Application of RG108 in heart transplantation immunosuppression by regulating T cell function

Abstract

The invention discloses application of RG108 in heart transplantation immunosuppression by regulating T cell functions, and relates to the technical field of medical bioengineering. The invention innovatively provides a method for inhibiting the activity of DNA methyltransferase 1 (DNMT1) by using a small molecule compound RG108, and by reducing the methylation level of a CDKN1A gene promoter, restoring the expression of the promoter and inducing T cells to enter an aging state, the proliferation and effect activity of the promoter is reduced, and rejection reaction is inhibited. The RG108 is a non-nucleoside DNMT1 selective inhibitor with a clear structure, the activity of the inhibitor is inhibited by combining with a catalytic structural domain, DNA is not doped, off-target toxicity is not caused, and the RG108 is an ideal molecule for realizing controllable and reversible epigenetic regulation. A new transplantation immune tolerance strategy is established, and the defects that an existing immunosuppressor is large in side effect and cannot induce immune memory tolerance are overcome.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of medical bioengineering, and particularly relates to application of RG108 in heart transplantation immunosuppression by regulating T cell function. BACKGROUND

[0002] Heart failure is a terminal cardiovascular disease with extremely high morbidity and mortality worldwide. According to the data released by the International Society for Heart and Lung Transplantation (ISHLT), heart transplantation is one of the most effective surgical methods for treating end-stage heart failure, with more than 5000 heart transplant surgeries performed worldwide each year, but the 10-year survival rate after surgery is still less than 60%. Immune rejection after transplantation is one of the main obstacles to long-term survival of the graft.

[0003] Current immunosuppressive therapy mainly uses calcineurin inhibitors (cyclosporine, tacrolimus), anti-proliferative drugs (mycophenolate mofetil), corticosteroids, and mTOR inhibitors (sirolimus). Although these drugs effectively prolong graft survival, long-term use still faces adverse reactions such as glucose and lipid metabolism disorders, nephrotoxicity, bone marrow suppression, and increased risk of infection. Therefore, finding new immune regulation targets and strategies has become a key problem in the field of transplantation immunology.

[0004] In recent years, epigenetics, especially DNA methylation modification, has been increasingly studied in the regulation of T lymphocyte function. DNA methyltransferase 1 (DNMT1), as the main enzyme for maintaining methylation, plays a core role in T cell activation, differentiation, and function. Studies have shown that DNMT1 is abnormally expressed in T cells of autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis, and affects T cell function and aging by regulating cycle regulators such as CDKN1A. However, in the field of organ transplantation immunology, DNMT1 as a target for regulating rejection has not been studied.

[0005] RG108 is a non-nucleoside small molecule DNMT1 inhibitor that can selectively inhibit its catalytic domain without DNA incorporation, with high safety. Studies have reported that RG108 has good epigenetic regulation in tumors and neurodegenerative diseases, but its application in transplantation immunoregulation, T cell aging induction, and anti-rejection therapy has not been reported.

[0006] Therefore, the main deficiencies of the current technology include: (1) lack of new anti-rejection drugs with novel mechanism of action and high safety; (2) there is no mature strategy for inducing T cell aging to achieve transplantation immunological tolerance; (3) existing DNMT1 inhibitors such as 5-Azac have integration toxicity, which is not conducive to clinical translation; (4) although RG108 is a high-quality epigenetic regulation molecule, its application in organ transplantation has not been established. SUMMARY

[0007] To solve the above problems in the prior art, the application aims to provide a T cell aging induction strategy based on DNMT1 inhibitor RG108 for reducing immune rejection after heart transplantation and prolonging the survival time of the graft. The application regulates the functional state of T cells through epigenetic regulation, especially induces the transformation of T cells to the aging phenotype, establishes a new transplantation immune tolerance strategy, and overcomes the defects of the existing immunosuppressive agents, such as large side effects and inability to induce immune memory tolerance.

[0008] To achieve the above-mentioned purpose, the application adopts the following technical solutions:

[0009] The application provides an application of RG108 in the preparation of an organ transplantation immunosuppressive drug, and the chemical structure of the RG108 is as follows:

[0010] .

[0011] Further, the organ includes a heart.

[0012] Further, the target of the drug includes T cells.

[0013] Further, the drug plays an organ transplantation immunosuppressive role by inhibiting DNMT1, reducing CDKN1A promoter methylation, up-regulating CDKN1A expression, blocking T cell cycle, and activating the aging phenotype.

[0014] Further, the dose range of the RG108 is 2.5-10 mg / kg / day.

[0015] Further, the optimal dose of the RG108 is 5 mg / kg / day.

[0016] Further, the RG108 is used once a day.

[0017] Further, the drug further comprises one or more pharmaceutically acceptable carriers and / or excipients.

[0018] Further, the drug can further comprise other immunosuppressive agents.

[0019] Further, the other immunosuppressive agents include CTLA4-Ig.

[0020] Compared with the prior art, the application has the following beneficial effects:

[0021] The application realizes effective control of transplant rejection by using the non-nucleoside small molecule DNMT1 inhibitor RG108 to intervene in heart transplant recipients, and has the following remarkable beneficial effects:

[0022] 1. Novel action mechanism, breaking through the traditional immunosuppression path

[0023] Unlike the traditional strategy of relying on inhibiting TCR / CD28 signals or immune checkpoints (such as CTLA4 and PD-1), the application proposes to induce T cell aging through DNMT1 inhibition to achieve immunoregulation. By inhibiting the activity of DNA methyltransferase 1 (DNMT1), the application reduces the methylation level of the CDKN1A promoter and promotes T cells to enter an irreversible cycle arrest state. This strategy can substantially reshape T cell fate, making it in a "immunosenescence" state with low reactivity and low proliferation ability. This mechanism is different from the passive blocking of traditional signal pathway inhibition, but a new way of actively inducing immune tolerance, providing a new treatment idea for the field of organ transplantation.

[0024] 2. Clear epigenetic target, more precise regulation, low off-target risk

[0025] RG108 directly binds to the catalytic domain of DNMT1 without the need for DNA incorporation, enabling selective demethylation and avoiding the toxicity and adverse reactions caused by non-specific demethylation of the whole genome. The application proves that RG108 can significantly reduce the methylation level of the CDKN1A promoter, and significantly up-regulate the expression of CDKN1A, promoting the formation of senescent T cells (CD27⁻CD49d⁺). It effectively avoids the problems of broad-spectrum toxicity, high off-target rate and genomic instability in the prior art.

[0026] 3. Significant anti-rejection effect, more durable tissue protection

[0027] Experiments on ectopic heart transplantation models have proved that the heart grafts of RG108-treated mice have significantly prolonged survival time, well-preserved tissue structure, and significantly reduced inflammatory cell infiltration and fibrosis. The survival time of the grafts is significantly prolonged (>25 days), which is significantly better than the CTLA4-Ig intervention group. The application has more significant effects and more durable maintenance time in controlling rejection and prolonging graft survival.

[0028] 4. High safety, low toxicity, good clinical transformation prospects

[0029] The RG108 used in the present application is a non-nucleoside structure, does not cause common side effects such as bone marrow suppression, DNA damage, off-target mutation, etc., does not affect T cell viability in the range of 10-30 μM in in vitro cell experiments, and does not show obvious toxicity when the injection dose is 5 mg / kg in vivo, so that the drug of the present application shows good tolerance and safety window in animal experiments, and has more clinical application potential.

[0030] 5. Strong generalizability, suitable for various organ transplantation or autoimmune models

[0031] The mechanism of the present application does not depend on specific antigen presentation systems or tissue environments, so it can be theoretically applied to liver transplantation, kidney transplantation, skin transplantation and other models, and is also expected to be applicable to epigenetic regulation and intervention of T cell related autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis.

[0032] In summary, the technical scheme of the present application for inducing T cell senescence by RG108 solves the problems of high toxicity, unstable effect, lack of specific regulation ability and the like of the existing immunosuppressants, significantly improves the precision, safety and sustainability of immunosuppression, and has outstanding scientific innovation value and broad clinical application prospect. BRIEF DESCRIPTION OF DRAWINGS

[0033] Figure 1 Figure 4 is the proportion change of CD4⁺CD27⁻CD49d⁺ senescent phenotype cells and the expression of PD-1 and TIM-3 in the RG108 group and the control group in Example 1.

[0034] Figure 2 Figure 6 is the methylation electrophoresis image results of the CDKN1A promoter region in the RG108 group and the control group in Example 2.

[0035] Figure 3 Figure 8 is the survival curve of the graft in the RG108 group, the CTLA4-Ig group and the control group in Example 3 and Comparative Example 1.

[0036] Figure 4 Figure 10 is the HE staining and EVG staining comparison results of the RG108 group and the control group in Example 3.

[0037] Figure 5 Figure 12 is the HE staining and EVG staining comparison results of the CTLA4-Ig group and the RG108 group in Comparative Example 1. DETAILED DESCRIPTION

[0038] The following examples are used to illustrate the present application but not to limit the scope of the present application. Modifications or substitutions to the methods, steps or conditions of the present application, without departing from the spirit and essence of the present application, are within the scope of the present application. The reagents, products and instruments used in the following examples are commercially available, and the methods used in the examples are consistent with the commonly used methods, unless otherwise specified.

[0039] The main purpose of the present application is to provide a T cell senescence induction strategy based on DNMT1 inhibitor RG108 for reducing immune rejection after heart transplantation and prolonging the survival time of the graft. The present application regulates the functional state of T cells through epigenetic regulation, especially induces the transformation of T cells to the senescent phenotype, establishes a new transplantation immune tolerance strategy, and overcomes the defects of existing immunosuppressive agents, such as large side effects and inability to induce immune memory tolerance.

[0040] The technical idea of the present application is as follows:

[0041] The present application innovatively proposes to use small molecule compound RG108 to inhibit the activity of DNA methyltransferase 1 (DNMT1), reduce the methylation level of the CDKN1A gene promoter, restore its expression, induce T cells to enter the senescent state, and thus reduce their proliferation and effector activity, and inhibit rejection.

[0042] RG108 is a non-nucleoside, structurally clear DNMT1 selective inhibitor (chemical name: N-Phthalyl-L-tryptophan), which inhibits its activity by binding to the catalytic domain, does not incorporate DNA, and does not cause off-target toxicity, and is an ideal molecule for realizing controllable and reversible epigenetic regulation.

[0043] The technical points of the present application are as follows:

[0044] ① Core drug: RG108 (MCE), CAS number: 48208-26-0, structural formula as follows:

[0045]

[0046] ② Key target: DNA methyltransferase 1 (DNMT1)

[0047] ③ Mechanism of action: inhibit DNMT1 → reduce CDKN1A promoter methylation → up-regulate CDKN1A expression → T cell cycle arrest → activate senescent phenotype

[0048] ④ Method of administration

[0049] A. Drug administration animal model

[0050] Model type: BALB / c→C57BL / 6 heterotopic heart transplantation model;

[0051] Animal grouping: the recipient C57BL / 6 mice were randomly divided into normal saline group (control), CTLA4-Ig group, RG108 group.

[0052] Operation procedure:

[0053] a. Donor heart acquisition (BALB / c mice, Rikangbao (Wuhan) Biotechnology Co., Ltd.): 8-12 weeks old male BALB / c mice were anesthetized (1% sodium pentobarbital), and the chest was opened under sterile conditions. The aorta and pulmonary artery were ligated and cut, and the heart and ascending aorta and pulmonary artery were taken out and placed in pre-cooled normal saline for standby.

[0054] b. Recipient operation site preparation (C57BL / 6 mice, Rikangbao (Wuhan) Biotechnology Co., Ltd.): The C57BL / 6 recipient mice were also anesthetized, fixed supine, and the abdominal aorta and inferior vena cava were exposed. The abdominal aorta and inferior vena cava were isolated to a length sufficient for anastomosis, and the blood vessels on both sides were temporarily clamped to control bleeding. The anterior wall of the abdominal aorta and inferior vena cava was incised to prepare for anastomosis with the donor heart.

[0055] c. Vascular anastomosis (microsurgery): The donor heart ascending aorta was end-to-side anastomosed with the recipient abdominal aorta, and the donor pulmonary artery was end-to-side anastomosed with the recipient inferior vena cava. The suture was completed under a microscope using 10-0 nylon thread, and no obvious leakage was ensured. The blood vessel clamp was loosened, and it was observed whether the donor heart was beating (≥80% of the donor heart could spontaneously beat within 2-5 minutes after the operation). The heart was gently replaced into the abdominal cavity with a wet gauze, and the peritoneum and skin were sutured.

[0056] d. Postoperative treatment: After the mice recovered, they were placed on a 37°C warming pad and observed for 1 hour. The graft was observed daily after the operation, and RG108 was injected intraperitoneally immediately after the operation.

[0057] B. RG108 administration regimen

[0058] Dose design: RG108 was used in a dose range of 2.5-10 mg / kg / day, and 5 mg / kg / day was used as the optimal dose.

[0059] Administration method: intraperitoneal injection;

[0060] Administration volume: 200 μL of drug solution (dissolved in PBS) was injected into each mouse according to the body weight;

[0061] Administration frequency: once a day;

[0062] Administration time course: starting from the 0th day after transplantation, continuous administration for 7-14 days, simulating the short-term induction regimen in clinic.

[0063] The technical solutions of the present application will be further described in detail in conjunction with the examples.

[0064] Example 1 RG108 induces T cell senescence phenotype Figure 1 )

[0065] (1) Experimental design

[0066] This experiment aims to verify whether the DNMT1 inhibitor RG108 can induce the appearance of senescence phenotype in primary T cells cultured in vitro, thereby constituting the basic mechanism of anti-rejection immune intervention.

[0067] The senescent T cell phenotype is mainly defined as CD4⁺CD27⁻CD49d⁺, accompanied by increased expression of senescence-associated receptors (such as PD-1, TIM-3), and low cell function and cycle arrest.

[0068] The experiment uses RG108 to treat mouse CD4+ T cells, and sets up a physiological saline negative control group to detect the changes in senescence phenotype and the expression of related markers.

[0069] (2) Experimental methods

[0070] ① Cell extraction and purification: Take the spleen of C57BL / 6 mice, use a cell screen to prepare a single cell suspension; use CD4⁺ T cell magnetic bead sorting kit (Miltenyi) to purify; purity is verified by flow cytometry, requiring >95%.

[0071] ② Cell culture and intervention: Seed CD4⁺ T cells in RPMI-1640 complete culture medium (supplemented with 10% FBS, 1% penicillin-streptomycin, 2 mM L-glutamine); at the same time, add anti-CD3 / CD28 antibodies (1 μg / mL each, CD3 (Proteintech, Cat No. 17617-1-AP), CD28 (Proteintech, Cat No. 65099-1-Ig)) to activate T cells;

[0072] ③ Group settings are as follows:

[0073] Control group: physiological saline;

[0074] RG108 intervention group: 30 μM;

[0075] Treatment time for each group: 72 hours.

[0076] ④ Flow cytometry detection of phenotype: collect treated cells, and flow cytometry detect the following indicators: CD4-APC, CD27-PE, CD49d-FITC, PD-1-PerCP, TIM-3-PE-Cy7; use BD FACS Canto II flow cytometer to collect data, and use FlowJo for analysis.

[0077] (3) Experimental results

[0078] ① CD4⁺CD27⁻CD49d⁺ senescent phenotype cell proportion change

[0079] The result data shows that the proportion of CD27⁻CD49d⁺ in the control group T cells is about 5.93%; the RG108 intervention group is increased to 31.6%;

[0080] ② PD-1 and TIM-3 expression

[0081] In the RG108 intervention group, the proportion of PD-1⁺ cells is significantly up-regulated; the proportion of TIM-3⁺ cells is also increased, indicating that T cells are in an exhausted / senescent state;

[0082] (4) Experimental conclusion

[0083] RG108 can effectively induce mouse CD4⁺T cells to transform into CD27⁻CD49d⁺ senescent phenotype; at the same time, up-regulate the expression of senescent / exhausted related receptors PD-1 and TIM-3; it is suggested that RG108 can regulate the immune activity of T cells by inducing them into a senescent state; and provide cell mechanism support for its application in heart transplant anti-rejection immune regulation.

[0084] Example 2 RG108 reduces CDKN1A promoter region methylation Figure 2 )

[0085] (1) Experimental design

[0086] The purpose of this experiment is to verify whether RG108 can reduce the DNA methylation level of the CDKN1A promoter region, thereby activating its transcription and expression. As a selective inhibitor of DNMT1, RG108 specifically inhibits methylation maintenance activity through a non-incorporation mechanism, which may regulate the expression of key cycle control genes without triggering whole genome demethylation side effects, providing an epigenetic basis for inducing T cell senescence.

[0087] The experimental mouse spleen T cells were used as a model, and RG108 was added in vitro for intervention. The MSP (methylation-specific PCR) technique was used to detect the methylation state change of the CDKN1A promoter region.

[0088] (2) Experimental method

[0089] ① Cell culture and treatment: C57BL / 6 mouse spleen was used, and a single cell suspension was prepared using a cell screen. CD4⁺T cell magnetic bead separation kit (Miltenyi) was used for purification. The purity was verified by flow cytometry, and the requirement was >95%. It was routinely cultured in 1640 medium supplemented with 10% FBS and 1% penicillin-streptomycin.

[0090] Group 2: 30 μM RG108, 72 hours;

[0091] Control group: treated with the same volume of normal saline;

[0092] RG108 group: the treatment concentration was 30 μM, and the culture time was 72 hours;

[0093] Three replicate wells were set up in each group.

[0094] ③ DNA extraction and sodium bisulfite treatment: after the culture, the TIANamp Genomic DNA Kit was used to extract the genomic DNA of the cells; the extracted DNA was subjected to sodium bisulfite conversion using the EZ DNA Methylation-Gold Kit, and the methylated cytosine was retained.

[0095] ④ MSP detection (methylation-specific PCR)

[0096] Methylation primers (M) and unmethylation primers (U) for the CDKN1A promoter region were designed:

[0097] CDKN1A (M) primers:

[0098] Forward: 5'-GGTTAGGTTTAGTTGGTTCGGC-3' (SEQ ID NO. 1)

[0099] Reverse: 5'-CCTCAACTAACGCAACTCAACG-3' (SEQ ID NO. 2)

[0100] CDKN1A (U) primers:

[0101] Forward: 5'-TTTGGTTAGGTTTAGTTGGTTTGGT-3' (SEQ ID NO. 3)

[0102] Reverse: 5'-CACACCTCAACTAACACAACTCAACA-3' (SEQ ID NO. 4)

[0103] PCR reaction system: 25 μL, HotStar Taq enzyme was used, and the amplification conditions were strictly in accordance with the MSP standard system;

[0104] The PCR products were detected by 2% agarose gel electrophoresis, and the band intensity was recorded by the Gel Doc imaging system.

[0105] (3) Experimental results

[0106] ① Electrophoresis image results

[0107] Control group: strong band amplified by M primer, weak band by U primer;

[0108] RG108 group: M band significantly weakened, U band enhanced, suggesting decreased methylation level.

[0109] ③ Changes in CDKN1A mRNA expression

[0110] RT-PCR detection showed that the expression of CDKN1A in the RG108 group was increased; the results were consistent with the MSP data, verifying that promoter demethylation led to gene activation.

[0111] (4) Experimental conclusion

[0112] RG108 treatment can significantly reduce the DNA methylation level in the CDKN1A promoter region; demethylation is accompanied by an increase in mRNA expression, suggesting transcriptional activation; it is shown that RG108 restores the expression of key cycle inhibitors through epigenetic regulation mechanisms; and it provides molecular mechanism support for its role in inducing T cell senescence and reducing immune rejection.

[0113] Example 3 RG108 alleviates cardiac transplant rejection Figure 3 、 Figure 4 )

[0114] (1) Experimental design

[0115] This experiment aims to verify whether RG108 can alleviate the immune rejection reaction after heart transplantation by inducing T cell senescence phenotype, and prolong the survival time of the graft.

[0116] The experiment uses the classic BALB / c→C57BL / 6 heterotopic heart transplantation mouse model. After the operation, the transplant recipients are intervened by systemic injection of RG108, and the differences in rejection degree, inflammatory cell infiltration, myocardial fibrosis, and graft survival time between the experimental and control groups are compared to evaluate the immunoprotective effect of RG108.

[0117] (2) Experimental method

[0118] ① Construction of transplantation model

[0119] Donor mice: 8-10 week old male BALB / c;

[0120] Recipient mice: 8-10 week old male C57BL / 6;

[0121] According to the "abdominal heterotopic heart transplantation" operation: the donor ascending aorta is anastomosed to the recipient abdominal aorta end-to-side; the pulmonary artery is anastomosed to the inferior vena cava; a successful graft can restore pulsation within 5 minutes after the operation.

[0122] ② Drug intervention settings

[0123] Experimental grouping:

[0124] Control group: postoperative injection of equal volume of normal saline;

[0125] RG108 group: postoperative intraperitoneal injection of RG108, 5 mg / kg;

[0126] Dosing time: 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 days after operation;

[0127] Dosing method: intraperitoneal injection, dose 5 mg / kg, 200 μL of PBS after dilution per injection;

[0128] Drug preparation: RG108 is dissolved in DMSO and diluted in PBS to the working concentration;

[0129] Number of animals in each group: n = 10.

[0130] ③Evaluation index and analysis method

[0131] Graft pulsation: daily record of graft pulsation, graft survival monitoring, survival curve drawing.

[0132] Histological analysis: HE staining, EVG staining of grafts on day 28, examination of myocardial necrosis, lymphocyte infiltration, collagen deposition.

[0133] (3) Experimental results

[0134] Graft survival time: compared with the control group, the heart graft survival time of the RG108 group was significantly prolonged, showing that RG108 can effectively inhibit graft failure Figure 3 ).

[0135] HE staining: reduced inflammatory cell infiltration, RG108 group graft myocardial structure was well preserved, lymphocyte infiltration was significantly reduced, and there was no obvious neutrophil aggregation or necrosis around the anastomotic stoma Figure 4 ).

[0136] EVG staining: compared with the control group, myocardial fibrosis was reduced, collagen deposition was reduced, suggesting that myocardial tissue damage was reduced Figure 4 ).

[0137] (4) Experimental conclusion

[0138] Systemic administration of DNMT1 inhibitor RG108 can significantly reduce mouse heterotopic heart transplant rejection; its effect is manifested as reduced myocardial tissue damage, reduced inflammation and fibrosis, and prolonged graft function maintenance time; compared with traditional immunosuppressants, RG108 has the advantages of clear epigenetic target, low toxicity, and long-lasting effect; the experimental results verify the effectiveness and scientificity of the application of RG108 to the heart transplant immunointervention strategy proposed by the application.

[0139] Comparative test of CTLA4-Ig control group and RG108 experimental group Figure 3 、 Figure 5 )

[0140] (1) Experimental design

[0141] To further illustrate the technical scheme of RG108 reducing heart transplant rejection proposed by the application, compared with the advantages of existing immune checkpoint inhibitors (such as CTLA4-Ig) in curative effect, this control test is designed.

[0142] (2) Experimental method

[0143] ① Model construction

[0144] Animal model: BALB / c→C57BL / 6 mouse abdominal heterotopic heart transplantation;

[0145] Evaluation of graft function: daily abdominal palpation to record heart beat.

[0146] ② Experimental grouping and drug administration

[0147] Control group (CTLA4-Ig group): use CTLA4-Ig recombinant protein (MCE);

[0148] Administration method: intraperitoneal injection;

[0149] Drug concentration: 1 μg / μL;

[0150] Drug volume: 250 μL / each;

[0151] Drug administration time: 0th day and 2nd day after operation;

[0152] Total dose: 2 times x 250 μg / each.

[0153] Experimental group (RG108 group)

[0154] Drug: RG108;

[0155] Administration method: intraperitoneal injection;

[0156] Drug concentration: 5 mg / kg;

[0157] Dosing time: 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 days after operation;

[0158] The volume of each injection is 200 μL, and the injection is continuously performed for 14 times.

[0159] 3. Detection index

[0160] Survival time of the graft;

[0161] HE staining and EVG staining (tissue structure and fibrosis): the heart graft is dyed 28 days after operation.

[0162] (3) Experimental results

[0163] Survival time of the graft: compared with the control group, the survival time of the heart graft in the RG108 group is significantly prolonged, and the function of the graft is maintained Figure 3 ).

[0164] Histopathology: HE staining of the CTLA4-Ig group shows that inflammatory cell infiltration and myocardial damage occur, and EVG staining shows obvious vascular fibrosis; the RG108 group shows that the myocardial structure is complete, and the degree of inflammation and fibrosis is obviously reduced Figure 5 ).

[0165] (4) Conclusion

[0166] The RG108 treatment scheme provided by the application can significantly prolong the survival time of the graft; in terms of improving the survival time of the graft, reducing tissue damage, and inhibiting inflammatory cell infiltration, RG108 shows more persistent and stable effects; meanwhile, RG108 is mild and has low toxicity, and has more clinical conversion prospects. Therefore, compared with the existing immunological intervention strategies, the application has substantial progress and obvious advantages.

[0167] The above-described embodiments are only used to describe the preferred modes of the application, and do not limit the scope of the application. Without departing from the design spirit of the application, various modifications and improvements to the technical solutions of the application made by those skilled in the art should fall within the protection scope of the claims of the application.

Claims

1. The use of RG108 in the preparation of immunosuppressive drugs for the prevention and treatment of organ transplantation, characterized in that: The chemical structure of RG108 is as follows: 。 2. The use according to claim 1, characterized in that The organ includes the heart.

3. The use according to claim 2, characterized in that The targets of the drug include T cells.

4. The use according to claim 3, characterized in that The drug inhibits DNMT1, reduces CDKN1A promoter methylation, upregulates CDKN1A expression, blocks T cell cycle, activates aging phenotype, and thus exerts an immunosuppressive effect on organ transplantation.

5. The use according to claim 4, characterized in that The dosage range of RG108 is 2.5–10 mg / kg / day.

6. The use according to claim 5, characterized in that The optimal dose of RG108 is 5 mg / kg / day.

7. The use according to claim 6, characterized in that The frequency of use of RG108 is once a day.

8. The use according to claim 7, characterized in that The medicine further comprises one or more pharmaceutically acceptable carriers and / or excipients.

9. The use according to claim 8, characterized in that The drug may also contain other immunosuppressants.

10. The use according to claim 9, characterized in that The other immunosuppressants include CTLA4-Ig.

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