCL12 as gastric cancer diagnostic markers. The results are shown in Fig.

As shown in Figure 2, the AUC of ENA78 / CXCL5 was 0.82 (95% CI, 0.76-0.87), SDF-1 / CXCL12 0.63 (95% CI, 0.55-0.71), CEA 0.52 -0.60). As a result, it was confirmed that ENA78 / CXCL5 and SDF-1 / CXCL12 improved diagnostic accuracy compared with CEA, which is used as a marker for gastric cancer diagnosis.

In addition, logistic regression was performed to measure the diagnostic accuracy of single marker and complex marker. The results are shown in Table 11.

Marker panel ^* Cut-off point ^† Sensitivity ^‡ Specificity ENA78 / CXCL5 0.43 71.0% 75.0% SDF-1 / CXCL12 0.46 56.0% 59.0% CEA 0.47 54.0% 46.0% ENA78 / CXCL5 + SDF-1 / CXCL12 0.44 71.0% 76.0% ENA78 / CXCL5 + CEA 0.46 71.0% 75.0% SDF-1 / CXCL12 + CEA 0.46 56.0% 64.0% ENA78 / CXCL5 + SDF-1 / CXCL12 + CEA 0.47 71.0% 76.0%

As shown in Table 11, the sensitivity and specificity of ENA78 / CXCL5 were all higher than 70%, indicating that ENA78 / CXCL5 and SDF-1 / CXCL12 were the highest among single markers When used, it was confirmed that the specificity was 76%. These results indicate that ENA78 / CXCL5 can be used as a single diagnostic marker and that diagnostic accuracy can be improved when ENA78 / CXCL5 and SDF-1 / CXCL12 are used together as a complex diagnostic marker.

3.4. Identification of potential as marker for distant metastasis of ENA78 / CXCL5, SDF-1 / CXCL12, and CEA

Through Example 3.2, it was confirmed that ENA78 / CXCL5, SDF-1 / CXCL12 and CEA were closely related to distant metastasis (M stage). Therefore, the ROC curve and AUC were calculated in the same manner as in Example 2.3 to confirm whether or not each marker can be used as a marker for predicting distant metastasis. In addition, logistic regression was performed to measure the diagnostic accuracy of single marker and multiple marker. The results are shown in FIG. 3 and Table 12.

As shown in Figure 3, the AUC of ENA78 / CXCL5 was 0.78 (95% CI, 0.68-0.88), SDF-1 / CXCL12 was 0.83 (95% CI, 0.73-0.94), CEA was 0.71 -0.87). From these results, it was confirmed that ENA78 / CXCL5, SDF-1 / CXCL12 and CEA can be used to diagnose distant metastasis of gastric cancer. In particular, SDF-1 / CXCL12 showed the highest diagnostic accuracy.

Marker panel ^* Cut-off point ^† Sensitivity ^‡ Specificity ENA78 / CXCL5 0.13 75.0% 61.4% SDF-1 / CXCL12 0.15 75.0% 75.9% CEA 0.47 68.8% 57.8% ENA78 / CXCL5 + SDF-1 / CXCL12 0.13 75.0% 78.3% ENA78 / CXCL5 + CEA 0.13 75.0% 77.0% SDF-1 / CXCL12 + CEA 0.22 75.0% 83.1% ENA78 / CXCL5 + SDF-1 / CXCL12 + CEA 0.20 75.0% 92.8%

In addition, as shown in Table 12, it was confirmed that ENA78 / CXCL5, SDF-1 / CXCL12 and CEA could be used as a single marker to diagnose distant metastasis of gastric cancer, and when used as a complex marker, Thus, it is confirmed that the diagnostic accuracy can be further increased. Based on the above results, it was confirmed that information on the distant metastasis can be obtained by using ENA78 / CXCL5, SDF-1 / CXCL12 and / or CEA as diagnostic markers of gastric cancer.

These results indicate that ENA78 / CXCL5 and SDF-1 / CXCL12 can be used to diagnose early gastric cancer more accurately than CEA, which is a single marker for gastric cancer diagnosis marker. ENA78 / CXCL5 and SDF-1 / It was confirmed that the accuracy of diagnosis of stomach cancer can be further improved by using it as a combined marker. In addition, ENA78 / CXCL5 or SDF-1 / CXCL12 can be used as a marker for gastric cancer diagnosis, and it can acquire various information related to penetration depth, metastasis, stage of cancer, cancer size, I can confirm that I can. Furthermore, since ENA78 / CXCL5, SDF-1 / CXCL12 and CEA all have a close relationship with distant metastasis, it was confirmed that distant metastasis can be diagnosed by using the markers as single or multiple markers.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

Claims (14)

The level of ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) was measured from biological samples isolated from patients and compared with the level of normal control sample, How to provide. The method according to claim 1,
The stage of the gastric cancer is composed of cancer size, T-stage of cancer, N-stage, distant metastasis, and an overall cancer stage Gt; information &lt; / RTI &gt; The method according to claim 1,
Wherein said biological sample is blood, plasma or serum. The level of SDF-1 / CXCL12 (stromal cell-derived factor-1 / chemokine (CXC motif) ligand 12) was measured from biological samples isolated from patients and compared with the level of normal control samples, / RTI &gt; 5. The method of claim 4,
The stage of the gastric cancer is composed of cancer size, T-stage of cancer, N-stage, distant metastasis, and an overall cancer stage Gt; information &lt; / RTI &gt; 5. The method of claim 4,
Wherein said biological sample is blood, plasma or serum. ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) and SDF-1 / CXCL12 (stromal cell-derived factor-1 / chemokine ) And comparing the level of the normal control sample with that of the normal control sample to provide information about the diagnosis of gastric cancer or the stage of gastric cancer. 8. The method of claim 7,
The stage of the gastric cancer is composed of cancer size, T-stage of cancer, N-stage, distant metastasis, and an overall cancer stage Gt; information &lt; / RTI &gt; 8. The method of claim 7,
Wherein said biological sample is blood, plasma or serum. From the biological samples isolated from patients, ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5), SDF-1 / CXCL12 (stromal cell- 12), and carcinoembryonic antigen (CEA), and comparing the level of one or more markers selected from the group consisting of CEA (carcinoembryonic antigen) with the level of a normal control sample to provide information about distant metastasis of gastric cancer. 11. The method of claim 10,
Wherein said biological sample is blood, plasma or serum. A diagnostic kit for stomach cancer or a stomach cancer staging kit comprising an antibody or an aptamer specifically binding to ENA78 / CXCL5 (epithelial-derived neutrophil-activating peptide 78 / chemokine (CXC motif) ligand 5) . 13. The method of claim 12,
The kit further comprises an antibody or an aptamer that specifically binds SDF-1 / CXCL12 (stromal cell-derived factor-1 / chemokine (CXC motif) ligand 12). 14. The method of claim 13,
Wherein the kit further comprises an antibody or an aptamer that specifically binds to carcinoembryonic antigen (CEA).

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