阅读说明：本技术 用于预测肺腺癌预后的标志物及其应用 (Marker for predicting lung adenocarcinoma prognosis and application thereof ) 是由 范涛 耿庆 潘世泽 杨硕 郝博 李东航 张霖 于 2021-08-17 设计创作，主要内容包括：本发明涉及一种用于预测肺腺癌预后的标志物及其应用,该标志物为IL7R、IL5RA、IL20RB、IL11和IL22RA1的组合,将该标志物的表达水平代入肺腺癌患者预后评分模型中,计算出风险评分,根据评分高低预测肺腺癌患者的总体生存,并为筛选出适合特定免疫检查点抑制剂治疗的肺腺癌患者提供依据。本发明需要检测的指标较少,仅需要检测患者样本内的IL7R、IL5RA、IL20RB、IL11和IL22RA1表达水平,即可预测肺腺癌预后,指导临床医生对高风险患者提供积极治疗；并且该发明指导临床医生对低风险患者采用CTLA4或TIM3免疫检查点抑制剂治疗提供依据。(The invention relates to a marker for predicting lung adenocarcinoma prognosis and application thereof, wherein the marker is a combination of IL7R, IL5RA, IL20RB, IL11 and IL22RA1, the expression level of the marker is substituted into a lung adenocarcinoma patient prognosis score model, a risk score is calculated, the overall survival of a lung adenocarcinoma patient is predicted according to the score, and a basis is provided for screening lung adenocarcinoma patients suitable for being treated by a specific immune checkpoint inhibitor. The invention has less indexes to be detected, and can predict the lung adenocarcinoma prognosis by detecting the expression levels of IL7R, IL5RA, IL20RB, IL11 and IL22RA1 in a patient sample, thereby guiding a clinician to provide positive treatment for a high-risk patient; and the invention guides clinicians to provide basis for low risk patients to be treated with CTLA4 or TIM3 immune checkpoint inhibitors.)

1. A marker, characterized by: the markers are a combination of IL7R, IL5RA, IL20RB, IL11 and IL22RA1 for predicting lung adenocarcinoma patient prognosis or for determining whether a lung adenocarcinoma patient is suitable for immunotherapeutic methods targeting CTLA4 or TIM 3.

2. A detection reagent, characterized in that: for detecting the marker of claim 1.

3. A kit, characterized in that: comprising the detection reagent of claim 2, for predicting lung adenocarcinoma prognosis or for predicting whether a lung adenocarcinoma patient is eligible for an immunotherapy targeting CTLA4 or TIM 3.

4. The kit of claim 3, wherein: the kit comprises a lung adenocarcinoma patient prognosis scoring model, wherein the lung adenocarcinoma patient prognosis scoring model comprises a mathematical formula: risk score-0.09948 × IL7R expression level + -0.51191 × IL5RA expression level +0.09591 × IL20RB expression level +0.28446 × IL11 expression level +0.2596 × IL22RA1 expression level.

5. The kit of claim 4, wherein: a method of predicting whether a lung adenocarcinoma patient is eligible for an immunotherapeutic method targeting CTLA4 or TIM3 comprising the steps of:

(1) detecting the expression level of IL7R, IL5RA, IL20RB, IL11 and IL22RA1 in a sample of a patient with lung adenocarcinoma;

(2) substituting the detection result of the step (1) into the mathematical formula to calculate a risk score;

(3) when the risk score is larger than or equal to the median value, the lung adenocarcinoma patient is judged to have poorer prognosis and is not suitable for the anti-CTLA 4 or TIM3 immune checkpoint inhibitor treatment; when the risk score is < median, the patient is judged to be better in prognosis, suitable for immune checkpoint inhibitor treatment against CTLA4 or TIM 3.

6. The kit of claim 5, wherein: the median value is 0.968578517.

7. A system for predicting prognosis of lung adenocarcinoma, comprising:

an obtaining module for obtaining the expression level of the marker of claim 1;

the prediction module is used for predicting the risk score of the lung adenocarcinoma patient according to the expression level obtained by the acquisition module and outputting the risk score; the prediction module comprises a lung adenocarcinoma patient prognosis scoring model, and the lung adenocarcinoma patient prognosis scoring model comprises a mathematical formula: risk score-0.09948 × IL7R expression level + -0.51191 × IL5RA expression level +0.09591 × IL20RB expression level +0.28446 × IL11 expression level +0.2596 × IL22RA1 expression level;

the acquisition module and the prediction module are connected in a wireless and/or wired mode.

8. The system for predicting prognosis of lung adenocarcinoma according to claim 7, characterized in that: the system is also useful for predicting whether a lung adenocarcinoma patient is eligible for an immunotherapeutic approach targeting CTLA4 or TIM 3.

9. Use of the marker of claim 1 for the preparation of a detection reagent of claim 2 or a kit of claim 3.

10. Use of a marker according to claim 1 for the preparation of a system according to claim 7 for predicting prognosis of lung adenocarcinoma.

Technical Field

The invention relates to the field of clinical application, in particular to a marker for predicting lung adenocarcinoma prognosis and application thereof.

Background

Currently, lung cancer remains the major type of malignancy worldwide and is a significant cause of cancer-related death. Lung adenocarcinoma (LUAD), the major molecular subtype of lung cancer, accounts for over 40% of lung cancers. Despite great advances in lung cancer treatment strategies, including molecularly targeted drugs and immune checkpoint inhibitors, the 5-year survival rate of lung cancer remains low. Therefore, there is a need to find a method that can specifically predict patient survival and can be used as a marker of tumor treatment response to give the most appropriate personalized treatment for different lung adenocarcinoma patients.

In recent years, with the development of multiomics, many studies using different expression profiles and bioinformatics have provided various prognostic evaluation methods for LUAD patients. However, the parameters used in these studies were derived from genome-wide and transcriptomic data, and do not take into account the intrinsic influence of various molecular families in tumor biological processes, which may lead to computational bias. Furthermore, these methods are simple mathematical models and may not reflect the intrinsic characteristics of the tumor itself.

With the successful application of various Immune Checkpoint Inhibitors (ICIs) such as programmed death receptor 1(PD1), programmed death receptor ligand 1(PD-L1), etc., the treatment of lung cancer has achieved many innovations. A number of large sample size cohort studies have established the role of PD1/PD-L1 in immunosuppression and its ability to serve as a prognostic biomarker for tumor progression or a biomarker for predicting immune response. However, still more than half of patients do not respond to anti-PD 1/PD-L1 immunotherapy, suggesting that more molecular signaling pathways are involved in the regulation of tumor immunity and that these molecules have relevance to immune checkpoint signaling pathways such as PD 1/PD-L1.

Disclosure of Invention

In order to accurately stratify patients with lung adenocarcinoma, take effective treatment measures for specific patients and solve the problem that more than half of patients with lung adenocarcinoma do not respond to anti-PD 1/PD-L1 immunotherapy, the invention provides a marker for accurately predicting the prognosis of patients with lung adenocarcinoma and provides a basis for selecting proper immune checkpoint inhibitor treatment for patients, and the marker can be used for accurately predicting whether patients with lung adenocarcinoma are suitable for cytotoxic T lymphocyte-associated antigen 4(CTLA4) or tenascin 3(TIM3) inhibitor treatment.

The technical scheme provided by the invention is as follows:

in a first aspect, a marker is provided that is a combination of interleukin-7 receptor (IL7R), interleukin-5 receptor alpha (IL5RA), interleukin-20 receptor beta chain antibody (IL20RB), human interleukin 11(IL11), and interleukin-22 receptor a1(IL22RA1) for predicting lung adenocarcinoma patient prognosis or determining whether a lung adenocarcinoma patient is eligible for an immunotherapeutic approach targeting CTLA4 or TIM 3.

In a second aspect, a detection reagent for detecting the above markers is provided, and a lung adenocarcinoma population suitable for anti-CTLA 4 or TIM3 immune checkpoint inhibitor treatment is screened out, so as to predict whether a lung adenocarcinoma patient is suitable for an immunotherapy targeting CTLA4 or TIM 3.

In a third aspect, a kit comprising the above detection reagent is provided for predicting prognosis of a lung adenocarcinoma patient or for predicting whether a lung adenocarcinoma patient is suitable for an immunotherapy targeting CTLA4 or TIM 3.

Specifically, the kit comprises a lung adenocarcinoma patient prognosis score model, which comprises a mathematical formula: risk score-0.09948 × IL7R expression level + -0.51191 × IL5RA expression level +0.09591 × IL20RB expression level +0.28446 × IL11 expression level +0.2596 × IL22RA1 expression level.

The prediction method of the kit comprises the following steps:

(1) detecting the expression level of IL7R, IL5RA, IL20RB, IL11 and IL22RA1 in a sample of a patient with lung adenocarcinoma;

(2) substituting the detection result of the step (1) into the mathematical formula to calculate a risk score;

(3) when the risk score is larger than or equal to the median value, the lung adenocarcinoma patient is judged to have poorer prognosis and is not suitable for the anti-CTLA 4 or TIM3 immune checkpoint inhibitor treatment; when the risk score is < median, the patient is judged to be better in prognosis, suitable for immune checkpoint inhibitor treatment against CTLA4 or TIM 3.

Specifically, the median value is 0.968578517.

Specifically, the sample of the lung adenocarcinoma patient is a tissue or a body fluid, and the body fluid is preferably blood.

In a fourth aspect, there is provided a system for predicting prognosis of lung adenocarcinoma, the system comprising:

an obtaining module for obtaining expression levels of IL7R, IL5RA, IL20RB, IL11 and IL22RA 1;

the prediction module is used for predicting the risk score of the lung adenocarcinoma patient according to the expression level obtained by the acquisition module and outputting the risk score; the prediction module comprises a lung adenocarcinoma patient prognosis score model which comprises a mathematical formula: risk score-0.09948 × IL7R expression level + -0.51191 × IL5RA expression level +0.09591 × IL20RB expression level +0.28446 × IL11 expression level +0.2596 × IL22RA1 expression level;

the acquisition module is connected with the prediction module in a wireless and/or wired mode.

The system can accurately predict the prognosis of the patient with the lung adenocarcinoma, and also provides a basis for selecting a proper immune checkpoint inhibitor for treatment of the patient. The system is also useful for predicting whether a lung adenocarcinoma patient is eligible for an immunotherapeutic approach targeting CTLA4 or TIM 3.

In a fifth aspect, there is provided the use of the above marker in the preparation of the above detection reagent or the above kit.

In a sixth aspect, there is provided the use of the above marker in the preparation of the above system for predicting prognosis of lung adenocarcinoma.

The invention has the following beneficial effects:

1. the marker provided by the invention is a combination of IL7R, IL5RA, IL20RB, IL11 and IL22RA1, is an interleukin and a receptor family thereof, screens out independent risk factors of lung cancer for the first time based on molecular expression of the interleukin and the receptor family thereof, and constructs a prognosis score model of a lung adenocarcinoma patient with tumor prognosis related to the interleukin and the receptor for the first time according to a screening result, only the expression levels of the 5 markers in a tumor tissue of the patient need to be detected and are brought into the prognosis score model of the lung adenocarcinoma patient, and the prognosis of the lung adenocarcinoma patient and whether the lung adenocarcinoma patient is suitable for an immunotherapy method targeting CTLA4 or TIM3 can be predicted according to risk scores.

2. The means for predicting the prognosis of the lung adenocarcinoma patient or predicting whether the lung adenocarcinoma patient is suitable for the immunotherapy method targeting CTLA4 or TIM3 based on the marker combination and the lung adenocarcinoma patient prognosis score model has strong capability of predicting the patient prognosis compared with the prior art based on the models with TIDE, TMB, IFNG, clarification, Merck18, CD8 and Dysfunction as markers.

Drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings needed to be used in the description of the embodiments will be briefly introduced below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings based on these drawings without creative efforts.

Figure 1 shows the relationship between the life span and risk score for 1312 independent cohorts of lung adenocarcinoma patients; fig. 1a represents the TCGA database, fig. 1b represents the GSE31210 queue, fig. 1c represents the GSE30219 queue, fig. 1d represents the GSE72094 queue, and fig. 1e represents the GSE13213 queue.

FIG. 2 illustrates the ability of the model established in example 1 to predict prognosis in patients with lung adenocarcinoma compared to prior art models; FIG. 2a shows a 1-year survival rate versus risk score for the model established in example 1; FIG. 2b represents the time dependent AUC values (area under ROC curve) of the model established in example 1; FIG. 2c is a graph of 1 year survival versus risk score for a representative prior art model; FIG. 2d represents the time-dependent AUC values of a prior art model; the arrows in fig. 2a and 2c represent ROC curves of the model, and a larger area below the ROC curve indicates better prediction performance; the arrows in fig. 2b and 2d represent the AUC values of the model, and the larger the AUC value, the better the prediction performance.

Figure 3 shows the correlation analysis of model risk scores with the expression levels of the immune checkpoints CTLA4 and TIM 3; figure 3a shows the relationship between model risk score and CTLA4 expression levels; figure 3b shows a comparison of CTLA4 expression levels in patients in the low risk group and high risk group; figure 3c shows the relationship between model risk score and TIM3 expression level; figure 3d shows a comparison of TIM3 expression levels in patients in the low risk group and the high risk group.

Detailed Description

In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be obtained by a person skilled in the art without any inventive step based on the embodiments of the present invention, are within the scope of the present invention.

The following examples screen independent interleukins and their receptor molecules as marker combinations affecting the prognosis of lung adenocarcinoma using a Cox risk ratio lung adenocarcinoma patient prognosis scoring model and a minimum absolute contraction selection algorithm based on lung adenocarcinoma transcriptome data. And constructing a lung adenocarcinoma patient prognosis score model according to the action proportion of each marker in lung adenocarcinoma prognosis. Through verification of a plurality of independent queues, the model is proved to have strong lung adenocarcinoma prognosis prediction value, and compared with the biomarker which is proved to be applicable to lung cancer immunotherapy reaction at present, the lung adenocarcinoma patient prognosis scoring model established by the invention shows higher prediction capability.

Example 1: screening of marker and establishment of model

(1) Downloading lung adenocarcinoma transcriptome data with prognosis information from TCGA database, extracting 87 interleukins and receptor molecule families thereof, and screening out 27 differentially expressed interleukin molecule family members in tumor tissues and tissues beside the cancer by utilizing R software ('RVersion 3.6.1') and a 'limma' program package; 7 dangerous interleukin molecules were identified using Cox one-way regression analysis.

(2) And (2) further performing LASSO regression analysis according to the patient data in the step (1) and the expression condition of the screened interleukin molecules, wherein the shrinkage variables are 5 (IL7R, IL5RA, IL20RB, IL11 and IL22RA1), and constructing a lung adenocarcinoma patient prognosis score model according to the five variables. The specific method adopts R language ("RVersion 3.6.1"), a "survival" program package and a "glmnet" program package.

(3) And constructing a lung adenocarcinoma patient prognosis score model. The risk score (risk score) of a patient is calculated as follows:

in the formula, n represents the number of independent risk factors, β represents a risk coefficient, x represents a value corresponding to each independent risk factor, β k represents a risk coefficient corresponding to the kth risk factor, and xk represents a value corresponding to the kth risk factor.

In this embodiment, we first calculate the proportion of each of the 5 risk factors (IL7R, IL5RA, IL20RB, IL11, IL22RA1) of the LUAD patient in the patient's disease progression (risk factor β) using the R language ("RVersion 3.6.1") and the "survival" package, and multiply each of the factors by the corresponding value of the risk factor, and the sum of the results is the risk score (risk score) of the patient; the resulting risk score calculation formula is as follows:

risk score＝-0.09948×IL7R+-0.51191×IL5RA+0.09591×IL20RB+0.28446×IL11+0.2596×IL22RA1

(1) patients were then ranked according to risk score and then divided into 2 groups according to median or joden index (best cut-off for risk score). In the training set for building the model, the included Low risk group (Low riskgroup) with the median value (0.968578517) or less and the included High risk group (High-risk group) with the median value (0.968578517) or more are included.

Example 2: reliability verification of lung adenocarcinoma patient prognosis score model

To validate the reliability of the prognostic scoring model for patients with lung adenocarcinoma established in example 1, we performed a systematic analysis of the expression levels and clinical features of the IL family in LUAD.

(2) The expression levels of IL7R, IL5RA, IL20RB, IL11 and IL22RA1 of 1312 lung adenocarcinoma samples in 5 independent cohorts (TCGA database, GSE31210 cohort, GSE30219 cohort, GSE72094 cohort and GSE13213 cohort) are substituted into the lung adenocarcinoma patient prognosis score model established in example 1 to obtain risk scores, and the survival years of the 1312 lung adenocarcinoma patients in the 5 independent cohorts and the relationship between the low risk group and the high risk group are analyzed, as shown in fig. 1, the analysis results of the 5 independent cohorts all show that the lung adenocarcinoma patients in the high risk group have significantly poor prognosis, and the patients in the low risk group show significant survival advantage, which shows that the lung adenocarcinoma patient prognosis model established in example 1 has a strong capability of predicting lung adenocarcinoma.

Example 3: comparison of the Lung adenocarcinoma patient prognosis score model with the prior art

At present, there are many markers of the treatment response of immune checkpoint inhibitors, and in order to show the superiority of the lung adenocarcinoma patient prognosis score model established in example 1, which uses PD1, PDL1, CTLA4, LAG3 and TIM3 as markers, we compared the model with clinically recognized models for predicting lung adenocarcinoma prognosis, which use TIDE, TMB, IFNG, Exclusion, Merck18, CD8 and Dysfunction as markers.

As shown in fig. 2a and fig. 2c, the lung adenocarcinoma patient prognosis score model established in example 1 has better prediction performance, the area under the ROC curve can directly reflect the prediction performance, and the larger the area is, the better the prediction performance is.

As shown in fig. 2b and fig. 2d, the time-dependent AUC value of the predicted efficacy of the prognostic scoring model for lung adenocarcinoma patients established in example 1 is larger (area under the curve), and the prognostic power of the lung adenocarcinoma patients is better compared with the existing model at all times in the model established in example 1.

Example 4: correlation analysis of risk score with expression levels of CTLA4 and TIM3

The expression level of the immune checkpoint is critical for the anti-tumor effect of the immune checkpoint inhibitor, so we compared the Risk score (Risk score) of this model with the expression level of the immune checkpoint and showed that fig. 3a and 3c performed a correlation analysis of the Risk score with the expression levels of CTLA4 and TIM3, and showed that the Risk score was negatively correlated with the expression levels of CTLA4 and TIM3, and fig. 3b and 3d compared the expression levels of CTLA4, TIM3 in high-Risk and low-Risk patients, and showed that CTLA4 and TIM3 expression was significantly increased in low-score patients, which indicated that the low-score patients were more suitable for the treatment with CTLA4 and 3 inhibitor. Through the correlation analysis of the risk score and the expression of the current immune checkpoint, IL7R, IL5RA, IL20RB, IL11 and IL22RA1 are indirectly proved to be used as the markers of the treatment response of the lung adenocarcinoma immune checkpoint inhibitor. The risk score of this model will help clinicians to personalize treatment for LUAD patients.

The foregoing are merely exemplary embodiments of the present invention, which enable those skilled in the art to understand or practice the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.