Exosome modified by Anti-HER2 and carrying STIL siRNA, preparation method and application

Abstract

The invention discloses an Anti-HER2 modified exosome carrying STIL siRNA, a preparation method and application, and belongs to the field of cell biology, and the exosome is prepared from the Anti-HER2 modified exosome and STIL siRNA molecules. The exosome can specifically target and inhibit proliferation and pulmonary metastasis of the gastric cancer, and has important scientific and clinical significance in treatment of the gastric cancer.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of cell biology, in particular to an Anti-HER2 modified and carrying STIL siRNA exosome, preparation method and application. BACKGROUND

[0002] Gastric cancer is insidious, and early symptoms are not obvious. Most patients have reached an advanced stage when they seek medical treatment, with peritoneal, liver, lung and other metastases, losing the opportunity for surgery. Chemotherapy combined with targeted therapy and immunotherapy is the main treatment for metastatic gastric cancer or locally advanced gastric cancer. However, due to off-targeting, drug resistance and other factors, its application in the treatment of gastric cancer is affected. In addition, due to the lack of potential targets, the number of patients who benefit from targeted therapy is small. In view of the above factors, the prognosis of gastric cancer is poor, and the 5-year survival rate is only 33.3%. Therefore, exploring the pathogenesis of gastric cancer and finding new targets are of great significance for the prevention and treatment of gastric cancer and improving the prognosis of patients.

[0003] Finding the key genes driving gastric cancer and designing drugs based on them is an important strategy to improve the efficacy of gastric cancer and improve the prognosis. SCL / TAL1 interrupting locus (STIL) is a key gene that promotes mitosis and cell cycle progression, and is related to the formation of primary cilium and spindle, and is a key gene affecting embryonic development. STIL knockout mice lack centrioles and primary cilia, leading to embryonic development stopping at the middle stage and death. Studies have shown that STIL is a cancer gene and is highly expressed in various cancer tissues, and is closely related to the occurrence and development of tumors. High expression of STIL is associated with poor prognosis in bladder cancer patients. STIL inhibits primary flagella formation through AURKA, activates the SHH signaling pathway, and promotes cell proliferation; STIL activates the PI3K / AKT / mTOR pathway to promote c-myc expression and induce bladder cancer. STIL is also related to the proliferation and invasion of osteosarcoma. After interfering with the expression of STIL, the proliferation, migration and invasion of osteosarcoma cells are reduced, and apoptosis is increased. Kasai K et al. found that STIL was significantly increased in pancreatic intraepithelial neoplasia (PanIN) and pancreatic ductal adenocarcinoma (PDA). Moreover, RAS promotes the binding of STIL to the suppressor of fusion (SUFU) and inhibits its activity, and SUFU affects the activity and nuclear localization of transcription factor GLI1, while GLI1 regulates the proliferation of stem cells. Therefore, STIL binds to SUFU, releases the inhibition of SUFU on transcription factor GLI1, GLI1 enters the nucleus, and promotes the proliferation of pancreatic cancer cells.

[0004] The research group identified the gene STIL that induces the proliferation of gastric cancer cells in vivo and in vitro. STIL is highly expressed in gastric cancer tissues, and knockdown of STIL inhibits the proliferation of gastric cancer cells, induces G2 / M arrest and apoptosis, and further inhibits the growth of gastric cancer. Moreover, STIL promotes the occurrence and development of gastric cancer by activating the PI3K / AKT pathway, and may be a new target for intervention in gastric cancer. Therefore, targeted inhibition of STIL expression is expected to be used for the treatment of gastric cancer. In addition to the oncogene property, STIL also has important biological functions, such as acting as a key factor for promoting mitosis and cell cycle progression, and is also expressed in small amounts in normal tissue cells. How to accurately target drug use and avoid the "off-target" effect of involving normal cells is a problem that needs to be solved.

[0005] Exosomes are extracellular vesicles with a diameter of about 40-150 nm, and their contents include important active molecules such as nucleic acids, proteins and lipids. They are released from tissue cells, mediate intercellular information transmission, and participate in various physiological and pathological processes. Therefore, exosomes can be used as effective drug carriers. As a nanoscale carrier, the exosome membrane is similar to the cell membrane and has a phospholipid bilayer, which can protect the contents from being degraded by enzymes in the blood; in addition, exosomes have the characteristics of low immunogenicity, high biocompatibility, good stability and easy availability. Compared with viral carriers, exosome-carrying genes are safer; compared with non-viral carriers, exosomes can release contents into the cytoplasm through membrane fusion, avoiding the endocytosis of nanocarriers into the cytoplasm, avoiding the acidification and destruction of the contents, and improving the transfection efficiency. At present, exosomes have been used as drug carriers for the treatment of various diseases including tumors, cardiovascular and cerebrovascular diseases, and have broad application prospects. However, there is no report on exosomes targeting HER2 and carrying siRNA STIL for the treatment of gastric cancer. SUMMARY

[0006] The purpose of the present application is to overcome the shortcomings of the prior art and provide an Anti-HER2 modified exosome carrying STIL siRNA, a preparation method and application. The exosome provided by the present application can specifically target and inhibit the proliferation and lung metastasis of gastric cancer cells, and has important scientific and clinical significance in the treatment of gastric cancer.

[0007] To achieve the above-mentioned purpose, the technical scheme adopted by the present application is:

[0008] The present application provides an Anti-HER2 modified exosome carrying STIL siRNA, which is prepared from Anti-HER2 modified exosome and STIL siRNA molecule.

[0009] The present application also provides a preparation method of the above-mentioned exosome, comprising the following steps:

[0010] S1, infect 293T cells with Anti-HER2 lentivirus, collect cell supernatant, centrifugal separation, discard supernatant, resuspend the precipitate to obtain Anti-HER2 modified exosomes;

[0011] S2, mix Anti-HER2 modified exosomes with STIL siRNA molecules, synthesize by electroporation system, wash, centrifugal, resuspend after synthesis to obtain Anti-HER2 modified exosomes carrying STIL siRNA.

[0012] The application also provides a use of the above-mentioned exosomes in the preparation of a drug for promoting tumor cell apoptosis or inhibiting gastric cancer proliferation and lung metastasis.

[0013] Preferably, the exosomes promote or inhibit gastric cancer proliferation and lung metastasis by regulating the expression of apoptosis-related genes.

[0014] Preferably, the apoptosis-related genes are Bcl-2, Bax, Caspase-3 and Caspase-9.

[0015] The application also provides a drug for regulating the expression of apoptosis-related genes, comprising the above-mentioned exosomes and a pharmaceutically or immunologically acceptable carrier or adjuvant.

[0016] The application also provides a drug for promoting tumor cell apoptosis or inhibiting gastric cancer proliferation and lung metastasis, comprising the above-mentioned exosomes and a pharmaceutically or immunologically acceptable carrier or adjuvant.

[0017] Compared with the prior art, the application has the beneficial effects that:

[0018] The Anti-HER2 modified exosomes carrying STIL siRNA provided by the application can be targeted and taken up by gastric cancer cells with high HER2 expression, and can target and deliver drugs to tumor cells with high HER2 expression as a drug carrier, thereby specifically targeting tumor cells, promoting tumor cell apoptosis, and inhibiting the proliferation and lung metastasis of gastric cancer, which has important scientific and clinical significance in the treatment of gastric cancer. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 Figure 1 is a characterization diagram of exosomes (Anti-HER2-Exo); A is a transfection efficiency and Western blot diagram of LAMP2b-Anti-HER2-scFv; B is a transmission electron microscope diagram of Anti-HER2-Exo; C is a particle size analysis diagram of Anti-HER2-Exo; D is a Western blot diagram of the expression levels of exosome markers Alix, CD63 and TSG101 of Anti-HER2-Exo.

[0020] Figure 2 Figure 4 is a graph of the uptake of Anti-HER2-Exo-siSTIL in SGC-7901 cells; A is a fluorescence microscope image of the uptake of exosomes by SGC-7901 cells at different time points; B is a qRT-PCR analysis result graph of STIL gene expression at different time points; C is a Western blot graph of STIL gene at different time points;

[0021] Figure 3 Figure 5 is a graph of the effect of different doses of Anti-HER2-Exo-siSTIL on tumor growth; A is a photo of the nude mice and tumors; B is a result graph of H&E staining for detecting the pathological morphological changes of tumor tissues; C is a graph of the change of body weight of nude mice; D is a graph of the change of tumor volume of nude mice; E is a graph of tumor mass of nude mice;

[0022] Figure 4 Figure 6 is a graph of the effect of Anti-HER2-Exo-siSTIL on tumor cell apoptosis; A is a result graph of RT-qPCR analysis; B is a Western blot graph of protein expression;

[0023] Figure 5 Figure 7 is a graph of the in vivo anti-tumor effect of Anti-HER2-Exo-siSTIL; A is a real-time imaging graph of tumor lung metastasis detection; B is the average fluorescence intensity. DETAILED DESCRIPTION

[0024] The technical solutions in the embodiments of the present application will be described below in conjunction with the drawings of the embodiments of the present application. Figures 1 to 5 It is apparent that the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by a person of ordinary skill in the art without creative labor fall within the protection scope of the present application.

[0025] I. Materials and methods

[0026] 1. Cell culture

[0027] The 293T cells and SGC-7901 gastric cancer cells in the embodiments of the present application were purchased from ATCC. The cells were cultured in DMEM medium (Thermo, USA) containing 10% fetal bovine serum (Gibco, USA) in a constant temperature incubator, and the conditions were as follows: 37℃, 5% carbon dioxide content.

[0028] 2. Animal experiments

[0029] The animal experiments were approved by the Animal Ethics Committee of the People's Hospital of Inner Mongolia Autonomous Region (Approval No.: 202202036K).

[0030] 3. Virus infection of 293T cells

[0031] 293T cells in logarithmic growth phase were diluted to 1 x 10 6 / mL, and 2 x 10 5 cells were inoculated in each well. The next day, the cells were cultured to 60-70% confluence. The experimental groups included:

[0032] Control group, i.e. blank group;

[0033] NC group, i.e. empty virus group;

[0034] LAMP2b-Anti-HER2 group, i.e. virus group carrying LAMP2b-Anti-HER2 plasmid;

[0035] Among them, the empty virus and the virus carrying LAMP2b-Anti-HER2 plasmid were constructed and packaged by Shanghai Jikai Gene Co., Ltd.

[0036] The transfection reagent was operated according to the instructions of Lipofectamine 3000 (Thermo, USA). The old culture medium was removed, and the 293T cells were washed with serum-free culture medium. The solutions of each group were added, incubated for 6-8 h, and then cultured with DMEM medium containing 10% fetal bovine serum for 48 h.

[0037] 4. Extraction of Anti-HER2-Exo

[0038] The culture supernatant of 293T cells after infection with the above LAMP2b-Anti-HER2 lentivirus was collected, centrifuged at 4°C, 100,000g for 90 min, and the supernatant was discarded. The precipitate was resuspended with PBS, washed again, and then resuspended with 200 μL of PBS. The exosome precipitate was stored in a -80°C refrigerator for standby use.

[0039] 5. Transmission electron microscopy and particle size analyzer identification of Anti-HER2-Exo

[0040] 10 μl of Anti-HER2-Exo was dropped on a copper grid, and after standing for 1 min, the excess liquid was absorbed with filter paper. 10 μl of uranyl acetate was added to the copper grid, and after standing for 1 min, the excess liquid was removed with filter paper. After the sample was air-dried at room temperature, transmission electron microscopy imaging was performed to observe the morphology of the exosomes.

[0041] Particle size analysis, 20 μl Anti-HER2-Exo was diluted 5000 times with PBS, the suspension was directly imaged using a particle size analyzer (ZetaView, Particle Metrix) to observe exosomes and small vesicles with a diameter range of 50-1000 nm. The Zetaview software was used to calculate the size distribution of exosomes.

[0042] After Anti-HER2-Exo was lysed using RIPA lysis buffer (Sigma, US), the expression of exosome markers Alix, CD63 and TSG101 was detected by Western blot.

[0043] 6. Preparation of Anti-HER2 modified exosomes carrying STIL siRNA (Anti-HER2-Exo-siSTIL)

[0044] Using the GENE Pulser X cell electroporation system, Anti-HER2-Exo (0.5 mg / ml) was mixed with STIL siRNA (100 nM) and added to the electroporation cup, and then electroporated at 400 V, 125 uF. After washing with PBS, centrifugation was performed at 12,000 g, and the precipitate was resuspended in PBS to obtain Anti-HER2 modified exosomes carrying STIL siRNA.

[0045] 7. SGC-7901 gastric cancer cell uptake Anti-HER2-Exo-siSTIL experiment

[0046] SGC-7901 cells were seeded in a 96-well plate, Anti-HER2-Exo-siSTIL was stained using a Dio kit, and the mixture was added to the SGC-7901 cell-seeded well plate under light shielding conditions (200 μl / well). Then, the 96-well plate was placed in a 5% CO2, 37°C incubator, and incubated in the dark. At 6, 12, 24, and 48 h, the 96-well plate was removed, and the culture supernatant was discarded. 100 μl of 4% paraformaldehyde solution was added to fix the cells, and after incubation at room temperature, the paraformaldehyde solution was discarded. Under light shielding conditions, 50 μl of 10 mg / L DAPI staining solution was added, and after incubation at room temperature for 5 min, the staining solution was discarded, and the cells were washed with PBS. Finally, 100 μl of PBS was added, and observed under a fluorescence inverted microscope. Images were taken to record the uptake of exosomes by SGC-7901 cells at different time points.

[0047] 8. Minimum Anti-tumor Dose of Anti-HER2-Exo-siSTIL

[0048] BALB / C nude mice, 4-5 weeks old, weighing 10-15 g, were cultured under the following conditions: 22±2℃, humidity 50-60%, light and dark cycle every 12 h. Logarithmic growth phase SGC-7901 cells (1×10 6 / mL) were collected and subcutaneously injected into the left and right dorsal regions of the nude mice (200 μl per mouse) to establish a xenograft tumor model. The mice were divided into three groups (3 mice per group) according to the dose: PBS group (Model group), low-dose group (low dose group, 1.5×1011 particles / mL), and high-dose group (high dose group, 3×1011 particles / mL). The low-dose and high-dose groups were injected intratumorally with exosomes (Anti-HER2-Exo-siSTIL) once every 3 days, for a total of 4 injections. During this period, the body weight, tumor volume (V=ab 2 / 2, where a is the long diameter and b is the short diameter), and tumor weight of the animals were monitored.

[0049] 9. Anti-HER2-Exo-siSTIL in vivo anti-metastasis effect

[0050] BALB / C nude mice, 4-5 weeks old, weighing 10-15 g, were cultured under the following conditions: 22±2℃, humidity 50-60%, light and dark cycle every 12 h. SGC-7901 cells carrying luciferase were injected into the tail vein to establish a lung metastasis tumor model, and a small animal in vivo imaging system was used for imaging. After successful modeling, the nude mice were randomly divided into four groups (3 mice per group): Anti-HER2-Exo-siSTIL group, Exo-siSTIL group, Anti-HER-Exo group, and PBS group. The mice were administered by tail vein injection twice a week for 3 weeks.

[0051] 10. Small animal imaging

[0052] The nude mice were injected intraperitoneally with D-Luciferin at a concentration of 0.15 g / L. After 15 min, the nude mice were placed in the anesthesia chamber of an animal gas anesthesia machine and induced anesthesia with isoflurane. Subsequently, in vivo imaging was performed, and the photon count was collected and analyzed using Lumina II LivingImage 4.0 software.

[0053] 11. H&E staining

[0054] The tumor tissue was fixed with 10% formalin for 24 h, dehydrated with ethanol gradient, cleaned with xylene, and embedded with paraffin. 2 μl thick sections were prepared. After deparaffinization and hydration of the sections, they were stained with hematoxylin for 40 s, rinsed with tap water, and then stained with eosin for 3 s. After dehydration with ethanol gradient, the sections were mounted with neutral resin and observed under an optical microscope.

[0055] 12. RT-qPCR

[0056] RNA was extracted from tissues, cells or exosomes using an RNA extraction kit (Tiangen, China), and then reverse-transcribed into cDNA for real-time quantitative PCR detection, with GAPDH as an internal reference. The RT-qPCR primer sequences are shown in Table 1 below.

[0057] Table 1 Gene primer sequences

[0058]

[0059] 13、Westernblot

[0060] Proteins were extracted from tissues, cells or exosomes using RIPA lysis buffer (Thermo, USA), and the protein concentration was determined using a BCA kit. 20 μg of sample was loaded onto a 10% SDS-PAGE for electrophoresis, and after 2 hours of constant voltage transfer, 5% skim milk was blocked at room temperature for 1 hour. The PVDF membrane was incubated with the antibody at 4°C overnight. The antibodies included LAMP2 (1:1000), Alix (1:1000), CD63 (1:1000), TSG101 (1:1000), Bcl-2 (1:1000), Bax (1:1000), Caspase-3 (1:1000), Caspase-9 (1:1000), STIL (1:1000), and GAPDH (1:4000). After TBST washing, the PVDF membrane was incubated with HRP-labeled secondary antibody at room temperature for 1 hour. ECL chemiluminescence solution was added dropwise, and exposed and developed.

[0061] II. Experimental results

[0062] 1. Exosome characterization

[0063] The present application uses lentiviral recombinant plasmid to infect 293T cells. Figure 1 A Western bolt result shows that the expression level of LAMP2 in the LAMP2b-Anti-HER2 group is increased (p<0.05), indicating successful transfection. Then, using ultracentrifugation method, Anti-HER2 modified exosomes (Anti-HER2-Exo) were extracted and purified from the culture supernatant of LAMP2b-Anti HER2-scFv infected 293T cells. Figure 1 B shows that Anti-HER2-Exo is a spherical vesicle with a membrane structure. Figure 1 C Particle size analysis results show that the diameter of Anti-HER2-Exo is 128.74±27.24 nm, which is consistent with the typical exosome size of 50-150 nm. Figure 1DWestern blot results showed that the expression levels of exosome markers Alix, CD63, and TSG101 were high. These results indicated that the ultracentrifugation method could successfully extract Anti-HER2-Exo.

[0064] 2. Relationship between Anti-HER2-Exo-siSTIL uptake efficiency and incubation time of SGC-790 cells

[0065] The present application uses the GENE Pulser X cell electroporation system to mix STIL siRNA with Anti-HER2-Exo to prepare Anti-HER2 modified and STIL siRNA carrying exosomes. DiO labeled Anti-HER2-Exo-siSTIL is co-incubated with SGC-7901 cells, and fluorescence images are taken at 6h, 12h, 24h, and 48h to record cell uptake. Figure 2 A immunofluorescence results show that SGC-7901 cells effectively internalize DiO labeled Anti-HER2-Exo-siSTIL, and the amount of uptake increases with time. Figure 2 B qRT-PCR results show that STIL expression is significantly reduced (P<0.01), and this result is further confirmed by Western blot detection (P<0.01), as shown in Figure 2 C. These findings indicate that Anti-HER2-Exo-siSTIL successfully enters SGC-7901 cells and inhibits STIL expression.

[0066] 3. Determination of the optimal concentration of Anti-HER2-Exo-siSTIL

[0067] To explore the optimal anti-tumor concentration of Anti-HER2-Exo-siSTIL, the present application inoculates SGC-7901 cells subcutaneously in nude mice to construct a xenograft tumor model. The low and high dose groups are injected with 1.5×10 11 particles / mL and 3×1011 particles / mL of exosomes, respectively. Figure 3 A in vitro imaging results show that compared with the model group, both the high and low doses of Anti-HER2-Exo-siSTIL can inhibit the growth of tumors in nude mice, with the high dose group being more effective. Figure 3 B H&E staining results show that tumor cells in the model group are tightly packed, with large and darkly stained nuclei, while in the exosome injection groups, especially in the high dose group, tumor cell structure is destroyed, with necrosis and fibrosis, indicating that Anti-HER2-Exo-siSTIL can affect tumor cell metabolism. Figure 3C shows that the body weight of the three groups of nude mice initially increased and then decreased during the 38-day treatment process, and the body weight of the model group and the high-dose group was always heavier than that of the low-dose group. Figure 3 D shows that Anti-HER2-Exo-siSTIL significantly inhibited the growth of tumor volume compared with the model group, and the high-dose group had better tumor inhibition effect (P < 0.001). Figure 3 E shows that the tumor weight of the Anti-HER2-Exo-siSTIL treatment group was significantly reduced compared with the model group, especially in the high-dose group (P < 0.05). These results further demonstrate the effectiveness of Anti-HER2-Exo-siSTIL in inhibiting tumor growth, and the high-dose group shows better anti-tumor effect.

[0068] 4. Anti-HER2-Exo-siSTIL promotes tumor cell apoptosis

[0069] To explore the effect of Anti-HER2-Exo-siSTIL on tumor cell apoptosis, the inventors detected the expression of Bcl-2, Bax, Caspase-3, and Caspase-9 using RT-qPCR and Western blot. The results of RT-qPCR showed that the mRNA level of Bcl-2 was significantly reduced, while the expression levels of Bax, Caspase-3, and Caspase-9 were significantly increased in the exosome treatment group, especially in the high-dose group, as shown in Figure 4 A. Western blot also confirmed this result at the protein level, as shown in Figure 4 B. Therefore, Anti-HER2-Exo-siSTIL induces cell apoptosis by reducing the expression of STIL and anti-apoptotic protein Bcl-2, and increasing the expression of pro-apoptotic proteins Bax, Caspase-3, and Caspase-9, and this effect is more pronounced in the high-dose group. The above results show that Anti-HER2-Exo-siSTIL can specifically target tumor cells, promote apoptosis, and inhibit tumor growth, and the high-dose group is superior to the low-dose group. Therefore, the inventors selected the high-dose (3 × 10^11 particles / mL) for subsequent experiments.

[0070] 5. Anti-HER2-Exo-siSTIL specifically inhibits lung metastasis of SGC-7901 cells

[0071] To study the in vivo anti-tumor effect of Anti-HER2-Exo-siSTIL, the inventors established a lung metastasis tumor model in nude mice by tail vein injection of luciferase-labeled SGC-7901 cells, and observed tumor lung metastasis using live imaging. Figure 5It is shown that the number of lung metastases in the Anti-HER2-Exo-siSTIL group, the Exo-siSTIL group and the Anti-HER-Exo-siCtrl group is significantly reduced compared with the PBS group, and the Anti-HER2-Exo-siSTIL group has the most obvious tumor inhibition effect (P<0.05). The above results show that the Anti-HER2-Exo-siSTIL can specifically inhibit the lung metastasis of SGC-7901 cells.

[0072] In summary, the exosome (Anti-HER2-Exo-siSTIL) provided by the application can inhibit the growth of SGC-7901 cell xenograft tumor, destroy tumor tissue and increase cell apoptosis; the exosome provided by the application can inhibit the expression of the Bcl-2 gene of SGC-7901 cells and promote the expression of the Bax, Caspase-3 and Caspase-9 genes, that is, the Anti-HER2-Exo-siSTIL can promote the apoptosis of tumor cells by regulating the expression of apoptosis-related genes.

[0073] Although the embodiments of the present application have been shown and described, those of ordinary skill in the art can understand that various changes, modifications, replacements and variations can be made to these embodiments without departing from the principles and purposes of the present application, and the scope of the present application is defined by the claims and their equivalents.

Claims

1. An exosome modified with Anti-HER2 and carrying STIL siRNA, characterized in that, The exosomes were prepared from Anti-HER2 modified exosomes and STIL siRNA molecules.

2. A method for preparing exosomes according to claim 1, characterized in that, Includes the following steps: S1. Infect 293T cells with Anti-HER2 lentivirus, collect cell culture supernatant, centrifuge, discard supernatant, resuspend precipitate to obtain Anti-HER2 modified exosomes; S2. The obtained Anti-HER2 modified exosomes were mixed with STIL siRNA molecules and synthesized using an electroporation system. After synthesis, the exosomes were washed, centrifuged, and resuspended to obtain Anti-HER2 modified exosomes carrying STIL siRNA.

3. The use of the exosomes according to claim 1 in the preparation of a drug that promotes tumor cell apoptosis or inhibits gastric cancer proliferation and lung metastasis.

4. The application according to claim 3, characterized in that, Exosomes promote tumor cell apoptosis by regulating the expression of apoptosis-related genes, thereby inhibiting gastric cancer proliferation and lung metastasis.

5. The application according to claim 3, characterized in that, Apoptosis-related genes are Bcl-2, Bax, Caspase-3, and Caspase-9.

6. A drug for regulating the expression of apoptosis-related genes, characterized in that, It includes the exosomes as described in claim 1, as well as pharmaceutically or immunologically binding carriers or excipients.

7. A drug that promotes tumor cell apoptosis or inhibits gastric cancer proliferation and lung metastasis, characterized in that, It includes the exosomes as described in claim 1, as well as pharmaceutically or immunologically binding carriers or excipients.

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