WO2023056961A1 - Prognosis assessment method for lung adenocarcinoma based on dna methylation - Google Patents

Abstract

Provided herein is a method for determining the prognosis of lung adenocarcinoma in a subject on the basis of degree of DNA methylation, comprising: 1) detecting a degree of methylation of at least one methylation site on the DNA of a tumor sample from the subject; and 2) comparing the detected degree of methylation with a degree of methylation of a corresponding methylation site in patients having a known lung adenocarcinoma prognosis, so as to assess and/or predict a lung adenocarcinoma prognosis of the subject. Also provided herein is a kit for determining the prognosis of lung adenocarcinoma in a subject.

Description

A prognostic assessment method for lung adenocarcinoma based on DNA methylation

This application claims the priority of the Chinese patent application submitted to the State Intellectual Property Office of China on October 9, 2021, with the application number 202111175625.8 and the title of the invention "A Method for Prognosis Evaluation of Lung Adenocarcinoma Based on DNA Methylation", It is hereby incorporated by reference in its entirety.

technical field

This article relates to a method for judging the prognosis of lung adenocarcinoma in a subject, especially using the degree of methylation of a methylation site to assess and/or predict the prognosis of a lung adenocarcinoma subject. This document also relates to a kit for determining the prognosis of a subject with lung adenocarcinoma.

Background technique

Lung adenocarcinoma (lung adenocarcinoma) is a type of lung malignancy and is the most common histological subtype of non-small cell carcinoma (NSCLC). Unlike squamous cell lung cancer, lung adenocarcinoma is more likely to occur in women and non-smokers. The incidence rate is lower than that of squamous cell carcinoma and undifferentiated carcinoma, and the age of onset is younger. Lung adenocarcinoma generally has no obvious clinical symptoms in the early stage, and is often found during chest X-ray examination. Despite recent advances in early diagnostic methods and treatment with chemotherapy, radiotherapy, and surgery, the prognosis of patients with lung adenocarcinoma remains poor.

Abnormal DNA methylation in the genome and the occurrence of tumors have always been one of the hotspots in the medical field. Cell cycle, DNA repair, angiogenesis and cell apoptosis are all related to the methylation of related genes. The most likely regulatory role of DNA hypermethylation is to inhibit the expression of key genes, thereby determining the fate of the cell. For example, the study of abnormal DNA methylation in tumor cells has made many significant progress in various tumors. In mammals, methylation usually affects only the cytosine (CpG) preceding the guanine on the DNA strand. The methylation distribution of CpG dinucleotides in normal cells is not uniform. About 50% of the genes have CpG islands with concentrated distribution of CpG in the promoter region, ranging in length from 0.5 to 2kb. This region is closely related to the transcriptional regulation of genes. relation. The methylation of CpG islands in certain gene regulatory regions in the human body frequently occurs in related cancer cell tissues, and has been shown to be related to the onset, progression and prognosis of certain tumors, and drug sensitivity.

Nevertheless, there is still a lack of specific methylation sites and methods for judging the prognosis of lung adenocarcinoma.

Contents of the invention

In one aspect, provided herein is a method for judging the prognosis of lung adenocarcinoma in a subject based on the degree of DNA methylation, comprising:

1) detecting the degree of methylation of at least one methylation site on DNA from a tumor sample from the subject; and

2) comparing the detected degree of methylation with the degree of methylation of corresponding methylation sites in patients with known prognosis of lung adenocarcinoma, so as to evaluate and/or predict the prognosis of lung adenocarcinoma,

其中所述至少一个甲基化位点选自cg22288737、cg06919800、cg12550203、cg27664120、cg06580065、cg10535478、cg09552761、cg08720365、cg04458206、cg19931644、cg00125544、cg16404170、cg03271650、cg26390965、cg24662823、cg22129323、cg19283298、cg18140857、cg19375810、 cg26709300、cg01081930、cg13960576、cg15224955、cg14980255、cg13219896、cg02441149、cg12180319、cg12141052、g07970040、cg16348385、cg19595277、cg08783616、cg07081759、cg18148530、cg03925072、cg07109046、cg26508444、cg04511534、cg03961551、cg02619478、cg21496187、cg01442843、cg05337753、cg01759110、 g03003256、cg08147389、cg26341831、cg05226740、cg02480298、cg01040786、cg14422315、cg13695646、cg19096475、cg23146197、cg08535361、cg12729166、cg23987322、cg26566236、cg24876969、cg14011327、cg25417917、cg24178440、cg11382133、cg24684576、cg10313043、cg17381426、cg27106909、cg02431562、cg02029665、 cg03356689、cg03046247、cg05305140、cg07929642、cg14519392、cg10660256、cg15021670、cg20218077、cg23336905、cg11837749、cg00657287、cg24287218、cg16511076、cg22809408、cg25134306、cg00161124、cg05968188、cg25319237、cg13655250、cg12033622、cg26501657、cg00560072、cg24038192、cg17518215、cg06804210、 cg26119877、cg13206028、cg05762309、cg24688655、cg03583016、cg03968618、cg21657704、cg06136054、cg25300386、cg11198596、cg26649504、cg02140384、cg03340356、cg02286091、cg11893955、cg16442712、cg09550809、cg02687352、cg12911122、cg00009922、cg00842549、cg03138928、cg22623319、cg24341129、cg01804934、 cg13609385、cg02168442、cg25886621、cg19570155、cg04844207、cg10811426、cg16265859、cg02215430、cg14080475、cg19559984、cg06546607、cg06457011、cg09807875、cg20883174、cg07465627、cg12309653、cg19856705、cg19646897、cg08317694、cg26281789、cg01101865、cg07907506、cg18516609、cg16988611、cg08825225、 cg08983217、cg05230834、cg01449674、cg17171407、cg26561413、cg05890484、cg17181043、cg01465596、cg23202468、cg16585234、cg04481181、cg23994112、cg14180029、cg03020554、cg02918577、cg19741167、cg14236976、cg01320579、cg08866634、cg03478969、cg18058747、cg22761483、cg03462901、cg13829680、cg21101743、 cg15960924、cg11620672、cg11770080、cg00457684、cg01544903、cg09775533、cg02184673、cg19218509、cg16655240、cg01798157、cg20092936、cg16661628、cg00225623、cg09168604、cg07614018、cg02093902、cg21553700、cg06291867、cg16332610、cg00637178、cg01239704、cg15050865、cg16465407、cg24127345、cg21576886、 cg09477407、cg07309576、cg05198244、cg05022368、cg23067090、cg00591421、cg01615475、cg02663137、cg02217063、cg05406101、cg19230755、cg06661612、cg23401796、cg08269733、cg16696400、cg03548170、cg06956786、cg01290565、cg12592815、 cg13306338、cg10603235、cg12072740、cg07993507、cg16788865、cg01376079、 cg19766178、cg22534097、cg11791155、cg08252585、cg16939187、cg05538161、cg25950564、cg06768361、cg13378643、cg23530473、cg25045113、cg03860013、cg15135773、cg18842107、cg06360543、cg14952312、cg18372960、cg05386977、cg06121514、cg06450634、cg12862820、cg09049140、cg04575058、cg14372037、cg03918756、 cg05364179、cg05388545、cg16105471、cg09439722、cg27144670、cg12157156、cg06900571、cg01899620、cg05479166、cg16866485、cg05779630、cg01339008、cg05445326、cg21845080、cg06306006、cg16255280、cg17077566、cg16359316、cg06742077、cg00474746、cg09291474、cg10741603、cg26757836、cg05556202、cg13887966、 cg08813325、cg15002541、cg05256179、cg16331565、cg09975995、cg03565584、cg22716738、cg17450815、cg18560144、cg15445281、cg21672109、cg10124440、cg27488680、cg08553950、cg11240431、cg27419119、cg23414001、cg04294383、cg23217940、cg07737682、cg23837623、cg03084652、cg06392241、cg20268054、cg27664343、 cg16634951, cg17490981, cg08538509, cg21475526, cg05478314, and cg00278107.

In some embodiments, the methylation degree of any of the following sites is positively correlated with the risk of poor prognosis of said lung adenocarcinoma in said subject: cg22288737, cg06919800, cg12550203, cg27664120, cg06580065, cg10535478, cg09552761, cg08720365 、cg04458206、cg19931644、cg00125544、cg16404170、cg03271650、cg26390965、cg24662823、cg22129323、cg19283298、cg18140857、cg19375810、cg26709300、cg01081930、cg13960576、cg15224955、cg14980255、cg13219896、cg02441149、cg12180319、cg12141052、cg07970040、cg16348385、cg19595277、cg08783616、cg07081759 、cg18148530、cg03925072、cg07109046、cg26508444、cg04511534、cg03961551、cg02619478、cg21496187、cg01442843、cg05337753、cg01759110、cg03003256、cg08147389、cg26341831、cg05226740、cg02480298、cg01040786、cg14422315、cg13695646、cg19096475、cg23146197、cg08535361、cg12729166、cg23987322、cg26566236 、cg24876969、cg14011327、cg25417917、cg24178440、cg11382133、cg24684576、cg10313043、cg17381426、cg27106909、cg02431562、cg02029665、cg03356689、cg03046247、cg05305140、cg07929642、cg14519392、cg10660256、cg15021670、cg20218077、cg23336905、cg11837749、cg00657287、cg24287218、cg16511076、cg22809408 、cg25134306、cg00161124、cg05968188、cg25319237、cg13655250、cg12033622、cg26501657、cg00560072、cg24038192、cg17518215、cg06804210、cg26119877、cg13206028、cg05762309、cg24688655、cg03583016、cg03968618、cg21657704、cg06136054、cg25300386、cg11198596、cg26649504、cg02140384、cg03340356、cg02286091 、cg11893955、cg16442712、cg09550809、cg02687352、cg12911122、cg00009922、cg00842549、cg03138928、cg22623319、cg24341129、cg01804934、 cg13609385、cg02168442、cg25886621、cg19570155、cg04844207、cg10811426、cg16265859、cg02215430、cg14080475、cg19559984、cg06546607、cg06457011、cg09807875、cg20883174 、cg07465627、cg12309653、cg19856705、cg19646897、cg08317694、cg26281789、cg01101865、cg07907506、cg18516609、cg16988611、cg08825225、cg08983217、cg05230834、cg01449674、cg17171407、cg26561413、cg05890484、cg17181043、cg01465596、cg23202468、cg16585234、cg04481181、cg23994112、cg14180029、cg03020554 、cg02918577、cg19741167、cg14236976、cg01320579、cg08866634、cg03478969、cg18058747、cg22761483、cg03462901、cg13829680、cg21101743、cg15960924、cg11620672、cg11770080、cg00457684、cg01544903、cg09775533、cg02184673、cg19218509、cg16655240、cg01798157、cg20092936、cg16661628、cg00225623、cg09168604 , cg07614018, cg02093902, cg21553700, cg06291867, and cg16332610.

In some embodiments, the degree of methylation at any of the following sites is inversely correlated with the risk of poor prognosis of said lung adenocarcinoma in said subject: cg00637178, cg01239704, cg15050865, cg16465407, cg24127345, cg21576886, cg09477407, cg07309576 、cg05198244、cg05022368、cg23067090、cg00591421、cg01615475、cg02663137、cg02217063、cg05406101、cg19230755、cg06661612、cg23401796、cg08269733、cg16696400、cg03548170、cg06956786、cg01290565、cg12592815、cg13306338、cg10603235、cg12072740、cg07993507、cg16788865、cg01376079、cg19766178、cg22534097 、cg11791155、cg08252585、cg16939187、cg05538161、cg25950564、cg06768361、cg13378643、cg23530473、cg25045113、cg03860013、cg15135773、cg18842107、cg06360543、cg14952312、cg18372960、cg05386977、cg06121514、cg06450634、cg12862820、cg09049140、cg04575058、cg14372037、cg03918756、cg05364179、cg05388545 、cg16105471、cg09439722、cg27144670、cg12157156、cg06900571、cg01899620、cg05479166、cg16866485、cg05779630、cg01339008、cg05445326、cg21845080、cg06306006、cg16255280、cg17077566、cg16359316、cg06742077、cg00474746、cg09291474、cg10741603、cg26757836、cg05556202、cg13887966、cg08813325、cg15002541 、cg05256179、cg16331565、cg09975995、cg03565584、cg22716738、cg17450815、cg18560144、cg15445281、cg21672109、cg10124440、cg27488680、cg08553950、cg11240431、cg27419119、cg23414001、cg04294383、cg23217940、cg07737682、cg23837623、cg03084652、cg06392241、cg20268054、cg27664343、cg16634951、cg17490981 , cg08538509, cg21475526, cg05478314, and cg00278107.

In some embodiments, the at least one methylation site is two or more of the methylation sites listed above.

In some embodiments, the at least one methylation site is three or more of the methylation sites listed above.

In some embodiments, the at least one methylation site is five or more of the methylation sites listed above.

In some embodiments, the methylation sites include cg03046247, cg03356689, cg07109046, cg11770080, and cg08813325.

In some embodiments, the method further comprises evaluating and/or predicting the prognosis of the lung adenocarcinoma in the subject based on the weight of each methylation site.

In some embodiments, the prognosis of lung adenocarcinoma includes risk of death, and/or risk of recurrence, and/or risk of disease progression.

In some embodiments, the degree of methylation at the methylation site is detected by methylation chip, pyrosequencing, methylation qPCR, or WGBS.

In another aspect, provided herein are methods of assessing the efficacy of a treatment for lung adenocarcinoma comprising

1) treating a subject with lung adenocarcinoma with a lung adenocarcinoma therapy; and

2) judging the prognosis of the lung adenocarcinoma by the method described above,

Wherein the therapeutic effect of the lung adenocarcinoma treatment method is evaluated according to the prognosis of the lung adenocarcinoma.

In some embodiments, the method of treatment includes drug therapy, surgery, and/or radiation therapy.

On the other hand, this paper provides a method for determining the follow-up strategy of lung adenocarcinoma subjects, which includes judging the risk of poor prognosis of lung adenocarcinoma by the method described above, and determining the risk of poor prognosis of lung adenocarcinoma according to the risk of poor prognosis of lung adenocarcinoma Determine the follow-up strategy.

In another aspect, provided herein is a kit for determining the prognosis of lung adenocarcinoma, which includes reagents for detecting the degree of methylation of at least one methylation site listed above.

In some embodiments, the kit includes reagents for detecting the degree of methylation of at least two, at least three, or at least five of the methylation sites listed above.

In some embodiments, the methylation sites include cg03046247, cg03356689, cg07109046, cg11770080, and cg08813325.

In some embodiments, the kit is in the form of a methylation chip.

The methylation sites and kits provided herein can be used to judge or evaluate the prognosis of lung adenocarcinoma of a subject.

Description of drawings

Figure 1 shows the survival curves and receiver operating characteristic curves based on the degree of methylation at a single methylation site.

Figure 2 shows the survival curves and receiver operating characteristic curves of a simple summation model based on the degree of methylation at 5 methylation sites.

Figure 3 shows the survival curve and receiver operating characteristic curve of the methylation degree weight model based on the 5 methylation sites.

Detailed ways

Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by one of ordinary skill in the art.

A "methylation site" herein refers to a position of a nucleotide whose base has a methylation modification. DNA methylation in vertebrates generally occurs at the CpG site (that is, the site where cytosine is immediately followed by guanine in the DNA sequence), and cytosine is converted into 5-methylcytosine by DNA methyltransferase catalysis. Most CpG sites in the human genome are methylated, but some specific regions, such as CpG islands rich in cytosine and guanine, are usually not methylated. CpG methylation can affect the transcriptional activity of related genes. For example, methylation can inhibit tumor suppressor genes, while demethylation can stimulate the expression of some oncogenes. These situations may lead to cancer. In this paper, the specific methylation site, such as the methylation site ID represented by "cg+number", is provided by Illumina's Infinium MethylationEPIC product file (https://webdata.illumina.com/downloads/productfiles/ methylationEPIC/infinium-methylationepic-v-1-0-b5-manifest-file-csv.zip), which has a one-to-one positional relationship in the human (Homo sapiens) genome (version number: GRCh37). In addition, in the following Table 1 and Table 2, further descriptions are also made in the form of "chromosome number + position number".

Herein, "degree of methylation", also referred to as "level of methylation", refers to the proportion (or frequency) of a specific methylation site in a DNA sample being modified by methylation. Whether a site is methylated can be detected by various methods, including but not limited to, methylation chip, pyrosequencing, methylation qPCR, wgbs, etc. In one specific example, bisulfite treatment can be used to cause the deamination of unmethylated cytosine residues in DNA double-stranded molecules to uracil; while the methylated cytosine residues remain unchanged, which can then be The presence or absence of such changes is detected by, for example, sequencing or PCR amplification reactions (eg, using primers that are complementary only to the target sequence that contains uracil).

In this paper, "prognosis" refers to the prediction of the possible outcome or consequences of a disease (such as recovery, the appearance or disappearance of certain symptoms, signs and complications, and the death of patients), as well as the possibility of these outcomes or consequences occurring. sexual size. Prognosis is related to various factors such as the type of disease, the physical condition of the patient, whether there is appropriate treatment and whether the measures are taken in time. For example, benign tumors have a complete capsule, do not metastasize, grow slowly, and are easier to treat, so the prognosis is better; malignant tumors often have no clear boundaries, proliferate quickly, easily metastasize, and are difficult to treat thoroughly, so the prognosis is poor. Judgment of prognosis may also include providing temporal cues, such as predicting the likelihood of a certain outcome occurring within a certain period of time.

In this paper, the "positive correlation" between the degree of methylation and the poor prognosis of lung adenocarcinoma means that the higher the degree of methylation at this site, the higher the risk of poor prognosis of the patient; The "negative correlation" of poor prognosis means that the lower the degree of methylation of the site, the higher the risk of poor prognosis of the patient. The prognosis of lung adenocarcinoma can include events such as recurrence, progression, metastasis or patient death of lung adenocarcinoma. High risk of poor prognosis refers to disease recurrence or progression, and/or high risk of patient death.

In some embodiments of the methods of determining the prognosis of lung adenocarcinoma provided herein, the degree of methylation of a single methylation site is used to assess the prognosis of lung adenocarcinoma of the subject. In a preferred embodiment, multiple (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40 1, 50, 100 or more) methylation sites to assess the prognosis of lung adenocarcinoma of the subject. For example, in some examples, the degree of methylation at 5 methylation sites is detected and used to assess the prognosis of lung adenocarcinoma in the subject. In some embodiments, the prognosis of lung adenocarcinoma is assessed by simple summation of methylation levels at multiple (two or more) methylation sites. In some embodiments, the plurality of methylated sites includes at least one methylated site positively associated with poor prognosis of lung adenocarcinoma and at least one methylated site negatively associated with poor prognosis of lung adenocarcinoma . In a more preferred embodiment, when the methylation degree of multiple (two or more) methylation sites is used to evaluate the prognosis of lung adenocarcinoma, the index coefficient of each methylation site is also considered (or weight), that is, a weighted model of these methylated sites is used to evaluate the prognosis of lung adenocarcinoma. In the case of known combinations of methylation sites, a training set can be constructed based on known or available methylation degree and prognosis data of subject samples (such as tumor biopsies or surgical samples) to determine specific methylation sites. The weight of the chemicalization site. In a specific example, the weight of the methylation site can be determined by a multiple linear regression model.

A "subject" as used herein refers to a diagnosis, treatment, or clinical trial object, which can be either a disease patient or a healthy individual.

The methylation sites provided herein can be used to judge the prognosis of patients with lung adenocarcinoma. For example, these methylation sites can be used to prepare a kit for judging the prognosis of lung adenocarcinoma patients. The kit may include, for example, bisulfite (such as sodium bisulfite) for processing the sample to be tested, amplification primers or hybridization probes for specific methylation sites, sequencing reagents, and the like. The kit can also exist in the form of a methylation chip that can simultaneously detect the methylation degree of multiple methylation sites.

The present invention is further illustrated below through specific examples.

Example

Example 1 Single methylation site for prognosis prediction of lung adenocarcinoma

Through the 450k methylation chip sequencing results of tumor tissue samples of lung adenocarcinoma patients published in the TCGA database and our own 67 tumor tissue samples of lung adenocarcinoma patients sequenced with an 850k methylation chip, we selected multiple sites as To determine the prognostic markers of patients with lung adenocarcinoma. These sites are listed in Table 1 and Table 2, respectively. The selected methylation sites that can be used to predict the prognosis of lung adenocarcinoma are divided into two categories, that is, the degree of methylation is positively correlated with prognosis (188, Table 1) and negatively correlated (112, Table 2 ). For each locus, we performed predictions in 449 TCGA patients and compared them with actual outcomes. By comprehensively comparing the prognosis of the judged high-risk and low-risk groups (225vs 224), we calculated the risk coefficient (HR) and AUC value of each site judgment result, and the larger the HR value, the better the judgment effect. Well, the closer the AUC is to 1, the better the judgment effect. See Table 1 and Table 2 for statistical results. In addition, referring to Figure 1, we also plotted survival curves and receiver operating characteristic curves (AOC) for each site.

Table 1 Methylation sites and corresponding statistical data positively correlated with poor prognosis

Marker number	chromosome	Location	hr	rocauc
cg22288737	chr3	52198012	1.56465	0.58466
cg06919800	Chr17	80829520	1.52945	0.61495
cg12550203	chr12	122059396	1.77447	0.61766
cg27664120	chr7	99306649	1.59463	0.61393
cg06580065	chr18	22418045	1.56392	0.63745
cg10535478	chr6	132538504	1.83016	0.63501
cg09552761	Chr19	12806435	1.40767	0.60758
cg08720365	chr10	99809897	1.60305	0.61687
cg04458206	chr10	133390997	1.53136	0.61236
cg19931644	chr8	12765976	1.76521	0.59733
cg00125544	chr2	235957287	1.45951	0.57682

cg16404170	chr10	124642307	2.08229	0.6498
cg03271650	chr10	112376123	1.4854	0.63279
cg26390965	chr6	151349313	2.33993	0.67028
cg24662823	chr11	128480740	1.75893	0.59703
cg22129323	chr12	6463316	1.61231	0.63326
cg19283298	chr13	97437972	2.05513	0.64132
cg18140857	chr8	56321315	2.05807	0.6342
cg19375810	chr3	42746435	2.03099	0.64625
cg26709300	chr16	30095361	2.23189	0.67515
cg01081930	chr1	91818723	2.03408	0.67718
cg13960576	chr13	43406272	1.88205	0.59961
cg15224955	chr7	134565488	1.79803	0.61623
cg14980255	chr15	36898493	1.57649	0.65129
cg13219896	chr7	129147757	1.69727	0.62464
cg02441149	chr13	106507375	1.87036	0.62158
cg12180319	chr15	48413465	1.49878	0.60763
cg12141052	chr12	65955823	1.90591	0.63485
cg07970040	chr1	9729107	1.54629	0.58724
cg16348385	chr16	30095501	1.76089	0.63081
cg19595277	chr6	43123071	1.48874	0.60384
cg08783616	Chr17	16485979	1.99761	0.64431
cg07081759	chr10	124642336	1.80141	0.6483
cg18148530	chr10	79611026	1.86263	0.62456
cg03925072	chr8	37610955	1.70045	0.62643
cg07109046	chr13	40630251	1.79709	0.67445
cg26508444	chr10	124642276	1.87895	0.65594
cg04511534	Chr17	4560076	2.02838	0.65531
cg03961551	chr1	24925239	1.81192	0.60848
cg02619478	chr8	30390502	2.01318	0.622
cg21496187	chr6	100176636	1.62277	0.59472
cg01442843	chr1	206555712	1.64908	0.57711
cg05337753	Chr17	77377325	1.72088	0.68672
cg01759110	chr8	24384181	1.42908	0.56624
cg03003256	chr6	116528394	1.7945	0.6622
cg08147389	chr14	73289543	1.96619	0.66713
cg26341831	chr1	225848578	1.79034	0.58651
cg05226740	chr1	206497848	2.0538	0.6119
cg02480298	chr11	36399680	1.75658	0.6221
cg01040786	chr5	78748072	1.34137	0.60711
cg14422315	chr10	97904247	1.81532	0.59793
cg13695646	chr5	79111686	1.76691	0.65151
cg19096475	chr11	123195821	1.59499	0.64755
cg23146197	chr12	65877222	1.74318	0.65594
cg08535361	chr12	95783746	1.56893	0.61066
cg12729166	chr1	99644834	2.0332	0.64083
cg23987322	chr5	79111743	1.54976	0.59013
cg26566236	chr6	139138564	2.04181	0.6637
cg24876969	chr6	29004125	1.78312	0.70355

cg14011327	chr4	98663810	1.77827	0.6563
cg25417917	chr2	150595809	1.48836	0.58692
cg24178440	chr3	149299511	2.09167	0.63736
cg11382133	chr3	16512348	1.73836	0.65586
cg24684576	Chr17	44806011	1.53923	0.63586
cg10313043	chr10	98260938	1.7049	0.65116
cg17381426	chr7	140674325	1.64794	0.62959
cg27106909	chr16	30095576	1.65841	0.62942
cg02431562	chr2	96144231	1.75941	0.63534
cg02029665	Chr22	46045919	1.97441	0.68242
cg03356689	chr11	128716470	1.66811	0.63646
cg03046247	chr7	55038357	1.67176	0.58203
cg05305140	chr8	56321263	1.71205	0.63617
cg07929642	chr16	89324277	1.54715	0.55676
cg14519392	chr6	4140024	1.65501	0.62738
cg10660256	chr5	79111860	1.40684	0.58819
cg15021670	chr15	42094528	1.86603	0.62511
cg20218077	chr8	74061140	1.86046	0.65771
cg23336905	chr5	35803334	1.91429	0.61558
cg11837749	chr1	54581659	1.53333	0.60873
cg00657287	chr5	72459744	1.8596	0.63865
cg24287218	chr6	33073491	1.6789	0.59226
cg16511076	chr4	48885463	1.90676	0.643
cg22809408	chr6	31441923	1.48012	0.61392
cg25134306	chr5	139565692	2.17689	0.62832
cg00161124	chr1	234230513	1.97519	0.67489
cg05968188	chr6	16483766	1.70413	0.64955
cg25319237	chr13	29483327	1.83467	0.6511
cg13655250	chr9	131669141	1.94271	0.63742
cg12033622	chr20	31637770	1.49624	0.65467
cg26501657	chr12	111435751	1.72822	0.62984
cg00560072	chr1	61284871	1.85931	0.64398
cg24038192	chr7	139635761	1.28442	0.58242
cg17518215	chr2	172108513	1.67263	0.65295
cg06804210	chr2	86809825	1.73001	0.60275
cg26119877	chr14	37061270	1.78662	0.63185
cg13206028	chr10	113020595	1.6266	0.65434
cg05762309	chr6	30154066	1.79166	0.6272
cg24688655	chr2	199467254	1.78618	0.65292
cg03583016	chr2	199466952	1.83065	0.65647
cg03968618	chr4	185837509	1.77701	0.65683
cg21657704	Chr17	82893131	1.70597	0.65926
cg06136054	chr11	131674401	1.61732	0.61263
cg25300386	chr7	94394676	1.3885	0.60915
cg11198596	chr3	66497693	2.0455	0.64696
cg26649504	chr12	111758593	1.80946	0.65862
cg02140384	chr1	61053225	2.08532	0.63772
cg03340356	chr1	67135152	1.55626	0.61272

cg02286091	chr5	79111855	1.56074	0.62705
cg11893955	chr8	29061304	1.39996	0.56079
cg16442712	chr10	24250393	1.76813	0.60344
cg09550809	chr5	79111739	1.56404	0.61062
cg02687352	chr2	96891548	1.64112	0.62134
cg12911122	chr6	41548748	1.83019	0.59801
cg00009922	chr13	110215029	1.7561	0.58568
cg00842549	Chr17	48574558	1.46562	0.62516
cg03138928	chr1	225817124	1.85871	0.60888
cg22623319	chr10	110668153	2.07448	0.63792
cg24341129	chr9	16727832	1.57551	0.60828
cg01804934	chr6	33073452	1.47063	0.62304
cg13609385	chr8	41723414	1.65758	0.57804
cg02168442	chr8	101224294	1.53043	0.6074
cg25886621	chr12	92736475	1.92564	0.62915
cg19570155	chr2	36538462	1.76555	0.63734
cg04844207	chr11	36070638	1.81614	0.61436
cg10811426	chr1	234531715	1.37489	0.64127
cg16265859	chr1	19074575	1.82061	0.6386
cg02215430	chr6	41748675	1.76634	0.61369
cg14080475	chr9	114388215	1.66514	0.61579
cg19559984	chr7	28809316	1.80026	0.57896
cg06546607	Chr19	33522113	1.51482	0.57122
cg06457011	chr20	41138850	1.78454	0.64556
cg09807875	chr7	34096500	1.46309	0.62563
cg20883174	chr2	144502443	1.38708	0.59247
cg07465627	Chr17	55090046	1.46946	0.61563
cg12309653	chr15	36878253	1.7179	0.62553
cg19856705	chr1	234531754	1.41909	0.66327
cg19646897	chr5	157109368	1.63394	0.56867
cg08317694	chr15	78339802	1.57133	0.59196
cg26281789	chr14	91272676	1.39535	0.60345
cg01101865	chr14	62117847	1.53427	0.59935
cg07907506	chr3	100491738	1.5065	0.60414
cg18516609	chr10	31458312	1.49971	0.62593
cg16988611	chr10	80465190	1.54419	0.6548
cg08825225	chr11	128717065	1.66695	0.60392
cg08983217	chr2	96325411	1.72925	0.61156
cg05230834	chr8	118614750	1.35119	0.59133
cg01449674	chr10	31242753	1.43939	0.5703
cg17171407	chr8	118948538	1.56088	0.6278
cg26561413	Chr17	5584728	1.83618	0.62314
cg05890484	chr5	79111729	1.40916	0.62498
cg17181043	chr1	64495852	1.31906	0.5993
cg01465596	chr1	41916245	1.54195	0.60945
cg23202468	chr3	42664336	1.70936	0.66159
cg16585234	chr1	25618311	1.29099	0.61015
cg04481181	chr3	165197208	1.80119	0.64494

cg23994112	Chr17	77377375	1.76759	0.65454
cg14180029	chr11	36109772	1.71226	0.60204
cg03020554	chr8	98939502	1.68701	0.6036
cg02918577	chr15	93089916	1.60514	0.61847
cg19741167	chr2	86036147	1.90705	0.59885
cg14236976	chr9	126615710	1.76842	0.63135
cg01320579	Chr17	77409760	1.82199	0.63117
cg08866634	chr5	176535337	1.62757	0.62358
cg03478969	chr5	80570612	1.50923	0.61968
cg18058747	chr5	79112430	1.69007	0.60506
cg22761483	chr2	215624503	1.54012	0.6079
cg03462901	chr15	93070224	1.57083	0.62647
cg13829680	chr9	133881853	2.02842	0.61375
cg21101743	chr7	39609655	1.58873	0.63013
cg15960924	chr3	32109790	1.50491	0.60586
cg11620672	chr6	19298245	1.50927	0.59074
cg11770080	chr5	79112356	1.60851	0.63423
cg00457684	chr6	28531598	1.47637	0.61939
cg01544903	chr16	56663317	1.61921	0.65384
cg09775533	chr20	34269345	1.55638	0.58116
cg02184673	chr11	123195852	1.45336	0.65748
cg19218509	chr20	34268941	1.60916	0.58565
cg16655240	chr20	34269266	1.56407	0.57432
cg01798157	chr1	203307467	1.79353	0.63971
cg20092936	chr3	195873440	1.63084	0.61755
cg16661628	chr20	34269421	1.65637	0.56072
cg00225623	chr13	24271753	1.54435	0.58463
cg09168604	chr10	86367431	1.47245	0.61974
cg07614018	Chr19	18700765	1.88686	0.61809
cg02093902	chr2	45274393	1.74308	0.63709
cg21553700	chr3	195873539	1.64756	0.60185
cg06291867	chr10	90857405	1.51237	0.56074
cg16332610	chr15	74366206	1.39544	0.61542

Table 2 Methylation sites and corresponding statistical data negatively correlated with poor prognosis

Marker number	chromosome	Location	hr	rocauc
cg00637178	chr5	16466997	1.19981	0.59895
cg01239704	chr7	157812877	1.52042	0.61304
cg15050865	chr5	19886491	1.53543	0.60448
cg16465407	chr7	9634160	1.30863	0.60201
cg24127345	chr10	1518276	1.66268	0.596
cg21576886	chr1	158930298	1.44222	0.57853
cg09477407	chr12	104303767	1.59452	0.57554
cg07309576	chr11	5442538	1.62695	0.61139
cg05198244	Chr21	25360604	1.62133	0.61111
cg05022368	chr8	39001033	1.76004	0.60087
cg23067090	chr15	25564371	1.52031	0.583

cg00591421	chr10	104708978	1.8293	0.60305
cg01615475	chr11	50268028	1.63934	0.59382
cg02663137	Chr19	6745706	1.43346	0.60337
cg02217063	chr16	6298708	1.44836	0.60699
cg05406101	Chr21	29019898	1.61413	0.60236
cg19230755	chr7	66413516	1.43628	0.6295
cg06661612	Chr21	45427666	1.6172	0.56326
cg23401796	chr8	140238568	1.69842	0.60213
cg08269733	chr16	62023275	1.64934	0.62932
cg16696400	chr16	7314321	1.81361	0.56467
cg03548170	chr1	71916962	1.5132	0.62137
cg06956786	chr7	3673716	1.5597	0.62886
cg01290565	chr12	111037434	1.54662	0.57813
cg12592815	chr7	4260674	1.57684	0.59519
cg13306338	chr7	4127975	1.30431	0.64119
cg10603235	Chr22	42940043	1.55144	0.58591
cg12072740	chr12	129720609	1.63081	0.5756
cg07993507	Chr22	47630818	1.49098	0.586
cg16788865	chr12	46385018	1.98203	0.60309
cg01376079	chr11	67302762	1.45824	0.56441
cg19766178	chr7	3174767	1.51921	0.64902
cg22534097	chr10	90921325	1.63229	0.60601
cg11791155	chr7	4208128	1.55262	0.62304
cg08252585	chr2	24708747	1.69517	0.59305
cg16939187	chr7	2572337	1.41109	0.56934
cg05538161	chr7	4203137	1.71946	0.62443
cg25950564	chr15	23447422	1.65108	0.61273
cg06768361	chr12	117042528	2.01995	0.65204
cg13378643	chr7	4107800	1.44241	0.60612
cg23530473	chr14	105624464	1.69264	0.56482
cg25045113	Chr17	8798781	1.56795	0.61591
cg03860013	chr20	33168207	1.42455	0.60162
cg15135773	chr7	4268740	1.43464	0.6084
cg18842107	chr10	113094096	1.68918	0.6052
cg06360543	chr1	156920264	1.6027	0.59959
cg14952312	Chr17	74354571	1.57701	0.59776
cg18372960	chr7	4266096	1.77117	0.62042
cg05386977	chr7	4260550	1.55539	0.61104
cg06121514	chr15	25117041	1.5075	0.60702
cg06450634	chr16	30418723	1.48426	0.58164
cg12862820	chr7	84188082	1.52541	0.61894
cg09049140	wxya	18993010	1.59314	0.57463
cg04575058	chr7	130440585	1.36965	0.62155
cg14372037	chr4	7524944	1.35864	0.60624
cg03918756	chr8	140238533	1.76568	0.58835
cg05364179	chr4	766251	1.54812	0.58812
cg05388545	Chr17	30825950	1.5207	0.59776
cg16105471	Chr17	28383618	1.78255	0.57257

cg09439722	chr7	4190511	1.69858	0.64111
cg27144670	chr15	51869241	1.91195	0.57824
cg12157156	chr12	124395627	1.45866	0.55006
cg06900571	chr8	140238483	1.70659	0.60942
cg01899620	chr13	113035108	1.68711	0.60138
cg05479166	Chr17	81642320	1.40391	0.59136
cg16866485	chr7	4236588	1.52748	0.60855
cg05779630	chr4	7471379	2.15714	0.62728
cg01339008	chr15	25080146	1.66982	0.65252
cg05445326	chr3	196338698	1.71515	0.60302
cg21845080	chr3	196338435	1.61002	0.61026
cg06306006	chr3	184300385	1.77275	0.64639
cg16255280	chr7	4179163	1.65128	0.59974
cg17077566	chr15	25090598	1.73289	0.65167
cg16359316	chr14	105624823	1.46579	0.56329
cg06742077	chr8	1816712	1.43036	0.59312
cg00474746	chr11	35663251	1.60837	0.63312
cg09291474	chr15	64930943	1.73667	0.55426
cg10741603	chr7	4203972	1.52584	0.6304
cg26757836	chr4	7303012	1.47619	0.6032
cg05556202	chr3	196338235	1.42959	0.5876
cg13887966	chr15	59276513	1.96904	0.62091
cg08813325	chr4	7697884	1.51588	0.61212
cg15002541	chr11	392903	1.65927	0.5843
cg05256179	chr6	31690399	1.59305	0.59248
cg16331565	chr8	141431488	1.46778	0.58512
cg09975995	chr4	174978052	1.89554	0.62922
cg03565584	Chr19	17296618	1.44364	0.57245
cg22716738	chr7	27745313	1.64625	0.58277
cg17450815	chr12	53190217	1.72149	0.58762
cg18560144	chr15	24985053	1.73508	0.61098
cg15445281	chr13	50589590	1.85554	0.59456
cg21672109	Chr22	42271889	1.45388	0.56611
cg10124440	chr12	6851131	1.69481	0.62855
cg27488680	chr4	7636495	1.66903	0.61688
cg08553950	chr20	37383613	1.43562	0.5726
cg11240431	chr8	139959000	1.54181	0.59325
cg27419119	chr12	57166118	1.55616	0.63031
cg23414001	chr20	35093267	1.72956	0.59285
cg04294383	chr15	22918276	1.45715	0.57173
cg23217940	chr15	22918250	1.72857	0.5941
cg07737682	chr16	2275985	1.55155	0.5844
cg23837623	Chr17	1574914	1.4052	0.58675
cg03084652	chr4	7636323	1.40175	0.60265
cg06392241	chr12	93377570	1.58989	0.56789
cg20268054	chr11	393029	1.76272	0.61619
cg27664343	chr12	132719638	1.40569	0.56672
cg16634951	chr12	130998756	1.55623	0.5877

cg17490981	chr5	154779440	1.94629	0.64071
cg08538509	chr13	30203413	1.51265	0.57772
cg21475526	chr12	917122	1.4373	0.60211
cg05478314	chr12	120737871	1.53223	0.58482
cg00278107	chr5	1061138	1.47345	0.6053

Example 2 Build a prognostic model by combining the above sites

Although a single locus has been able to preliminarily predict the prognosis of patients with lung adenocarcinoma, a combination of multiple loci can better predict the prognosis. Here we take a model including 5 sites as an example cg03046247+cg03356689+cg07109046-cg08813325+cg11770080, among which the degree of methylation of four sites is positively correlated with poor prognosis, and the fourth site is negatively correlated.

Taking 3 samples as an example to calculate: sample TCGA-49-AAR9, the methylation degree of the five sites is 0.833717, 0.596238, 0.467905, 0.28985, 0.492296, and the sum score is 2.100306; sample TCGA-05-4415, five The degree of methylation of each site was 0.794832, 0.434625, 0.75571, 0.378951, 0.579038 respectively, and the sum score was 2.185254. Both are in the prognostic high risk group based on the score. For sample TCGA-49-AAR2, the methylation degrees of the five sites were 0.287222, 0.096838, 0.192056, 0.547331, and 0.041958, respectively, and the sum score was 0.070743. This sample was in the low prognostic risk group based on the score.

Figure 2 is the result of predicting the prognosis of 449 patients through this model. The judgment result, HR is 2.9, AUC is 0.796, far better than the judgment of a single site.

Example 3 Establishing a prognostic model by combining the above sites and assigning different weight coefficients

Although simply combining multiple sites can predict the prognosis of patients with lung adenocarcinoma well, if the importance of each site in prediction is considered during model training, the accuracy of prediction can be further improved. Here we also take the 5 sites in Example 2 as an example, and use the formula 4.04731*cg03046247+2.04107*cg03356689+2.18439*cg07109046-4.10464*cg08813325+1.87320*cg11770080 to judge.

Also take the three samples mentioned in Example 2 as examples for calculation: sample TCGA-49-AAR9, the methylation degrees of the five sites are 0.833717, 0.596238, 0.467905, 0.28985, 2.100306, and the summed score is 5.345800612; For sample TCGA-05-4415, the methylation degrees of the five sites were 0.794832, 0.434625, 0.75571, 0.378951, and 0.579038, respectively, and the summed score was 5.283993467. Both are in the prognostic high risk group based on the score. For sample TCGA-49-AAR2, the methylation degrees of the five sites were 0.287222, 0.096838, 0.192056, 0.547331, and 0.041958, respectively, and the sum score was -0.388346175. This sample was in the low prognostic risk group based on the score.

Figure 3 shows the results of predicting the prognosis of 449 patients through this model. The judgment result, HR is 3.4, AUC is 0.810, the result is better than the judgment of simple sum of five sites.

Claims (17)

1. A method for judging the prognosis of lung adenocarcinoma in a subject based on the degree of DNA methylation, including:

   1) detecting the degree of methylation of at least one methylation site on DNA from a tumor sample from the subject; and

   2) comparing the detected degree of methylation with the degree of methylation of corresponding methylation sites in patients with known prognosis of lung adenocarcinoma, so as to evaluate and/or predict the prognosis of lung adenocarcinoma,

   其中所述至少一个甲基化位点选自cg22288737、cg06919800、cg12550203、cg27664120、cg06580065、cg10535478、cg09552761、cg08720365、cg04458206、cg19931644、cg00125544、cg16404170、cg03271650、cg26390965、cg24662823、cg22129323、cg19283298、cg18140857、cg19375810、 cg26709300、cg01081930、cg13960576、cg15224955、cg14980255、cg13219896、cg02441149、cg12180319、cg12141052、g07970040、cg16348385、cg19595277、cg08783616、cg07081759、cg18148530、cg03925072、cg07109046、cg26508444、cg04511534、cg03961551、cg02619478、cg21496187、cg01442843、cg05337753、cg01759110、 g03003256、cg08147389、cg26341831、cg05226740、cg02480298、cg01040786、cg14422315、cg13695646、cg19096475、cg23146197、cg08535361、cg12729166、cg23987322、cg26566236、cg24876969、cg14011327、cg25417917、cg24178440、cg11382133、cg24684576、cg10313043、cg17381426、cg27106909、cg02431562、cg02029665、 cg03356689、cg03046247、cg05305140、cg07929642、cg14519392、cg10660256、cg15021670、cg20218077、cg23336905、cg11837749、cg00657287、cg24287218、cg16511076、cg22809408、cg25134306、cg00161124、cg05968188、cg25319237、cg13655250、cg12033622、cg26501657、cg00560072、cg24038192、cg17518215、cg06804210、 cg26119877、cg13206028、cg05762309、cg24688655、cg03583016、cg03968618、cg21657704、cg06136054、cg25300386、cg11198596、cg26649504、cg02140384、cg03340356、cg02286091、cg11893955、cg16442712、cg09550809、cg02687352、cg12911122、cg00009922、cg00842549、cg03138928、cg22623319、cg24341129、cg01804934、 cg13609385、cg02168442、cg25886621、cg19570155、cg04844207、cg10811426、cg16265859、cg02215430、cg14080475、cg19559984、cg06546607、cg06457011、cg09807875、cg20883174、cg07465627、cg12309653、cg19856705、cg19646897、cg08317694、cg26281789、cg01101865、cg07907506、cg18516609、cg16988611、cg08825225、 cg08983217、cg05230834、cg01449674、cg17171407、cg26561413、cg05890484、cg17181043、cg01465596、cg23202468、cg16585234、cg04481181、cg23994112、cg14180029、cg03020554、cg02918577、cg19741167、cg14236976、cg01320579、cg08866634、cg03478969、cg18058747、cg22761483、cg03462901、cg13829680、cg21101743、 cg15960924、cg11620672、 cg11770080、cg00457684、cg01544903、cg09775533、cg02184673、cg19218509、cg16655240、cg01798157、cg20092936、cg16661628、cg00225623、cg09168604、cg07614018、cg02093902、cg21553700、cg06291867、cg16332610、cg00637178、cg01239704、cg15050865、cg16465407、cg24127345、cg21576886、 cg09477407、cg07309576、cg05198244、cg05022368、cg23067090、cg00591421、cg01615475、cg02663137、cg02217063、cg05406101、cg19230755、cg06661612、cg23401796、cg08269733、cg16696400、cg03548170、cg06956786、cg01290565、cg12592815、cg13306338、cg10603235、cg12072740、cg07993507、cg16788865、cg01376079、 cg19766178、cg22534097、cg11791155、cg08252585、cg16939187、cg05538161、cg25950564、cg06768361、cg13378643、cg23530473、cg25045113、cg03860013、cg15135773、cg18842107、cg06360543、cg14952312、cg18372960、cg05386977、cg06121514、cg06450634、cg12862820、cg09049140、cg04575058、cg14372037、cg03918756、 cg05364179、cg05388545、cg16105471、cg09439722、cg27144670、cg12157156、cg06900571、cg01899620、cg05479166、cg16866485、cg05779630、cg01339008、cg05445326、cg21845080、cg06306006、cg16255280、cg17077566、cg16359316、cg06742077、cg00474746、cg09291474、cg10741603、cg26757836、cg05556202、cg13887966、 cg08813325、cg15002541、cg05256179、cg16331565、cg09975995、cg03565584、cg22716738、cg17450815、cg18560144、cg15445281、cg21672109、cg10124440、cg27488680、cg08553950、cg11240431、cg27419119、cg23414001、cg04294383、cg23217940、cg07737682、cg23837623、cg03084652、cg06392241、cg20268054、cg27664343、 cg16634951, cg17490981, cg08538509, cg21475526, cg05478314, and cg00278107.
2. The method of claim 1, wherein the degree of methylation at any of the following sites is positively correlated with the risk of poor prognosis of the lung adenocarcinoma of the subject:

   cg22288737、cg06919800、cg12550203、cg27664120、cg06580065、cg10535478、cg09552761、cg08720365、cg04458206、cg19931644、cg00125544、cg16404170、cg03271650、cg26390965、cg24662823、cg22129323、cg19283298、cg18140857、cg19375810、cg26709300、cg01081930、cg13960576、cg15224955、cg14980255、cg13219896、 cg02441149、cg12180319、cg12141052、cg07970040、cg16348385、cg19595277、cg08783616、cg07081759、cg18148530、cg03925072、cg07109046、cg26508444、cg04511534、cg03961551、cg02619478、cg21496187、cg01442843、cg05337753、cg01759110、cg03003256、cg08147389、cg26341831、cg05226740、cg02480298、cg01040786、 cg14422315、cg13695646、cg19096475、cg23146197、cg08535361、cg12729166、cg23987322、cg26566236、cg24876969、cg14011327、cg25417917、cg24178440、cg11382133、cg24684576、cg10313043、cg17381426、cg27106909、cg02431562、cg02029665、cg03356689、cg03046247、cg05305140、 cg07929642、cg14519392、cg10660256、 cg15021670、cg20218077、cg23336905、cg11837749、cg00657287、cg24287218、cg16511076、cg22809408、cg25134306、cg00161124、cg05968188、cg25319237、cg13655250、cg12033622、cg26501657、cg00560072、cg24038192、cg17518215、cg06804210、cg26119877、cg13206028、cg05762309、cg24688655、cg03583016、cg03968618、 cg21657704、cg06136054、cg25300386、cg11198596、cg26649504、cg02140384、cg03340356、cg02286091、cg11893955、cg16442712、cg09550809、cg02687352、cg12911122、cg00009922、cg00842549、cg03138928、cg22623319、cg24341129、cg01804934、cg13609385、cg02168442、cg25886621、cg19570155、cg04844207、cg10811426、 cg16265859、cg02215430、cg14080475、cg19559984、cg06546607、cg06457011、cg09807875、cg20883174、cg07465627、cg12309653、cg19856705、cg19646897、cg08317694、cg26281789、cg01101865、cg07907506、cg18516609、cg16988611、cg08825225、cg08983217、cg05230834、cg01449674、cg17171407、cg26561413、cg05890484、 cg17181043、cg01465596、cg23202468、cg16585234、cg04481181、cg23994112、cg14180029、cg03020554、cg02918577、cg19741167、cg14236976、cg01320579、cg08866634、cg03478969、cg18058747、cg22761483、cg03462901、cg13829680、cg21101743、cg15960924、cg11620672、cg11770080、cg00457684、cg01544903、cg09775533、 cg02184673, cg19218509, cg16655240, cg01798157, cg20092936, cg16661628, cg00225623, cg09168604, cg07614018, cg02093902, cg215653700, cg3763 and 2.29
3. The method according to claim 1 or 2, wherein the degree of methylation at any of the following sites is negatively correlated with the risk of poor prognosis of the lung adenocarcinoma of the subject:

   cg00637178、cg01239704、cg15050865、cg16465407、cg24127345、cg21576886、cg09477407、cg07309576、cg05198244、cg05022368、cg23067090、cg00591421、cg01615475、cg02663137、cg02217063、cg05406101、cg19230755、cg06661612、cg23401796、cg08269733、cg16696400、cg03548170、cg06956786、cg01290565、cg12592815、 cg13306338、cg10603235、cg12072740、cg07993507、cg16788865、cg01376079、cg19766178、cg22534097、cg11791155、cg08252585、cg16939187、cg05538161、cg25950564、cg06768361、cg13378643、cg23530473、cg25045113、cg03860013、cg15135773、cg18842107、cg06360543、cg14952312、cg18372960、cg05386977、cg06121514、 cg06450634、cg12862820、cg09049140、cg04575058、cg14372037、cg03918756、cg05364179、cg05388545、cg16105471、cg09439722、cg27144670、cg12157156、cg06900571、cg01899620、cg05479166、cg16866485、cg05779630、cg01339008、cg05445326、cg21845080、cg06306006、cg16255280、cg17077566、cg16359316、cg06742077、 cg00474746、cg09291474、cg10741603、cg26757836、cg05556202、cg13887966、cg08813325、cg15002541、cg05256179、 cg16331565、cg09975995、cg03565584、cg22716738、cg17450815、cg18560144、cg15445281、cg21672109、cg10124440、cg27488680、cg08553950、cg11240431、cg27419119、cg23414001、cg04294383、cg23217940、 cg07737682, cg23837623, cg03084652, cg06392241, cg20268054, cg27664343, cg16634951, cg17490981, cg08538509, cg21475526, cg05478314, and cg07027.
4. The method according to any one of claims 1-3, wherein said at least one methylation site is two or more of the methylation sites listed in claim 1.
5. The method according to any one of claims 1-4, wherein said at least one methylation site is three or more of the methylation sites listed in claim 1.
6. The method according to any one of claims 1-5, wherein said at least one methylation site is five or more of the methylation sites listed in claim 1.
7. The method according to any one of claims 1-6, wherein the methylation sites include cg03046247, cg03356689, cg07109046, cg11770080 and cg08813325.
8. The method according to any one of claims 1-7, further comprising evaluating and/or predicting the prognosis of the subject's lung adenocarcinoma according to the weight of each methylation site.
9. The method according to any one of claims 1-8, wherein the prognosis of lung adenocarcinoma includes risk of death, and/or risk of recurrence, and/or risk of disease progression.
10. The method according to any one of claims 1-9, wherein the methylation degree of the methylation site is detected by methylation chips, pyrosequencing, methylation qPCR, or WGBS.
11. Methods of assessing the therapeutic effect of a treatment for lung adenocarcinoma, including

    1) treating a subject with lung adenocarcinoma with a lung adenocarcinoma therapy; and

    2) judge the prognosis of described lung adenocarcinoma with the method described in any one of claim 1-10,

    Wherein the therapeutic effect of the lung adenocarcinoma treatment method is evaluated according to the prognosis of the lung adenocarcinoma.
12. The method of claim 11, wherein said treatment method comprises drug treatment, surgery treatment and/or radiation treatment.
13. A method for determining a follow-up strategy for lung adenocarcinoma subjects, comprising judging the risk of poor prognosis of the lung adenocarcinoma according to any one of claims 1-10, and determining the risk of poor prognosis of the lung adenocarcinoma according to the risk of poor prognosis of the lung adenocarcinoma Determine the follow-up strategy.
14. The kit for judging the prognosis of lung adenocarcinoma comprises reagents for detecting the degree of methylation of at least one methylation site listed in claim 1.
15. The kit according to claim 14, comprising reagents for detecting the degree of methylation of at least two, at least three, or at least five methylation sites listed in claim 1.
16. The kit according to claim 14 or 15, wherein the methylation sites include cg03046247, cg03356689, cg07109046, cg11770080 and cg08813325.
17. The kit according to any one of claims 14-16, which is in the form of a methylation chip.

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