Disclosure of Invention
The invention aims to provide a method for realizing gastric adenocarcinoma metastasis diagnosis, gastric adenocarcinoma prediction prognosis and gastric adenocarcinoma prognosis evaluation by detecting the difference of LRRTM1 gene expression.
The invention also aims to provide a method for treating gastric adenocarcinoma metastasis by promoting the expression of LRRTM1 gene.
The invention also aims to provide a method for screening a gastric adenocarcinoma metastasis treatment drug.
The fourth purpose of the invention is to provide a medicine for treating gastric adenocarcinoma metastasis.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention provides an application of a reagent for detecting LRRTM1 in preparation of tools for gastric adenocarcinoma metastasis diagnosis, gastric adenocarcinoma prediction prognosis and gastric adenocarcinoma prognosis evaluation.
Further, the reagent for detecting LRRTM1 comprises a reagent for detecting the expression level of LRRTM1 gene.
Further, the reagent for detecting LRRTM1 includes a reagent capable of quantifying mRNA of LRRTM1 gene, and/or a reagent capable of quantifying LRRTM1 protein.
The agent for quantifying LRRTM1 gene mRNA of the present invention can exert its function based on a known method using a nucleic acid molecule: such as PCR, e.g., Southern hybridization, Northern hybridization, dot hybridization, Fluorescence In Situ Hybridization (FISH), DNA microarray, ASO methods, high throughput sequencing platforms, etc. The assay can be performed qualitatively, quantitatively, or semi-quantitatively using the reagent.
Further, the PCR method is a known method, for example, ARMS (Amplification Refractorymutation System) method, RT-PCR (reverse transcriptase-PCR) method, nested PCR method, or the like. The amplified nucleic acid can be detected by using a dot blot hybridization method, a surface plasmon resonance method (SPR method), a PCR-RFLP method, an in situ RT-PCR method, a PCR-SSO (sequence specific oligonucleotide) method, a PCR-SSP method, an AMPFLP (amplifiable fragment length polymorphism) method, an MVR-PCR method, and a PCR-SSCP (single strand conformation polymorphism) method.
The reagent capable of quantifying mRNA of LRRTM1 gene can be a specific primer of LRRTM1 gene or transcript, can also be a specific recognition probe of LRRTM1 gene or transcript, or comprises the primer and the probe.
The specific primers for LRRTM1 gene or transcript described above include primers for specific amplification of LRRTM1 gene used in real-time quantitative PCR. In a specific embodiment of the invention, the primer sequences are shown as SEQ ID NO.1 and SEQ ID NO. 2.
The primer can be prepared by chemical synthesis, appropriately designed by referring to known information using a method known to those skilled in the art, and prepared by chemical synthesis.
The probe may be prepared by chemical synthesis, by appropriately designing with reference to known information using a method known to those skilled in the art, and by chemical synthesis, or may be prepared by preparing a gene containing a desired nucleic acid sequence from a biological material and amplifying it using a primer designed to amplify the desired nucleic acid sequence.
The reagent for quantifying LRRTM1 protein of the present invention can exert its function based on a known method using an antibody: for example, ELISA, radioimmunoassay, immunohistochemistry, Western blotting, etc. may be included.
The reagent for quantifying LRRTM1 protein of the present invention includes an antibody or a fragment thereof that specifically binds to LRRTM1 protein. An antibody or fragment thereof of any structure, size, immunoglobulin class, origin, etc., may be used so long as it binds to the target protein. The antibodies or fragments thereof included in the assay products of the invention may be monoclonal or polyclonal. An antibody fragment refers to a portion of an antibody (partial fragment) or a peptide containing a portion of an antibody that retains the binding activity of the antibody to an antigen. Antibody fragments may include F (ab')2Fab', Fab, single chain fv (scfv), disulfide-bonded fv (dsfv) or polymers thereof, dimerized V regions (diabodies), or CDR-containing peptides. The reagent for quantifying LRRTM1 protein of the present invention may include an isolated nucleic acid encoding an amino acid sequence of an antibody or encoding a fragment of an antibody, a vector containing the nucleic acid, and a cell carrying the vector.
Antibodies can be obtained by methods well known to those skilled in the art. For example, mammalian cell expression vectors that retain all or part of the target protein or incorporate polynucleotides encoding them are prepared as antigens. After immunizing an animal with an antigen, immune cells are obtained from the immunized animal and myeloma cells are fused to obtain hybridomas. The antibody is then collected from the hybridoma culture. Finally, a monoclonal antibody against LRRTM1 protein can be obtained by subjecting the obtained antibody to antigen-specific purification using LRRTM1 protein or a part thereof used as an antigen. Polyclonal antibodies can be prepared as follows: an animal is immunized with the same antigen as above, a blood sample is collected from the immunized animal, serum is separated from the blood, and then antigen-specific purification is performed on the serum using the above antigen. The antibody fragment can be obtained by treating the obtained antibody with an enzyme or by using sequence information of the obtained antibody.
Binding of the label to the antibody or fragment thereof can be carried out by methods generally known in the art. For example, proteins or peptides may be fluorescently labeled as follows: the protein or peptide is washed with phosphate buffer, a dye prepared with DMSO, a buffer, or the like is added, and the solution is mixed and left at room temperature for 10 minutes. In addition, labeling may be carried out using commercially available labeling kits, such as biotin labeling kit, e.g., biotin labeling kit-NH 2, biotin labeling kit-SH (Dojindo laboratories); alkaline phosphatase labeling kits such as alkaline phosphatase labeling kit-NH 2, alkaline phosphatase labeling kit-sh (dojindo laboratories); peroxidase labeling kits such as peroxidase labeling kit-NH 2, peroxidase labeling kit-NH 2(Dojindo Laboratories); phycobiliprotein labeling kits such as phycobiliprotein labeling kit-NH 2, phycobiliprotein labeling kit-SH, B-phycoerythrin labeling kit-NH 2, B-phycoerythrin labeling kit-SH, R-phycoerythrin labeling kit-NH 2, R-phycoerythrin labeling kit SH (dojindo laboratories); fluorescent labeling kits such as fluorescein labeling kit-NH 2, HiLyte Fluor (TM)555 labeling kit-NH 2, HiLyte Fluor (TM)647 labeling kit-NH 2(Dojindo Laboratories); and DyLight 547 and DyLight647(Techno Chemical Corp.), Zenon (TM), Alexa Fluor (TM) antibody labeling kit, Qdot (TM) antibody labeling kit (Invitrogen Corporation), and EZ-marker protein labeling kit (Funakoshi Corporation). For proper labeling, a suitable instrument can be used to detect the labeled antibody or fragment thereof.
The obtaining of the sample for detecting the expression level of LRRTM1 gene according to the present invention is a routine technique in the art, and is preferably obtained by a method that can be selected to be non-invasive or minimally invasive.
The sample may be (but is not limited to): peripheral blood, bone marrow, lymph nodes, peritoneal lavage fluid, parietal cells or gastric juice. In a specific embodiment of the invention, the sample is from a tissue of a subject.
It is well known to those skilled in the art that cells from tumor tissue are shed into body fluids, these shed cells are called circulating tumor cells, which have the same properties as the tumor cells in tumor tissue, and thus the detection of the properties of the circulating tumor cells in body fluids, especially in blood, can represent the properties of the tumor tissue. In the invention, the diagnosis of gastric adenocarcinoma metastasis can be realized by detecting the expression of LRRTM1 gene in tumor tissue, and the diagnosis of gastric adenocarcinoma metastasis can be easily obtained or can be realized by detecting the expression of LRRTM1 gene in circulating tumor cells.
The invention also provides a tool for gastric adenocarcinoma metastasis diagnosis, gastric adenocarcinoma prediction prognosis and gastric adenocarcinoma prognosis evaluation, wherein the tool can detect the expression level of the LRRTM1 gene.
Further, the tool comprises an agent capable of quantifying LRRTM1 gene mRNA, and/or an agent capable of quantifying LRRTM1 protein.
Typically, the reagents are present in suitable containers. Each of the primers or probes can be adjusted to at least one desired amount of concentration using a diluent such as deionized water and dispensed into a container.
Further, the reagent capable of quantifying mRNA of the LRRTM1 gene comprises a primer for specifically amplifying the LRRTM1 gene used in real-time quantitative PCR, and the sequence of the primer is shown as SEQ ID NO.1 and SEQ ID NO. 2.
Further, the tools for gastric adenocarcinoma metastasis diagnosis, gastric adenocarcinoma prognosis prediction and gastric adenocarcinoma prognosis evaluation include, but are not limited to, a chip, a kit, a test paper or a high throughput sequencing platform; the high-throughput sequencing platform is a special tool, and with the development of high-throughput sequencing technology, the construction of a gene expression profile of a person becomes very convenient work. By comparing the gene expression profiles of patients with diseases and normal people, the abnormality of which gene is related to the disease can be easily analyzed. Therefore, the knowledge that the abnormality of the LRRTM1 gene is related to gastric adenocarcinoma metastasis in high-throughput sequencing also belongs to a new application using the LRRTM1 of the invention and is also within the protection scope of the invention.
The kit of the present invention may further comprise a reagent for extracting nucleic acid, a reagent for PCR, a reagent for staining or developing color, and the like. For example, such agents include, but are not limited to: an extraction solution, an amplification solution, a hybridization solution, a color development solution, a washing solution, and the like.
In addition, the kit can also comprise instructions and the like for describing a method for detecting gastric adenocarcinoma metastasis.
The kit of the present invention may contain a plurality of different reagents suitable for practical use (e.g., for different detection methods), and is not limited to the reagents listed so far, and any reagent that determines metastasis of gastric adenocarcinoma based on the detection of LRRTM1 gene or transcript is included in the scope of the present invention.
The invention also provides a method for diagnosing gastric adenocarcinoma metastasis, predicting gastric adenocarcinoma prognosis or evaluating gastric adenocarcinoma prognosis, which comprises the following steps:
(1) obtaining a sample from a subject;
(2) detecting the expression level of LRRTM1 gene in the sample of the subject;
(3) correlating the measured LRRTM1 gene expression level with a disease association of the subject.
(4) The statistically decreased expression level of LRRTM1 gene compared to normal controls indicates that the subject is judged to have metastasized gastric adenocarcinoma, or that the prognosis of the subject is poor for predicting metastasis of gastric adenocarcinoma, or that the subject being assessed for metastasis of gastric adenocarcinoma has relapsed.
The invention also provides a therapeutic method for gastric adenocarcinoma metastasis, which comprises promoting the expression of LRRTM1 gene, or promoting the expression of LRRTM1 protein or enhancing the activity of LRRTM1 protein. .
The invention also provides a screening method of the tumor drug, which can measure the effect of the tumor drug on improving tumor prognosis by measuring the expression level of LRRTM1 gene or LRRTM1 protein at a certain period after adding the test drug to cancer cells or after applying the test drug to tumor model animals. More specifically, when the expression level of LRRTM1 gene or LRRTM1 protein is elevated or restored to a normal level after addition or administration of a test drug, the drug can be selected as a therapeutic drug for improving the prognosis of tumor.
The invention also provides a medicament for treating gastric adenocarcinoma metastasis, which comprises an agonist of LRRTM 1.
The activator of LRRTM1 gene or LRRTM1 protein of the present invention is not limited as long as it is a drug that can promote or enhance the expression or activity of LRRTM1 or a substance involved in the upstream or downstream pathway of LRRTM1 and is effective for treating tumors.
The activator also includes a vector or host cell comprising a vector carrying the LRRTM1 gene.
The activator can be used for supplementing the deletion or deficiency of endogenous LRRTM1 protein and treating gastric adenocarcinoma caused by LRRTM1 protein deficiency by improving the expression of LRRTM1 protein. On the other hand, the protein can be used for enhancing the activity of LRRTM1 protein, thereby treating gastric adenocarcinoma.
The invention also provides application of the agonist in preparing a medicament for treating gastric adenocarcinoma metastasis.
The agonists of the invention may be used by formulating pharmaceutical compositions by any means known in the art. Such compositions comprise the active ingredient in admixture with one or more pharmaceutically acceptable carriers, diluents, fillers, binders and other excipients, depending on the mode of administration and the dosage form envisaged. Therapeutically inert inorganic or organic carriers known to those skilled in the art include, but are not limited to, lactose, corn starch or derivatives thereof, talc, vegetable oils, waxes, fats, polyols such as polyethylene glycol, water, sucrose, ethanol, glycerol and the like, various preservatives, lubricants, dispersants, flavoring agents. Moisturizers, antioxidants, sweeteners, colorants, stabilizers, salts, buffers and the like may also be added as needed to aid in the stability of the formulation or to aid in the enhancement of the activity or its bioavailability or to produce an acceptable mouthfeel or odor upon oral administration, formulations which may be used in such compositions may be in the form of their original compounds as such, or optionally in the form of their pharmaceutically acceptable salts, and the agonists of the invention may be administered alone, or in various combinations, as well as in combination with other therapeutic agents. The compositions so formulated may be administered in any suitable manner known to those skilled in the art, as desired. In using the pharmaceutical compositions, a safe and effective amount of an inhibitor of the present invention is administered to a human, wherein the safe and effective amount is typically at least about 100 micrograms per kilogram of body weight for oral administration. Of course, the particular dosage will depend upon such factors as the route of administration, the health of the patient, and the like, and is within the skill of the skilled practitioner.
The medicine of the present invention may be prepared into various preparation forms. Including, but not limited to, tablets, solutions, granules, patches, ointments, capsules, aerosols or suppositories for transdermal, mucosal, nasal, buccal, sublingual or oral use.
The route of administration of the drug of the present invention is not limited as long as it exerts the desired therapeutic or prophylactic effect, and includes, but is not limited to, intravenous, intraperitoneal, intraocular, intraarterial, intrapulmonary, oral, intravesicular, intramuscular, intratracheal, subcutaneous, transdermal, transpleural, topical, inhalation, transmucosal, dermal, gastrointestinal, intraarticular, intraventricular, rectal, vaginal, intracranial, intraurethral, intrahepatic, intratumoral. In some cases, the administration may be systemic. In some cases topical administration.
The dose of the drug of the present invention is not limited as long as the desired therapeutic effect or prophylactic effect is obtained, and can be appropriately determined depending on the symptoms, sex, age, and the like. The dose of the therapeutic agent or prophylactic agent of the present invention can be determined using, for example, the therapeutic effect or prophylactic effect on a disease as an index.
The "LRRTM 1 gene" (NC _000002.12(80301878..80304738) sequences of the present invention can be queried in the NCBI database.
In the context of the present invention, "diagnosis of gastric adenocarcinoma metastasis" includes determining whether a subject has developed gastric adenocarcinoma metastasis, determining whether a subject is at risk for gastric adenocarcinoma metastasis, and determining whether a patient with gastric adenocarcinoma metastasis has relapsed metastasis.
As used herein, "treatment" encompasses treatment-related diseases or disease states in a mammal, such as a human, having the associated disease or disorder, and includes:
(1) preventing the occurrence of a disease or condition in a mammal, particularly when the mammal is susceptible to said disease condition but has not been diagnosed as having such a disease condition;
(2) inhibiting a disease or disease state, i.e., preventing its occurrence; or
(3) Alleviating the disease or condition, i.e., causing regression of the disease or condition.
The term "treatment" generally refers to the treatment of a human or animal (e.g., as applied by a veterinarian) wherein some desired therapeutic effect is achieved, e.g., inhibiting the progression of a condition (including slowing the progression, stopping the progression), ameliorating the condition, and curing the condition. Treatment as a prophylactic measure (e.g., prophylaxis) is also included. The use of a patient who has not yet developed a condition but who is at risk of developing the condition is also encompassed by the term "treatment".
The invention has the advantages and beneficial effects that:
the invention discovers and confirms the close correlation between LRRTM1 gene expression and gastric adenocarcinoma tissue metastasis for the first time, and has a large number of verified samples and accurate results. The proposal of the correlation provides a new way for diagnosing and treating the metastasis of the gastric adenocarcinoma.
The invention develops a reagent or a kit suitable for gastric adenocarcinoma metastasis detection, and the detection sensitivity is good.
Example 1 differential expression of LRRTM1 Gene
1. Sample collection
50 cases of metastatic gastric adenocarcinoma tissues and paracancerous normal tissues thereof are collected, and 45 cases of non-metastatic gastric adenocarcinoma tissues and paracancerous normal tissues thereof are collected. Tissue samples were sampled with the assistance of a pathologist, and the sample collection criteria were as follows:
(1) primary gastric adenocarcinoma without other diseases, and patients do not undergo radiotherapy and chemotherapy before operation; (2) the sample is diagnosed as gastric adenocarcinoma by a pathology department, and tumor cells are not detected in paracancerous normal tissues; (3) in order to avoid unnecessary cross contamination, two sets of equipment are adopted during sampling, a paracancer normal tissue sample which is separated from cancer tissues by at least 5cm and has no obvious lesion is observed by naked eyes, then a full-layer gastric adenocarcinoma tissue sample is taken, the sample is immediately subpackaged and put into an EP tube added with 1ml of RNAlater after being collected, the temperature is kept overnight at 4 ℃, and then the sample is placed in a refrigerator at 70 ℃ below zero for long-term storage. And numbering and labeling on each EP tube. And simultaneously making remarks of related information on the sample record book.
2. RNA (ribonucleic acid) for extracting paired gastric adenocarcinoma and tissue sample beside gastric adenocarcinoma
RNA extraction kit (Trans Zol) from Beijing Quanji Biotech Ltd is usedTMUp PlusRNA Kit) tissue sample RNA was extracted. The method comprises the following specific steps:
(1) weighing the ultra-low temperature frozen sample, quickly transferring to a mortar precooled by liquid nitrogen, sufficiently grinding with a pestle until the sample is ground into powder, transferring the sample ground into powder into a centrifuge tube, and adding 1ml of Trans into every 50-100mg of the sampleZolTMHomogenizing Up with homogenizer, or repeatedly blowing and sucking with gun. The mixture was allowed to stand at room temperature for 5 minutes.
(2) 1ml of Trans Zol is usedTMUp, 0.2ml of chloroform was added thereto, followed by vigorous shaking for 30 seconds and incubation at room temperature for 3 minutes.
(3) Centrifugation at 10000 Xg for 15 minutes at 4 ℃. The sample now separated into three layers, a colorless aqueous phase (upper layer), an intermediate layer, and a pink organic phase (lower layer). RNA was in the colorless aqueous phase and 500. mu.l of the colorless aqueous layer was pipetted.
(4) The aspirated 500. mu.l of colorless aqueous phase was transferred to a new centrifuge tube, 500. mu.l of absolute ethanol was added, and the mixture was gently inverted and mixed. (centrifugation can be carried out at room temperature thereafter)
(5) Mu.l of the resulting solution was added to the column together with the pellet, centrifuged at 12000 Xg for 30 seconds at room temperature, and the effluent discarded (which could be done in several portions if the volume was larger than the column capacity).
(6) Add 500. mu.l of CB9, centrifuge at 12000 Xg at RT for 30 seconds and discard the effluent.
(7) Repeating the step (6) once.
(8) Adding 500 μ l WB9 (checking whether absolute ethanol is added before use), centrifuging at 12000 Xg for 30 s at room temperature, and discarding the effluent
(9) Repeating the step (8) once.
(10) Centrifuging at 12000 Xg room temperature for 2min, completely removing residual ethanol, standing at room temperature for several min, and completely air drying the column.
(11) The column was placed in an RNase-free Tube (kit prepared), and 30. mu.l of RNase-free Water was added to the center of the column, and the column was allowed to stand at room temperature for 1 minute.
(12) The RNA was eluted by centrifugation at 12000 Xg for 1 minute at room temperature.
(13) The RNA was stored in a freezer at-80 ℃.
3. Determination of concentration and purity of extracted RNA
RNA concentration and purity were determined using an ND-1000 instrument from Nano Drop, USA.
Using an ultraviolet spectrophotometer Nano Drop 1000 to double click a Nano Drop icon on a computer screen and entering a system menu; after selecting a nucleic acid measurement option, adding 2 mu l of double distilled water into the sample adding hole according to system prompt; clicking to determine to initialize the system; add 2. mu.l of double distilled water to the wells, select RNA items, and click Blank to measure system background. And (4) wiping the double distilled water with a piece of rear lens wiping paper, then dropping the RNA sample to be detected, and clicking Measure. Reading the value and recording the RNA concentration (ng/. mu.l), wherein the care of the A value 260/280 is necessary, and if the value is between 1.8 and 2.0, the RNA sample quality is better. The RNA concentration of each sample was labeled completely and then frozen in a freezer at-70 ℃.
4. Extracted RNA integrity assay
Agarose gel detection of RNA samples the procedure:
1) electrophoresis tank, cleaning of glue making tool: cleaning with detergent (generally soaking overnight), washing with water, and washing with 3% H2O2Filling the electrophoresis tank, standing at room temperature for 10min, washing with 0.1% (V/V) DEPC water, and air drying for use.
2) Preparing glue: 0.5g of agarose powder was weighed into an Erlenmeyer flask containing 45ml of DEPC water, and heated to completely dissolve the agarose. After a little cooling, 5ml of 10 XTAE electrophoresis buffer and a final concentration of 0.5. mu.g/ml ethidium bromide were added. Then pouring gel into the gel groove, inserting a comb, horizontally placing for use after solidification.
3) Sample adding: the sample is mixed with 6 Xelectrophoresis buffer solution and loaded into the gel sample application hole.
4) Electrophoresis: and opening the electrophoresis apparatus, and stabilizing the voltage of 80V electrophoresis.
5) After electrophoresis (about 30 minutes), the gel was stored by UV light and photographed with a gel imaging system.
Integrity determination by agarose gel electrophoresis detection, if can clearly see 28s rRNA, 18s rRNA, 5srRNA three bands, and 28s rRNA brightness should be 18s rRNA two times. The completeness of the extracted total RNA is better, and the RNA quality meets the requirement.
5. Design and preparation of primers
The primer sequence for detecting LRRTM1 gene expression by qRT-PCR and the primer sequence for amplifying internal reference GAPDH by qRT-PCR are designed and synthesized by biological engineering (Shanghai) corporation, and the sequence information is recorded into NCBI software design primer by searching UCSC database, and is confirmed to be correct in blast in the gene bank.
LRRTM1 gene primers:
an upstream primer: 5'-TCTGCCCAGGAATACTAC-3' (SEQ ID NO. 1);
downstream primer 5'-AGCCATACTCGTTGATGA-3' (SEQ ID NO.2),
GAPDH gene primers:
an upstream primer: 5'-GGGAGCCAAAAGGGTCA-3' (SEQ ID NO. 3);
a downstream primer: 5'-GAGTCCTTCCACGATACCAA-3' (SEQ ID NO. 4).
6. Real-time fluorescent quantitative cDNA reverse transcription detection step
Mu.g of total RNA was reverse transcribed with reverse transcription buffer to synthesize cDNA. A25-mu-l reaction system is adopted, 1 mu g of total RNA is taken from each sample as template RNA, and the following components are respectively added into a PCR tube: DEPC water, 5 Xreverse transcription buffer, 10mmol/L dNTP, 0.1mmol/L DTT, 30. mu. mmol/L Oligo dT, 200U/. mu. L M-MLV, template RNA. Incubate at 42 ℃ for 1h, 72 ℃ for 10min, and centrifuge briefly.
7、PCR
(1) A Bio-RAD real-time fluorescence quantitative PCR instrument was used to prepare a reaction system as shown in Table 1.
TABLE 1 PCR reaction System
(2) The following qPCR reaction parameters were used: pre-denaturation at 95 deg.C for 10min, then denaturation at 95 deg.C for 15s, annealing at 57 deg.C, and extension for 1min for 40 cycles; then, the fluorescence signal was collected and the product dissolution curve was prepared, and 3 replicates of each sample were averaged. By using 2-△△CtThe expression level of LRRTM1 was analyzed by relative quantification method, and Ct is the intensity value of fluorescence signal detected by thermal cycler.The calculation method comprises the following steps: delta Ct ═ (Ct target gene-Ct reference gene) tumor tissue experimental group- (Ct target gene-Ct reference gene) normal tissue control group, 2-△△CtThe expression of the target gene in the experimental group is shown as the fold change relative to the control group, and the analysis of the experimental data is performed by the Bio-RAD analysis software.
5. Statistical analysis statistical software SPSS19.0 is used for data analysis, and a paired T test is used for judging whether the expression of LRRTM1 in gastric adenocarcinoma tissue and a tissue sample beside cancer has a difference of statistical significance, wherein the difference is that P is less than 0.05, and the difference has statistical significance.
6. Results
As shown in FIG. 1, in the metastatic group, the expression level of mRNA of LRRTM1 gene was reduced compared with that of the para-carcinoma tissue, and the difference was statistically significant; in the non-metastatic group, compared with the para-carcinoma tissue, the mRNA expression level of the LRRTM1 gene is not changed statistically in the stomach adenocarcinoma tissue, and the same mRNA expression level of the LRRTM1 gene is detected in the para-carcinoma tissue of the metastatic group and the non-metastatic group. The above results indicate that the low expression of LRRTM1 gene is involved in metastasis of gastric adenocarcinoma tissue.