Use of acs extract in the preparation of a medicament for treating icis-induced liver injury
By preparing total saponin extracts from *Abrus precatorius*, the problem of liver damage caused by immune checkpoint inhibitors was solved, achieving effective prevention and treatment of liver damage, reducing liver inflammation and immune infiltration, and providing a safe treatment option.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- NANFANG HOSPITAL OF SOUTHERN MEDICAL UNIV
- Filing Date
- 2026-02-10
- Publication Date
- 2026-06-09
AI Technical Summary
There is a lack of effective treatments for liver damage caused by immune checkpoint inhibitors in the current technology, especially since hormone therapy has serious side effects, affecting treatment efficacy and patient health.
The total saponin extract of *Abrus precatorius* was used as the drug component. It was extracted by hot reflux of ethanol and purified by macroporous resin to prepare oral or injectable dosage forms for the prevention and treatment of liver damage caused by immune checkpoint inhibitors.
It significantly reduced liver injury-related ALT and AST levels, decreased hepatic immune infiltration, improved liver inflammation, and provided a safe and effective alternative treatment, avoiding the side effects of hormone therapy.
Smart Images
Figure CN122163677A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of traditional Chinese medicine, specifically relating to the application of ACS extract (total saponin extract of chicken bone grass) in the preparation of a drug for treating liver damage caused by ICIs (immune checkpoint inhibitors). Background Technology
[0002] Immune checkpoint inhibitors (ICIs), as an important means of cancer immunotherapy, significantly enhance T cell-mediated anti-tumor immune responses by blocking key immune checkpoints such as PD-1 / PD-L1 and CTLA-4, and have become the standard treatment for many advanced solid tumors and hematologic malignancies. However, the widespread use of ICIs has also led to a significant increase in immune-related adverse events (irAEs). Among them, liver injury caused by immune checkpoint inhibitors is a common clinical complication, which not only forces treatment interruption and affects the efficacy of anti-tumor therapy, but also seriously threatens the quality of life of patients. Therefore, finding safe and effective treatment methods is particularly important.
[0003] For liver injury caused by immunosuppressants, clinical practice typically involves dose reduction or medication replacement. However, this may affect the effectiveness of immunosuppression and increase the risk of rejection. Furthermore, high-dose corticosteroids (such as prednisone) are the standard first-line treatment for liver injury caused by immunosuppressants. However, long-term or high-dose use of hormones is often accompanied by various serious side effects, including osteoporosis, hyperglycemia, a significantly increased risk of secondary infections, and the potential weakening of the body's anti-tumor immune response. Therefore, developing highly effective, low-toxicity, and especially non-immunosuppressive alternative prevention and treatment strategies is a major clinical need that urgently needs to be addressed.
[0004] Traditional Chinese medicine, with its characteristics of "multiple components, multiple targets, and multiple pathways," may have significant advantages in the treatment of ICI liver injury. *Abrus cantoniensis*, the dried whole plant of the legume *Abrus cantoniensis*, is a commonly used specialty medicinal material in the Lingnan region and is included in the Chinese Pharmacopoeia (2020 edition). *Abrus cantoniensis* is sweet and slightly bitter in taste, and cool in nature. It has the effects of clearing heat and detoxifying, promoting diuresis and relieving jaundice, and soothing the liver and relieving pain. It is often used to treat damp-heat jaundice, hypochondriac discomfort, stomach distension and pain, and mastitis. Clinically, compound preparations based on *Abrus cantoniensis* have been widely used to treat hepatitis and liver fibrosis, showing certain efficacy and suggesting its great potential in liver disease treatment. However, its main active ingredients and specific mechanisms of action remain unclear. Furthermore, there is no experience in treating liver injury caused by immune checkpoint inhibitors with *Abrus cantoniensis*, therefore, research on the therapeutic effect of *Abrus cantoniensis* in ICI liver injury is urgently needed. Summary of the Invention
[0005] The purpose of this invention is to overcome at least one deficiency of the prior art and to provide the application of total saponin extract of *Abrus precatorius* in the preparation of a medicament for treating liver damage caused by immune checkpoint inhibitors.
[0006] The technical solution adopted in this invention is:
[0007] In a first aspect, the present invention provides the use of total saponin extract of *Abrus precatorius* in the preparation of a medicament for treating liver injury caused by immune checkpoint inhibitors.
[0008] Secondly, the present invention provides the use of total saponin extract of *Abrus precatorius* in the preparation of a medicament for preventing liver damage caused by immune checkpoint inhibitors.
[0009] In some embodiments, the total saponin extract of *Abrus precatorius* provided by the present invention can be prepared by the following method:
[0010] 1) After coarsely crushing the chicken bone grass, extract it using the hot reflux method with ethanol. After concentrating the extract, obtain the ethanol concentrate, and add water to obtain the crude extract A.
[0011] 2) Petroleum ether was added to crude extract A for extraction, and the aqueous layer was collected to obtain crude extract B;
[0012] 3) Add n-butanol to crude extract B for extraction, collect the n-butanol layer, evaporate the n-butanol to dryness, and dissolve it in water to obtain crude extract C;
[0013] 4) Purify the crude extract C through a macroporous resin, elute with water and ethanol, collect the eluent, recover the ethanol, concentrate and dry to obtain the final product.
[0014] In some embodiments, the immune checkpoint inhibitor is selected from anti-PD-L1 antibody, anti-PD-1 antibody, anti-CTLA-4 antibody, and combinations thereof.
[0015] In some embodiments, the total saponin extract of *Abrus precatorius* contains daidzein I, sophora japonica saponin III, dehydrodaidzein I, absinthecin F, and absinthecin SO1.
[0016] In some embodiments, the medicament comprises an effective amount of total saponin extract of *Abrus precatorius* and a pharmaceutically acceptable carrier or excipient.
[0017] In some embodiments, the dosage form of the drug is an oral liquid, pill, granule, capsule, tablet, drop pill, injection, or powder.
[0018] In some embodiments, the total saponin extract of *Abrus precatorius* is an organic solvent extract of total saponins of *Abrus precatorius*.
[0019] In some embodiments, the organic solvent extract of total saponins from *Abrus precatorius* is selected from the n-butanol extract of total saponins from *Abrus precatorius*.
[0020] The beneficial effects of this invention are:
[0021] Clinically, there is a lack of effective interventions for liver injury caused by immunosuppressants. Generally, treatment involves discontinuing the medication or using hormone therapy, but hormone therapy has serious side effects. This invention provides a novel preventative treatment method, using total saponins from *Abrus precatorius* for the first time to preventively treat liver injury caused by immunosuppressants, and demonstrating its significant therapeutic effect. After preventative treatment with total saponins from *Abrus precatorius*, liver injury caused by immunosuppressants was significantly alleviated in Pdcd1 knockout C57BL / 6 mice, with a significant decrease in ALT and AST levels and a significant reduction in hepatic immune infiltration, indicating that the preventative treatment of liver injury caused by immunosuppressants with total saponins from *Abrus precatorius* is effective. Attached Figure Description
[0022] Figure 1 This is a schematic diagram of an animal experimental protocol for the preventive treatment of ICI liver injury with total saponins from *Heliotropium indicum*.
[0023] Figure 2 The pharmacodynamics of total saponins from *Aristolochia debilis* in the preventive treatment of ICI liver injury (n=6) is shown in the following figures: (A) End-stage AST level in mouse serum; (B) Baseline ALT level in mouse serum; (C) End-stage ALT level in mouse serum; (D) Changes in ALT level in mouse serum; (E) Trend of ALT changes in mouse serum; (F) HE staining results of mouse liver; Compared with the normal control group, *P<0.05, **P<0.01, ***P<0.001; Compared with the model group, #P<0.05, ##P<0.01, ###P<0.001.
[0024] Figure 3 The study investigated the effect of ACS on ICI-induced liver inflammation (n=3), specifically as follows: (A) mRNA expression levels of liver inflammatory factors Tnfa, Ifng, and Ccl2; (B) immunohistochemical staining results of mouse liver CD8+; (C) flow cytometry gate ratio of liver CD8+ T cells (n=3); (D) flow cytometry gate ratio of liver Ly6c Hi cells; Compared with the normal control group, *P<0.05, **P<0.01, ***P<0.001; Compared with the model group, # P<0.05, ## P<0.01, ### P<0.001.
[0025] Figure 4 This is a PCA diagram from the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0026] Figure 5 This is a bar chart showing the differential gene expression of MOD vs NC and ACS vs MOD in the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0027] Figure 6 This is a differential expression volcano plot of the MOD vs NC groups in the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0028] Figure 7 This is a volcano plot showing the differential gene expression between the ACS and MOD groups in the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0029] Figure 8 This is a Venn diagram showing the differential gene expression of MOD vs NC and ACS vs MOD in the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0030] Figure 9 This is a heatmap of gene expression intersections among the NC, MOD, and ACS groups in the transcriptome sequencing differential gene expression analysis of ICI-induced liver inflammation by ACS.
[0031] Figure 10 This is a bubble chart of GO functional enrichment analysis in the transcriptome sequencing differential gene enrichment analysis of ICI-induced liver inflammation by ACS.
[0032] Figure 11 This is a bubble chart showing the enrichment of the KEGG pathway in the transcriptome sequencing differential gene enrichment analysis of ICI-induced liver inflammation by ACS.
[0033] Figure 12 This is a graph showing the differential gene enrichment analysis of the cGAS-STING pathway and the NOD-like receptor pathway GSEA in the transcriptome sequencing of ACS for ICI-induced liver inflammation.
[0034] Figure 13 This is a gene expression box plot from the transcriptome sequencing differential gene enrichment analysis of ICI-induced liver inflammation by ACS.
[0035] Figure 14 The expression levels of hepatic Cxcl10 and Rsad2 mRNA in the transcriptome sequencing differential gene enrichment analysis of ICI-induced liver inflammation by ACS were *P<0.05, **P<0.01, ***P<0.001 compared with the normal control group; compared with the model group, # P<0.05, ## P<0.01, ### P<0.001. Detailed Implementation
[0036] The following disclosure provides many different implementations or examples for different ways of implementing the present invention.
[0037] Example 1
[0038] 1. Laboratory animals and grouping
[0039] This embodiment sets up the following 5 groups for experiments:
[0040] Normal control group (NC group): Wild-type C57BL / 6 mice were given solvent control.
[0041] Model group (Mod group): Pdcd1- / - C57BL / 6 mice (i.e., Pdcd1 knockout C57BL / 6 mice) were used to construct an ICIs liver injury model.
[0042] Dexamethasone positive control group (DXM group): Pdcd1- / - C57BL / 6 mice were given dexamethasone (3 mg / kg).
[0043] ACS low-dose group (ACS-L group): Pdcd1- / - C57BL / 6 mice were given total saponins of *Abrus precatorius* (75 mg / kg).
[0044] High-dose ACS group (ACS-H group): Pdcd1- / - C57BL / 6 mice were given total saponins of *Abrus precatorius* (120 mg / kg).
[0045] ACS preparation: Extraction of total saponins from *Abrus precatorius* according to the established process. [1] Further purification yielded total saponins with higher purity. LC-MS analysis showed that the saponin purity was 61.25%, with the top five saponin monomers being absinin F, daidzein I, dehydro-daidzein I, sophoraecin III, and absinin SO1.
[0046] References: [1] Yao Xiangcao. Extraction process and anti-HBV activity and mechanism of action of total saponins of chicken bone grass [D]. Guangdong: Southern Medical University, 2019.
[0047] 2. Modeling and Dosing Regimen
[0048] The experiment lasted for 2 weeks (14 days), and the experimental protocol was as follows: Figure 1 As shown:
[0049] Week 1 (preventive administration period): The treatment group (ACS-L, ACS-H) and the positive control group were given the corresponding drugs such as ACS and dexamethasone for pretreatment; the normal control group and the model group were given the same amount of carrier solution.
[0050] Week 2 (Coexistence of Modeling and Treatment): Continue administration according to the above protocol. Simultaneously, except for the normal control group, mice in all other groups were given anti-CTLA4 antibody combined with ODN1668 to induce liver injury; the normal control group was treated with an equal amount of PBS combined with ODN1668.
[0051] Sample collection: After 2 weeks of drug administration, mice were anesthetized with isoflurane, samples were collected, and the mice were sacrificed. Serum biochemistry, HE staining, enzyme-linked immunosorbent assay (ELISA), immunohistochemical staining, and flow cytometry were used to evaluate the efficacy of ACS intervention in treating ICI-related liver injury. The mechanism of action of ACS was explored using Western blot and RT-qPCR.
[0052] 3. Experimental Results
[0053] 3.1 Improvement of the pathological state of ICI-related liver injury by ACS treatment
[0054] The degree of liver damage was assessed by detecting serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and HE-stained tissue sections. Figure 2 ).
[0055] AST level: such as Figure 2 As shown in Figure A, on day 15 of modeling, the AST level in the model group (94.00±19.08) was significantly higher than that in the normal control group (43.09±3.94), indicating that the liver injury model was successfully established. Compared with the model group, the AST levels in the positive drug dexamethasone group (70.03±6.40) and the high-dose ACS group (79.56±13.60) were significantly lower.
[0056] Changes in ALT levels: such as Figure 2 B and Figure 2 As shown in C, due to differences in baseline ALT levels among groups, the degree of ALT increase (D15-D0 difference) was analyzed for evaluation. The results showed ( Figure 2 (D) The increase in ALT in the model group (48.41±14.43) was significantly higher than that in the normal control group (14.64±3.21). After drug intervention, the increase in ALT in the dexamethasone group (19.33±12.45), ACS-L group (24.22±8.90), and ACS-H group (27.35±9.63) was significantly reduced compared with the model group. Figure 2 E shows the overall trend of serum ALT changes.
[0057] HE staining results: as shown Figure 2As shown in Figure F, the normal control group showed no immune infiltration around the central vein and intact hepatic cord structure; the model group showed extensive immune infiltration in the liver, with a higher density around the central vein and damaged hepatic cord structure; the positive dexamethasone group showed less immune infiltration and no obvious structural damage to the liver; the ACS-L and ACS-H groups showed reduced immune infiltration and smaller hepatocyte damage foci compared to the model group, but the number of immune cells in the hepatic sinusoids was still higher than in the normal control group. Blood biochemistry results and pathological section results showed that the mice had liver damage, which improved after ACS treatment.
[0058] 3.2 Improvement of inflammation levels caused by ICIs liver injury by ACS treatment
[0059] Clinically, ICI-induced liver injury is characterized by an excessive inflammatory response and extensive infiltration of CD8+ T cells in the liver, leading to liver damage. Therefore, we assessed the liver inflammation status and the role of ACS treatment in an animal model of ICI-induced liver injury by detecting the expression levels of inflammatory factors and the degree of immune cell infiltration.
[0060] The results are as follows Figure 3 As shown. The mRNA expression levels of Tnfa, Ifng, and Ccl2 in the model group (see...) Figure 3 A) Both were significantly elevated compared to the normal control group, suggesting the occurrence of liver inflammation. Figure 3 B showed extensive CD8+ T cell infiltration in the liver injury lesion. Flow cytometry detected Ly6C hi pro-inflammatory monocytes (...). Figure 3 D) and CD8+ T cells ( Figure 3 C) The proportion of liver infiltration in the model group was significantly higher than that in the normal control group, indicating that ICIs successfully induced hepatic immune accumulation. After ACS treatment, the expression levels of inflammatory factors, CD8+ T cell infiltration, and the levels of Ly6C hi pro-inflammatory monocytes and CD8+ T cells in the liver were all improved, and the proportion of Ly6C hi pro-inflammatory monocytes in the liver was significantly lower in the ACS treatment group than in the model group.
[0061] 3.3 Mechanism study of ACS treatment for ICI-related liver injury
[0062] To further investigate the mechanism by which ACS plays a role in ICI-related liver injury, we performed liver transcriptomics analysis, and the results are as follows: Figures 4-14 As shown.
[0063] exist Figure 4 PCA analysis revealed a clear separation between the control and model groups, with the ACS group falling between the control and model groups, suggesting that ACS intervention can reverse the expression of pathogenic genes. Differential gene screening was performed using logFC ≤ 0.585 and Padj < 0.05 as criteria.
[0064] The results are as follows Figure 5 As shown, there were 4103 differentially expressed genes in the MOD group versus the NC group, of which 2516 were upregulated and 1587 were downregulated; there were 721 differentially expressed genes in the ACS group versus the MOD group, of which 268 were upregulated and 453 were downregulated.
[0065] like Figure 6 The chart shows the differentially expressed genes between the MOD and NC groups, presented as a volcano plot. Among the 4103 differentially expressed genes, the top ten, sorted by Padj (corrected p-value) from smallest to largest, are Gm42031, Dbp, Wdfy1, Tmem254, Gbp2, Tef, Tlr2, Cd44, Npas2, and Cybb. Dbp, Tmem254, and Tef are downregulated, while Gm42031, Wdfy1, Gbp2, Tlr2, Cd44, Npas2, and Cybb are upregulated.
[0066] like Figure 7 The chart shows differentially expressed genes between the ACS and MOD groups using a volcano plot. Among the 721 differentially expressed genes, the top ten, sorted by Padj (corrected p-value) from smallest to largest, are Cyp2a4, Cyp2b9, Cyp3a41a, Xist, Mbl1, Cadm4, Cyp4a32, Cyp2b13, Slc22a27, and Serpina11. Cyp2a4, Cyp2b9, Cyp3a41a, Xist, Cyp2b13, and Slc22a27 are downregulated, while Mbl1, Cadm4, Cyp4a32, and Serpina11 are upregulated.
[0067] like Figure 8 The results show the intersection of differentially expressed genes between the MOD group and the NC group with those between the ACS group and the MOD group, yielding a total of 533 differentially expressed genes. Additionally, 188 differentially expressed genes are specific to the ACS group vs. MOD group, and 3570 genes are specific to the MOD group vs. NC group.
[0068] like Figure 9 The image shows a heatmap illustrating the intersection of differentially expressed genes between the MOD group and the NC group, and between the ACS group and the MOD group, comparing the expression differences of each gene between the groups. Cluster analysis was performed on each sample for comparison. The results show good clustering and small intra-group differences. Different colors were used to indicate gene upregulation / downregulation between groups: red for upregulation and blue for downregulation. The image shows that genes downregulated in the MOD group compared to the NC group tended to be upregulated again after ACS treatment. Conversely, genes upregulated in the MOD group compared to the NC group tended to be downregulated again after ACS treatment. This indicates that ACS treatment can reverse the upregulation / downregulation of pathogenic genes in the MOD group.
[0069] To investigate how ACS reverses excessive liver inflammation induced by ICI, the intersection of differentially expressed genes from the two groups was analyzed, and the results are as follows: Figure 10 As shown. First, GO functional enrichment analysis was performed, revealing that the overlapping genes were concentrated in inflammatory response functions such as cellular response to interferon β, positive regulation of defense responses, and responses to stimuli. For example... Figure 11 As shown, KEGG enrichment analysis mainly focused on inflammatory pathways such as the Toll-like receptor pathway, NK cell-mediated cytotoxicity, T cell receptor pathway, NOD-like receptor pathway, and cytoplasmic DNA sensor pathway. Based on the results of GO functional enrichment and KEGG pathway enrichment analysis, the cytoplasmic DNA sensor pathway, namely the cGAS-STING pathway, was found to play an important role in the action of ACS. Therefore, we used GSEA analysis to determine the status of the pathway, and the results are as follows. Figure 12 As shown, the cGAS-STING pathway was inhibited after ACS treatment. We first analyzed the expression levels of its downstream factors (Cxcl10, Rsad2, Ifit1) using transcriptome expression data, such as... Figure 13 As shown, the expression levels of Cxcl10, Rsad2, and Ifit1 in the model group were significantly higher than those in the control group, while those in the ACS group were significantly lower than those in the model group. To further verify the role of ACS in the cGAS-STING pathway, we used qPCR to analyze the mRNA expression levels of Cxcl10 and Rsad2 in the liver. The results are as follows. Figure 14 As shown, the expression levels of Cxcl10 and Rsad2 in the model group were significantly higher than those in the control group, and decreased after treatment with high and low doses of ACS. The expression level of Cxcl10 in the high-dose ACS group showed a statistically significant difference. Therefore, we preliminarily conclude that ACS can improve liver inflammation by inhibiting the activation of the cGAS-STING pathway.
[0070] In summary, prophylactic administration of total saponins from *Abrus precatorius* can alleviate the excessive level of liver inflammation and the pathological state of liver damage caused by ICI by regulating the cGAS-STING pathway.
[0071] The above is a further detailed description of the present invention and should not be considered as a limitation on the specific implementation of the present invention. For those skilled in the art, simple deductions or substitutions without departing from the concept of the present invention are all within the protection scope of the present invention.
Claims
1. Application of total saponin extract of *Abrus precatorius* in the preparation of drugs for treating liver damage caused by immune checkpoint inhibitors.
2. Application of total saponin extract of *Abrus precatorius* in the preparation of drugs for preventing liver damage caused by immune checkpoint inhibitors.
3. The application according to claim 1 or 2, characterized in that, The immune checkpoint inhibitor is selected from anti-PD-L1 antibody, anti-PD-1 antibody, anti-CTLA-4 antibody, and combinations thereof.
4. The application according to claim 1 or 2, characterized in that, The total saponin extract of *Abrus precatorius* contains daidzein I, sophora japonica saponin III, dehydrodaidzein I, absinthecin F, and absinthecin SO1.
5. The application according to claim 1 or 2, characterized in that, The drug comprises an effective amount of total saponin extract of *Abrus precatorius* and a pharmaceutically acceptable carrier or excipient.
6. The application according to claim 1 or 2, characterized in that, The dosage form of the drug is oral liquid, pill, granule, capsule, tablet, drop pill, injection or powder.
7. The application according to claim 1 or 2, characterized in that, The total saponin extract of *Abrus precatorius* is an organic solvent extract of total saponins from *Abrus precatorius*.
8. The application according to claim 7, characterized in that, The organic solvent extract of total saponins from *Abrus precatorius* is selected from the n-butanol extract of total saponins from *Abrus precatorius*.