Use of ptprj agonists in the preparation of a medicament for the prevention and / or treatment of renal fibrosis

Abstract

The application discloses application of a Ptprj agonist GJ103 in preparation of a medicine for preventing and / or treating renal fibrosis. It is disclosed that active up-regulation of Ptprj can obviously inhibit proliferation and activation of fibroblasts caused by TGF-beta 1, and knockout of Ptprj aggravates renal fibrosis caused by UUO and promotes activation of fibroblasts. As a Ptprj small molecule agonist, GJ103 treatment not only obviously inhibits renal fibroblast activation induced by TGF-beta 1, but also obviously improves pathological damage of renal tissues caused by obstruction and ischemia and reduces deposition of extracellular matrix. The results show that Ptprj plays a protective role in the process of renal fibrosis, and its agonist GJ103 provides a new target for prevention and diagnosis and treatment of CKD.

Description

Technical Field

[0001] This invention relates to the pharmaceutical field, specifically to the use of Ptprj agonists in the preparation of medicaments for the prevention and / or treatment of renal fibrosis. Background Technology

[0002] Chronic kidney disease (CKD) is a long-term disease characterized by progressive and irreversible loss of kidney function or persistent kidney damage[1]. It is projected to become the fifth leading cause of death worldwide by 2040. The high incidence and high mortality of CKD have made it a global public health problem. Regardless of the cause of CKD, the common pathological manifestation is renal fibrosis. It is characterized by excessive production / deposition of extracellular matrix (ECM)[2], leading to tissue remodeling and renal parenchymal scarring. In this process, the proliferation and activation of fibroblasts are the clear effects and driving factors of renal fibrosis[3]. The pathophysiological mechanism of CKD is relatively complex, and there is a lack of effective and specific prevention and treatment methods in clinical practice. Therefore, in-depth exploration of the molecular mechanism behind renal fibrosis and finding effective targets are the key to delaying the chronic progression of kidney disease and blocking renal fibrosis.

[0003] Protein tyrosine phosphatase receptor J (Ptprj) is a 220 kDa transmembrane protein belonging to the receptor-type protein tyrosine phosphatase family and participating in multiple signaling pathways [4]. Ptprj is expressed in various cell types, including hematopoietic cells, endothelial cells, fibroblasts, thyroid cells, and mammary cells. Its structure consists of a single intracellular catalytic PTP domain, a transmembrane domain, and an extracellular domain composed of nine fibronectin type III repeat sequences. It regulates multiple cellular processes, including cell growth, differentiation, mitosis, and oncogenic transformation, through dephosphorylation or participation in the dephosphorylation of other protein tyrosine kinases [5-7]. Studies have found that fibrosis involves multiple pathways, and PDGF signal transduction is one of the central mediators. Ptprj can downregulate PDGFRβ phosphorylation levels and also has a negative regulatory effect on cell proliferation and migration [8-10]. The role of Ptprj in renal fibrosis is still unclear.

[0004] GJ103 is a general-reading compound with CAS number 1459687-89-8 and structural formula as follows:

[0005]

[0006] GJ103 can induce premature codon readthrough and has research potential in genetic diseases caused by nonsense mutations. As a GJ072 analog, GJ103 can induce ATM kinase activity in AT153LA cells with homozygous TGA and TAA mutations. Currently, there are no studies on GJ103 in kidney diseases.

[0007] The references are as follows:

[0008] 1.Zhong,J.,HCYang,and ABFogo,A perspective on chronic kidneydisease progression.Am J Physiol Renal Physiol, 2017.312(3):p.F375-F384.

[0009] 2. Bulow, RD and P. Boor, Extracellular Matrix in Kidney Fibrosis: MoreThan Just aScaffold.J Histochem Cytochem, 2019.67(9):p.643-661.

[0010] 3. Falke LL, Gholizadeh S, Goldschmeding R, et al. Diverse origins of themyofibroblast-implications for kidney fibrosis[J]. Nat Rev Nephrol, 2015, 11(4): 233-244.

[0011] 4.Keane MM, Lowrey GA, Ettenberg SA, et al. The protein tyrosinephosphatase DEP-1 is induced during differentiation and inhibits growth ofbreast cancer cells[J]. Cancer Res, 1996, 56(18):4236-4243.

[0012] 5.Grazia Lampugnani,M.,et al.,Contact inhibition of VEGF-inducedproliferation requires vascular endothelial cadherin,beta-catenin,and thephosphatase DEP-1 / CD148.JCell Biol,2003.161(4):p.793-804.

[0013] 6.Schwarz,M.,et al.,Disrupting PTPRJ transmembrane-mediatedoligomerization counteracts oncogenic receptor tyrosine kinase FLT3 ITD.FrontOncol,2022.12:p.1017947.

[0014] 7.Hendriks,W.,et al.,Proteinaceous Regulators and Inhibitors ofProtein Tyrosine Phosphatases.Molecules,2018.23(2).

[0015] 8.Sala M,Spensiero A,Scala MC,et al.Design,Synthesis,BiologicalActivity,and Structural Analysis of Lactam-Constrained PTPRJ AgonistPeptides.ChemMedChem 2018;13:1673-1680.

[0016] 9.Buhl EM,Djudjaj S,Klinkhammer BM,et al.Dysregulated mesenchymalPDGFR-beta drives kidney fibrosis.EMBO Mol Med 2020;12:e11021.

[0017] 10. Chen YT, Chang FC, Wu CF, et al. Platelet-derived growth factor receptor signaling activates pericyte-myofibroblast transition in obstructive and post-ischemic kidney fibrosis. Kidney international 2011;80:1170-1181. Summary of the Invention

[0018] Purpose of the invention: The purpose of this invention is to propose the application of the Ptprj agonist GJ103 in the preparation of drugs for the prevention and / or treatment of renal fibrosis, providing a new pharmaceutical use for GJ103, and at the same time, providing a new treatment strategy for chronic kidney disease.

[0019] This application reveals that actively upregulating Ptprj significantly inhibits TGF-β1-induced fibroblast proliferation and activation, while knocking out Ptprj exacerbates renal fibrosis induced by UUO and promotes fibroblast activation. As a small molecule Ptprj agonist, GJ103 treatment not only significantly inhibits TGF-β1-induced renal fibroblast activation but also significantly improves renal tissue pathological damage caused by obstruction and ischemia and reduces extracellular matrix deposition.

[0020] Specifically, this application first identified the expression and localization of Ptprj in the renal tissue of CKD patients. Immunohistochemistry revealed that Ptprj expression was low in normal renal tissue, but significantly increased in the renal tubules and interstitium of CKD patients. Further double staining with Ptprj and Fsp1 confirmed increased expression in the interstitium. In vitro and in vivo experiments were conducted to explore the potential function of Ptprj in CKD. Overexpression of Ptprj in cultured rat fibroblasts (NRK49F) significantly inhibited TGF-β1-induced Collagen I and Collagen III mRNA expression and FN1 and Vimentin protein expression, while also inhibiting the expression of α-SMA, a marker of fibroblast activation. By constructing Ptprj heterozygous knockout mice, the function of Ptprj in CKD was further clarified. qRT-PCR analysis showed that Ptprj knockdown exacerbated the deposition of FN1, an extracellular matrix component, and Western blot analysis revealed a significant increase in the expression of FN1, Collagen I, and Collagen III proteins. In UUO mice, Ptprj knockdown also increased the proliferation and activation of fibroblasts characterized by high α-SMA expression. Similarly, we found that Ptprj knockdown increased PDGFRβ phosphorylation levels in fibrotic kidneys compared to the control group, suggesting that Ptprj may regulate renal fibrosis through PDGFRβ.

[0021] The above data suggest that targeting Ptprj has therapeutic potential in the molecular intervention mechanism of CKD. Therefore, we screened for effective Ptprj activators through molecular docking and further screened small molecule agonists with high docking scores by detecting the dephosphorylation activity of Ptprj on PDGFRβ. First, after stimulating NRK49F cells with 10 small molecules, we found that AS252424 and GJ103 significantly inhibited the phosphorylation level of PDGFRβ, and also promoted the dephosphorylation of PDGFRβ after co-stimulation with TGF-β1. Next, through in vitro dephosphorylation of PDGFRβ, we found that GJ103 could better activate Ptprj. Finally, quantitative analysis using surface plasmon resonance (SPR) revealed that GJ103 has a high affinity for recombinant Ptprj protein.

[0022] To further verify the protective effect of GJ103 in renal fibrosis caused by chronic kidney disease (CKD), we pretreated NRK49F cells with different concentrations of GJ103 (5, 10, 20, and 40 μM) for 2 h, followed by TGF-β1 stimulation. The results showed that when the GJ103 concentration was higher than 20 μM, the phosphorylation level of PDGFRβ was significantly inhibited. With the decrease in p-PDGFRβ expression, the protein level of the fibrosis marker FN1 also decreased further. Subsequently, the function of GJ103 was verified in animal models. The results indicated that high-dose GJ103 (30 mg / kg / day) significantly improved renal fibrosis induced by UUO and UIRI. This was mainly manifested in the reduction of renal interstitial collagen deposition and extracellular matrix, as well as the inhibition of fibroblast activation. In the UUO and UIRI models, the therapeutic dose of GJ103 showed no significant toxicity to the kidneys, liver, heart, or systemic system.

[0023] The results in summary indicate that GJ103 and its sodium salt have great potential in the preparation of drugs for the prevention and / or treatment of chronic kidney disease, and can effectively alleviate renal fibrosis caused by chronic kidney disease. Attached Figure Description

[0024] Figure 1 A spatiotemporal expression relationship diagram of Ptprj and the development of renal fibrosis in CKD;

[0025] Figure 2 Figure showing the effect of Ptprj overexpression on TGF-β1-induced renal fibroblast activation;

[0026] Figure 3 Figure showing the effect of Ptprj knockout on UUO-induced extracellular matrix deposition and fibroblast activation;

[0027] Figure 4 Mechanism diagram for screening the Ptprj small molecule agonist GJ103;

[0028] Figure 5 Figure showing the effect of GJ103 treatment on TGF-β1-induced renal fibroblast activation;

[0029] Figure 6 Figure showing the effect of GJ103 treatment on UUO-induced extracellular matrix deposition and fibroblast activation;

[0030] Figure 7 Figure showing the effect of GJ103 treatment on UIRI-induced extracellular matrix deposition and fibroblast activation. Detailed Implementation

[0031] The present invention is further illustrated below with reference to specific embodiments. Unless otherwise specified, all reagents used in the present invention are commercially available. Rat fibroblasts (NRK-49F) were obtained from ATCC (American Type Culture Collection). Ptprj-KO mice and C57BL / 6J mice were obtained from Jicui Yaokang (Nanjing, China), and GJ103 was obtained in its sodium salt form.

[0032] The experimental methods and materials used in the following embodiments are as follows:

[0033] Animal model preparation: A unilateral ureteral obstruction (UUO) model was constructed to evaluate the effect of Ptprj on CKD. Ptprj heterozygous knockout (Ptprj + / - Mice were randomly divided into 4 groups (sham-operated group: WT+Sham group and Ptprj group). + / - +Sham group; Model groups: WT+UUO group and Ptprj + / - (UUO group). Mice were anesthetized with isoflurane. After successful anesthesia, the mice were immobilized with their limbs facing upwards. The abdomen was disinfected, and the abdominal cavity was opened and exposed along the midline of the abdomen using ophthalmic scissors. The left ureter was located and isolated. The left ureter was ligated with 4-0 sutures near the renal pelvis. The mouse organs were repositioned completely, the abdominal cavity was closed, and the peritoneum and skin were sutured. In the sham surgery group, only the abdominal cavity needed to be opened and the abdomen sutured. After the surgery, the mice were fed in the SPF-grade animal facility of the Experimental Animal Center of Nanjing Medical University, and their health was monitored daily postoperatively. Seven days after the UUO surgery, the mice were euthanized, blood was drawn from the heart, and kidney tissue was collected. Kidney tissue used for histological analysis was fixed in 4% paraformaldehyde, and the remaining kidney tissue was stored at -80°C for mRNA and protein analysis. All animal husbandry and handling were approved by the Institutional Animal Care and Use Committee of Nanjing Medical University.

[0034] A unilateral ureteral obstruction (UUO) model was established to evaluate the effect of GJ103 on CKD. Eight-week-old male C57BL / 6J mice were randomly divided into four groups (sham-operated group: Sham group; model group: UUO group, UUO + GJ103 - 15 mg / kg / day group, UUO + GJ103 - 30 mg / kg / day group). Mice were pretreated with two concentrations of GJ103 (15 mg / kg / day and 30 mg / kg / day) intraperitoneally for one day each. UUO surgery was then performed, following the same procedure. Mice were subsequently treated with two concentrations of GJ103 (15 mg / kg / day and 30 mg / kg / day) for seven consecutive days. Seven days after UUO surgery, the mice were euthanized, and the remaining procedures were the same.

[0035] A unilateral ischemia-reperfusion (UIRI) model was constructed to evaluate the effect of GJ103 on CKD. Eight-week-old male C57BL / 6J mice were randomly divided into three groups (sham-operated group: Sham group; model groups: UIRI group and UIRI+GJ103-30mg / kg / day group). Mice were pretreated with GJ103 (30mg / kg / day) intraperitoneally for one day. Then, the UIRI procedure was performed as follows: Mice were anesthetized with isoflurane. After successful anesthesia, the mice were fixed with their limbs facing upwards on a 37°C incubator. The abdomen was disinfected, and the peritoneal cavity was carefully opened and exposed using ophthalmic scissors. The left renal pedicle was freed, and the left renal pedicle was clamped using a non-invasive arterial clamp. The kidney gradually changed from bright red to dark red, indicating successful clamping. The mouse abdomen was covered with warm saline-moistened gauze. The clamp was removed after 32 minutes. The left kidney gradually changed from dark red to bright red, indicating restored reperfusion. The abdominal organs were repositioned, and 1 mL of warm saline was injected to replenish lost fluids. The abdominal cavity of the mouse was closed, and the peritoneum and skin were sutured. In the control group, only the abdominal cavity of the mouse needed to be opened and the abdomen sutured. After the surgery, the mice were fed in an SPF-grade animal facility, and their condition was monitored daily postoperatively. The mice were then treated with GJ103 (30 mg / kg / day) for 14 consecutive days. Two weeks after the UIRI surgery, the mice were euthanized, blood was extracted from the heart, and kidney tissue was collected. The remaining steps were the same as above.

[0036] Histological analysis: Kidney tissue was fixed in 4% paraformaldehyde for more than 24 hours, then dehydrated and embedded. Paraffin sections (3 μm) were dewaxed and hydrated, and then stained with Masson's trichrome and Sirius red to visualize collagen fibers. The fibrosis area was analyzed against the total area using Image Pro Plus software.

[0037] Immunohistochemical staining: Paraffin sections (3 μm) were dewaxed, hydrated with xylene and graded ethanol, incubated with 3% hydrogen peroxide for 20 minutes, and then boiled in citrate antigen extraction solution for 20 minutes. After blocking with immunostaining blocking solution for 1 hour, the sections were incubated with primary antibody overnight at 4°C. DAB was used for colorimetric reaction, and hematoxylin was used to counterstain cell nuclei. Image analysis and quantification were performed using Image Pro Plus.

[0038] Cell culture: NRK-49F cells were grown adherently in DMEM medium supplemented with 10% FBS and incubated at 37°C with 5% CO2. Cells were passaged with 0.25% trypsin at 70-80% confluence. In specific experiments, cells were pretreated with GJ103 for 2 hours or transfected with Ptprj plasmid for 6 hours, followed by stimulation with TGF-β1. Cells were collected for mRNA analysis after 24 hours or for protein analysis after 48 hours.

[0039] EdU staining: NRK-49F cells were cultured in 6-well plates with EdU working solution (10 μM) for 2 hours. The culture medium was removed, and 1 ml of fixative was added, incubating at room temperature for 15 min. Then, 1 ml of osmotic solution was added, and the plates were incubated at room temperature for 10 min. Finally, 0.5 ml of Click reaction solution was added to each well, and the plates were incubated at room temperature in the dark for 30 min. Cell nuclei were stained with DAPI.

[0040] Cell proliferation and cytotoxicity assays: NRK-49F cells were seeded in 96-well plates and treated with different concentrations of GJ103 (0-40 nM). After 24 hours of culture, 10 μl of CCK-8 reagent was added to each well and incubated for 2 hours. The absorbance at 450 nm was measured using a microplate reader.

[0041] Real-time quantitative PCR: Total RNA was extracted from tissues or cells using RNA isoPlus reagent. qPCR reverse transcription was performed using HiScript IIQ RT SuperMix. Real-time PCR amplification was performed on a LightCycler96 real-time PCR detection system using AceQ qPCR SYBR Green Master Mix. GAPDH was used as an internal control, and relative values ​​were calculated using the ΔΔCt method.

[0042] Western blotting (WB): Tissues or cells were lysed with RIPA buffer containing 1×Protease inhibitor and 1×Phosphatase inhibitor. Samples were centrifuged at 5000 rpm for 30 min. Protein concentrations were determined using a BCA protein assay kit. Equal masses of samples were spotted onto a 10% polyacrylamide gel, which was then transferred to a PVDF membrane. The PVDF membrane was blocked with 5% skim milk powder for 1 h, followed by overnight incubation with antibodies against FN1 (1:1000), type I collagen (1:000), type III collagen (1:1000), and GAPDH (1:10000). Bands were visualized using an Amersham Biosciences ECL detection system. Relative quantitative analysis of grayscale values ​​was performed using ImageJ.

[0043] Immunofluorescence staining: NRK-49F cells were cultured on glass-bottomed cell culture dishes (NEST, China), pretreated with Apcin (50 nm) for 0.5 h, and then treated with recombinant human TGF-β1 (10 ng / ml). After washing repeatedly with PBS, the cells were fixed with 4% paraformaldehyde at room temperature for 20 min, washed extensively with PBS, infiltrated with PBS + 1% Triton X-100 for 10 min, and blocked with 2% BSA for 1 h. Primary antibody α-SMA was added to the culture dishes at a 1:100 dilution, along with the cytoskeletal protein dye phalloidin, and the cells were cultured overnight in a humidified chamber at 4°C. After washing with PBST, secondary antibody was applied. The cells were then observed, photographed, and recorded under a laser confocal scanning microscope (Zeiss).

[0044] SPR: The binding affinity between GJ103 and Ptprj was detected using the BIAcore T200 SPR biosensor system and the NTA sensor chip (Cytiva). First, the purified protein Ptprj was diluted to 20 μg / mL, at which point the coupling between Ptprj and the chip was optimal. Next, the binding and dissociation efficiencies of GJ103 and Ptprj were measured. Seven analytical concentrations of GJ103 were set: 0 μM, 3.125 μM, 6.25 μM, 12.5 μM, 25 μM, 50 μM, and 100 μM. The flow rate was set to 30 μL / min. The binding time and dissociation time were both 120 s. Finally, the dynamic parameters were determined. The experiment was conducted in multiple cycles, and the data were analyzed using BIAcore T200 software.

[0045] In vitro PDGFRβ dephosphorylation: NRK49F cells were stimulated with TGF-β1 (10 ng / ml) at 70% confluence. After 24 h, the cells were washed twice with pre-chilled PBS. After lysis and centrifugation, the cells were inverted overnight at 4°C with PDGFRβ antibody and magnetic beads. The immune complexes on the magnetic beads were collected and washed four times with washing buffer. The PDGFRβ immune complexes immobilized on the magnetic beads were aliquoted into 40 μl buffer and incubated with purified proteins Ptprj, GJ103, and AS252424 at 30°C for 30 min. The total volume was 60 μl. Dephosphorylation was then terminated by adding SDS buffer, and Western blot analysis was performed.

[0046] Statistical analysis: Graphpad 6.0 statistical software was used to perform statistical analysis on all experimental data. Experimental data are expressed as mean ± standard error of mean (SEM). Two-tailed t-tests were used to compare two groups, and one-way ANOVA was used to compare data from multiple groups. A p-value < 0.05 was considered statistically significant.

[0047] Example 1: Spatiotemporal expression relationship between Ptprj and the development of renal fibrosis in CKD.

[0048] Immunohistochemical results using renal tissue specimens from clinical CKD patients showed that, compared with normal human renal tissue, Ptprj expression was significantly upregulated in the renal tissue of CKD patients, mainly increasing in the renal tubules and interstitium. Figure 1 (a and b). Further immunofluorescence co-localization revealed that double staining with Fsp1 (a marker of mesenchymal fibroblasts) and Ptprj confirmed increased Ptprj expression in the mesenchyme. Figure 1 (c) The above results indicate that Ptprj may be involved in the progression of CKD.

[0049] Example 2: Overexpression of Ptprj inhibits TGF-β1-induced activation of renal fibroblasts.

[0050] We validated this by overexpressing Ptprj in rat fibroblasts (NRK-49F). EdU staining results showed that overexpression of Ptprj significantly inhibited TGF-β1-induced fibroblast proliferation and activation. Figure 2 a). WB and qRT-PCR showed that the protein levels of PAI1, FN1, and Vimentin were ( Figure 2 b and c), mRNA levels of fibrosis-related molecules Collagen I and Collagen III ( Figure 2 d and e) were also significantly downregulated, and immunofluorescence also showed a significant decrease in α-SMA protein levels. Figure 2 f). Preliminary in vitro experiments suggest that Ptprj plays a certain protective role in the progression of renal fibrosis in CKD.

[0051] Example 3: Downregulation of Ptprj exacerbates UUO-induced extracellular matrix deposition and fibroblast activation.

[0052] Renal fibrosis is the common final pathway and histological manifestation of CKD, characterized by excessive extracellular matrix deposition, leading to tissue remodeling and renal parenchymal scarring. Numerous studies have shown that a series of signal transduction processes induced by the proliferation and activation of renal tubulointerstitial fibroblasts are the main factors initiating and advancing renal fibrosis. To further elucidate the role of Ptprj in renal fibrosis, we bred a Ptprj heterozygous knockout strain (Ptprj...). + / - A UUO model was established in mice. Masson staining and Sirius red staining showed that, compared with the WT group, Ptprj + / - In mice, UUO-induced renal tissue pathological damage was further aggravated, with a significant increase in collagen fiber deposition. Figure 3 qRT-PCR analysis showed that Ptprj knockdown upregulated FN1 expression. Figure 3 e), Western blot analysis showed a significant increase in the protein expression levels of FN1, Collagen I, and Collagen II. Figure 3 f and g). In UUO mice, knockdown of Ptprj increased the proliferation and activation of fibroblasts characterized by high α-SMA expression. Figure 3 Similarly, we found that, compared with the control group, Ptprj knockdown increased the level of PDGFRβ phosphorylation in the kidneys of fibrotic lesions (hj). Figure 3 f&g), which suggests that Ptprj may regulate renal fibrosis through PDGFRβ.

[0053] Example 4: Screening of Ptprj small molecule agonists.

[0054] The above data suggest that targeting Ptprj has therapeutic potential in the molecular intervention mechanism of CKD. Therefore, we screened for effective Ptprj activators through molecular docking and further screened small molecule agonists with high docking scores by detecting the dephosphorylation activity of Ptprj on PDGFRβ. After stimulating NRK49F cells with 10 small molecule compounds, we found that AS252424 and GJ103 significantly inhibited the phosphorylation level of PDGFRβ. Figure 4 a) When co-stimulated with TGF-β1, it also significantly promoted the dephosphorylation of PDGFRβ. Figure 4 b&c). Next, through in vitro dephosphorylation of PDGFRβ, we found that GJ103 could better activate Ptprj ( Figure 4 d). As shown in the combined model of 2D and 3D diagrams, GJ103 can form two conventional hydrogen bonds with GLY1244 and VAL1243, a salt bridge with ARG1245, and a Pi-cation and a Pi-Pi stack (…). Figure 4 Finally, surface plasmon resonance (SPR) quantitative analysis revealed that GJ103 has a high affinity for the Ptprj recombinant protein, with a binding affinity (KD value) of 6.399 μM (e&f). Figure 4 g). The effect of GJ103 on Ptprj was further verified in NRK49F cells. Under stimulation with different concentrations of GJ103, the expression level of Ptprj remained essentially unchanged ( Figure 4 h&i).

[0055] Example 5: GJ103 treatment inhibits TGF-β1-induced renal fibroblast activation.

[0056] GJ103 is a readthrough compound that can induce readthrough of premature stop codons, showing potential in genetic diseases caused by nonsense mutations. Currently, the role of GJ103 in kidney diseases has not been reported. We found that different concentrations of GJ103 had little effect on cell viability. Figure 5 a). Subsequently, we evaluated the protective effect of GJ103 in vitro. NRK49F cells were pretreated with different concentrations of GJ103 (5, 10, 20, and 40 μM) for 2 h, followed by stimulation with TGF-β1. We found that when the concentration of GJ103 was higher than 20 μM, the phosphorylation level of PDGFRβ was significantly inhibited (a). Figure 5 (b&c). With the decrease in p-PDGFRβ expression, the protein expression level of the fibrosis marker FN1 was also suppressed. Figure 5 d&e).

[0057] Example 6: GJ103 treatment reduced UUO-induced extracellular matrix deposition and fibroblast activation.

[0058] To further validate the protective effect of GJ103 in animal models, we established a classic unexamined kidney fibrosis (UUO) model. C57BL / 6J mice were pretreated with GJ103 at 15 mg / kg / day and 30 mg / kg / day, respectively, before UUO surgery. The mice were then administered GJ103 daily for 7 consecutive days. Serum levels of BUN, Scr, AST, ALT, CK-MB, and LDH were measured. The results showed that therapeutic doses of GJ103 did not cause significant renal, hepatic, cardiac, or systemic toxicity. Figure 6 ag). Masson staining and Sirius red staining showed that GJ103 stimulation, especially at high concentrations, significantly reduced renal interstitial fibrosis induced by UUO. Figure 6 Similarly, the mRNA (Figure kn) and protein (hj) of FN1, Collagen I, Collagen III, and α-SMA Figure 6 The expression of o) was also reduced, especially in the high-dose group.

[0059] Example 7: GJ103 treatment inhibits UIRI-induced extracellular matrix deposition and fibroblast activation.

[0060] We also examined the effect of GJ103 in the UIRI model, and the results were consistent with those in the UUO model; therapeutic doses of GJ103 did not cause significant renal, hepatic, cardiac, or systemic toxicity. Figure 7 Similarly, high-dose GJ103 (30 mg / kg / day) also significantly improved UIRI-induced renal histopathological damage (af). Figure 7 g&h), reduced extracellular matrix deposition ( Figure 7The expression of i&j) and α-SMA, a marker of fibroblast activation ( Figure 7 (k&l). In vitro and in vivo experiments have confirmed that Ptprj agonists play a certain protective role in renal fibrosis in CKD.

[0061] The above experimental results indicate that treatment with the Ptprj agonist GJ103 significantly reduced obstructed or ischemic kidney tissue, as well as TGF-β1-induced extracellular matrix deposition and fibroblast activation. In conclusion, Ptprj plays a crucial role in the occurrence and progression of CKD, providing a new approach for the prevention and targeted treatment of renal fibrosis.

Claims

1. The use of Ptprj agonists in the preparation of medicaments for the prevention and / or treatment of renal fibrosis, characterized in that, The Ptprj agonist is GJ103.

2. The application according to claim 1, characterized in that, The renal fibrosis described is renal fibrosis caused by the proliferation and activation of fibroblasts induced by TGF-β1.

3. The application according to claim 1, characterized in that, The renal fibrosis mentioned refers to renal tissue pathological damage and extracellular matrix deposition caused by unilateral ureteral obstruction.

4. The application according to claim 1, characterized in that, The renal fibrosis mentioned refers to renal fibrosis caused by pathological damage to renal tissue and deposition of extracellular matrix due to unilateral ischemia-reperfusion.

5. The application according to claim 1, characterized in that, The renal fibrosis described is renal fibrosis caused by unilateral ureteral obstruction and unilateral ischemia-reperfusion induced by fibroblast proliferation and activation.

6. The application according to claim 1, characterized in that, The drug also includes its pharmaceutically acceptable salts.

7. The application according to claim 1, characterized in that, The drug is GJ103 sodium salt.

Images