Application of neuropeptide Y1 receptor antagonist in preparation of hereditary polycystic kidney disease treatment medicine

Abstract

The invention relates to the technical field of medicines, in particular to application of a neuropeptide Y1 receptor antagonist in preparation of a hereditary polycystic kidney disease treatment medicine. The neuropeptide Y1 receptor antagonist is selected from a compound BIBO3304 which plays a role in blocking combination of neuropeptide Y (NPY) and a Y1 receptor subtype thereof. On the basis that early-stage histopathology, high-throughput sequencing, cytobiology and molecular biology are combined with in-vitro cell culture and in-vivo animal experiments, the potential treatment effect of the neuropeptide Y1 receptor antagonist in preparation of hereditary polycystic kidney disease treatment drugs is provided, and a new basis is provided for hereditary polycystic kidney disease treatment.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to the application of neuropeptide Y1 receptor antagonists in the preparation of drugs for the treatment of hereditary polycystic kidney disease. Background Technology

[0002] Autosomal dominant polycystic kidney disease (ADPKD) is a disease characterized by multiple fluid-filled vesicles in both kidneys, with an incidence of 1 / 1000 to 1 / 400. It is the most common monogenic inherited kidney disease. ADPKD is the fourth leading cause of end-stage renal disease (ESRD), with approximately half of patients progressing to ESRD by age 60. Clinically, ADPKD is primarily treated symptomatically, and its natural course cannot be altered. Therefore, exploring new treatment methods remains a key focus and challenge in current ADPKD research.

[0003] Most cases of ADPKD are caused by mutations in the PKD1 or PKD2 genes, accounting for 86% and 10% of ADPKD cases, respectively. The expression products of the PKD1 and PKD2 genes are called polycystin 1 (PC1) and polycystin 2 (PC2), respectively. PC1 and PC2 are mainly located on the primary cilia of cells. Primary cilia are organelles present on the surface of most cells and play a cellular sensory role in promoting cell-environment interaction. Therefore, polycystic kidney disease is considered a ciliary-related disease, and abnormalities in the structure or function of cilia play an important role in the formation and development of renal cysts. PC1 acts as a mechanosensor, and PC2 is a calcium-regulated non-selective cation channel; together, they regulate intracellular calcium ion concentration. Defective ciliary-polycystin complexes in renal tubular epithelial cells cannot sense fluid flow, which may reduce cellular calcium ion influx, thereby stimulating cAMP signaling and leading to cyst growth. However, the specific regulatory mechanisms of ADPKD occurrence and development remain unclear.

[0004] Neuropeptide Y (NPY) is a hormone widely distributed in the central and peripheral nervous systems, maintaining homeostasis. NPY's effects are primarily achieved through binding to its receptor. The small molecule drug BIBO3304, as a specific neuropeptide Y1 receptor antagonist, can block the series of reactions caused by the binding of NPY to its Y1 receptor subtype.

[0005] Currently, there are no reports on the relationship between NPY and the treatment of hereditary polycystic kidney disease. Summary of the Invention

[0006] The purpose of this invention is to address the above-mentioned challenges in the treatment of hereditary polycystic kidney disease by providing new pharmaceutical applications for neuropeptide Y1 receptor antagonists, and to provide the application of NPY as a target in the treatment of hereditary polycystic kidney disease. Specifically, it provides the application of neuropeptide Y1 receptor antagonists in the preparation of drugs for the treatment of hereditary polycystic kidney disease.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] Firstly, the study found that the expression of NPY in the lining epithelial cells of polycystic kidney disease patients was significantly upregulated. A polycystic kidney disease mouse model was constructed, and administration of a neuropeptide Y1 receptor antagonist significantly reduced the kidney-to-body ratio and cystic index in mice, with no significant side effects observed in the mouse model group. This invention discloses the mechanism of action of neuropeptide Y1 receptor antagonists in inhibiting polycystic kidney disease, and for the first time, reveals that BIBO3304 significantly inhibits the kidney-to-body ratio and cystic index in polycystic kidney disease mice, providing a novel perspective and target for inhibiting the progression of polycystic kidney disease.

[0009] Based on the above technical solutions, the first aspect of the present invention provides the application of a neuropeptide Y1 receptor antagonist in the preparation of a therapeutic drug for hereditary polycystic kidney disease.

[0010] Furthermore, the neuropeptide Y1 receptor antagonist is a substance that blocks the binding of neuropeptide Y (NPY) to its Y1 receptor subtype.

[0011] Furthermore, the hereditary polycystic kidney disease mentioned above is autosomal dominant polycystic kidney disease (ADPKD).

[0012] Furthermore, the aforementioned hereditary polycystic kidney disease treatment drug is a drug that reduces the volume and cystic index of polycystic kidneys.

[0013] Furthermore, the hereditary polycystic kidney disease treatment drug is a pharmaceutical composition consisting of a neuropeptide Y1 receptor antagonist as the sole active monomer component, or a neuropeptide Y1 receptor antagonist used in combination with other drugs.

[0014] Furthermore, the drug also includes pharmaceutically acceptable carriers or excipients.

[0015] Furthermore, the neuropeptide Y1 receptor antagonist is BIBO3304, with the chemical formula C. 29 H 35 N7O3 has the following structural formula, as shown in Formula I:

[0016]

[0017] Formula I

[0018] Animal model experiments of the present invention show that after administration of the neuropeptide Y1 receptor antagonist BIBO3304, the kidney-to-body ratio and cystic index of polycystic kidney mouse models were significantly reduced.

[0019] Furthermore, the concentration of BIBO3304 in the hereditary polycystic kidney disease treatment drug is 100-250 μM, preferably 160 μM.

[0020] In a second aspect, the present invention provides a pharmaceutical composition for treating hereditary polycystic kidney disease, comprising a neuropeptide Y1 receptor antagonist as the sole active monomeric component, or a neuropeptide Y1 receptor antagonist used in combination with other drugs.

[0021] The advantages of this invention are:

[0022] Based on previous in vitro and in vivo experiments combining histopathology, cell biology, and molecular biology, this invention proposes the potential therapeutic effect of the neuropeptide Y1 receptor antagonist BIBO3304 on hereditary polycystic kidney disease, thus demonstrating that BIBO3304 can be used to prepare a therapeutic drug for hereditary polycystic kidney disease. Therefore, this invention provides new evidence for the treatment of hereditary polycystic kidney disease. Attached Figure Description

[0023] Figure 1 Results of tissue immunofluorescence analysis of NPY protein.

[0024] Figure 2 NPY mRNA and protein expression levels in the lining epithelial cells of the polycystic kidney mouse model: A, Fluorescence in situ hybridization analysis of NPY mRNA; B, Immunofluorescence staining analysis of NPY protein in endometrial tissue.

[0025] Figure 3 The neuropeptide Y1 receptor antagonist BIBO3304 downregulated the kidney-to-body ratio and cystic index in a polycystic kidney mouse model: A, HE staining analysis; B, cystic index analysis results. Detailed Implementation

[0026] The specific implementation methods provided by the present invention will be described in detail below with reference to the embodiments.

[0027] Example 1: Collection of human kidney tissue

[0028] 1) Study Groups: Referring to the "Chinese Clinical Practice Guidelines for Autosomal Dominant Polycystic Kidney Disease (Second Edition)" (Journal of Clinical Nephrology, 2019, Vol. 4), the study subjects were divided into:

[0029] ① Normal control group (Control): Patients with other kidney diseases but without polycystic kidney disease (peripheral tissue of renal cell carcinoma);

[0030] ② Polycystic kidney disease group (ADPKD): Patients with polycystic kidney disease;

[0031] Study participants were those who had not used systemic or local hormones within one month prior to surgery; patients with systemic diseases such as immunodeficiency, neurological or psychiatric disorders were excluded. All enrolled patients signed informed consent forms.

[0032] 2) Sample Collection: Kidney tissues were collected from patients in the Control and ADPKD groups for later research use; all collected samples were uniformly numbered and managed. Tissue samples were fixed in 4% paraformaldehyde for immunofluorescence detection.

[0033] The expression of NPY in ADPKD was detected using human kidney tissue and tissue immunofluorescence. Figure 1 The study showed that NPY protein expression levels were significantly upregulated in the cyst lining epithelial cells of patients with polycystic kidney disease. Figure 2 The study showed that NPY mRNA and protein expression levels were significantly upregulated in the cyst lining epithelial cells of a polycystic kidney mouse model.

[0034] Experimental data showed that the expression level of neuropeptide Y protein was significantly upregulated in the ADPKD group compared with the normal control group.

[0035] Example 2: Animal Experiment

[0036] Pkd1 was obtained through gene editing using CRISPR-Cas9 technology. L3984R Heterozygous mutant mice (Wang, H., Yang, H., Shivalila, CS, Dawlaty, MM, Cheng, AW, Zhang, F., &Jaenisch, R. (2013). One-step generation of mice carrying mutations in multiple genes by CRISPR / Cas-mediated genome engineering. Cell, 153(4), 910–918.) were then bred together to obtain homozygous mice, thus establishing an ADPKD mouse model. One group of ADPKD mice was treated with the neuropeptide Y1 receptor antagonist BIBO3304, while the other group received an equal volume of saline as a control. Changes in the overall bilateral renal appearance, kidney weight-to-body weight ratio, and renal cystic index, as well as the mechanism of action, were examined in ADPKD mice.

[0037] ADPKD mouse model administration method:

[0038] By Pkd1 L3984RThe ADPKD mouse model established by homozygous mutations exhibits early phenotype (observable even in newborn mice) and progresses rapidly (dying at four weeks of age). Direct drug administration using traditional methods is not feasible; therefore, we adopted an indirect intervention approach by gavage of the mother mice: Pkd1 L3984R Heterozygous mutant mice were housed together, and the presence or absence of vaginal plugs was used to determine successful mating. Successfully mated females were administered medication via gavage (300 μL of 160 μM BIBO3304 per mouse) once daily from day 11.5 of gestation until weaning (three weeks of age), for a total of 30 days. Genotyping was performed on the mice seven days after birth to determine their genotype. Pkd1 mice were selected three weeks after birth. L3984R Mutant homozygous mice were euthanized by overdose anesthesia, and kidney specimens were collected, weighed, and the kidney weight to body weight ratio was calculated. A portion of the kidney specimens were subjected to routine hematologic and epithelial studies to observe the histopathological condition; the other portion of kidney tissue was collected and stored at -80°C for subsequent RT-qPCR, Western blotting, and other experiments.

[0039] The results showed that the bilateral kidney size of ADPKD mice in the experimental group ( Figure 3 A) and kidney weight to body weight ratio ( Figure 3 B) were significantly lower than the control group. After administration of the neuropeptide Y1 receptor antagonist BIBO3304, the cystic index of the kidneys in the ADPKD mouse model ( Figure 3 C and D) were significantly reduced.

[0040] Conclusion: The neuropeptide Y1 receptor antagonist BIBO3304 has a therapeutic effect on hereditary polycystic kidney disease, and a therapeutic drug for hereditary polycystic kidney disease can be prepared.

[0041] The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.

Claims

1. Application of neuropeptide Y1 receptor antagonists in the preparation of drugs for the treatment of hereditary polycystic kidney disease.

2. The application of the neuropeptide Y1 receptor antagonist according to claim 1 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The hereditary polycystic kidney disease mentioned is autosomal dominant polycystic kidney disease.

3. The application of the neuropeptide Y1 receptor antagonist according to claim 1 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The neuropeptide Y1 receptor antagonist is a substance that blocks the binding of neuropeptide Y to its Y1 receptor subtype.

4. The application of the neuropeptide Y1 receptor antagonist according to claim 1 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The aforementioned medication for treating hereditary polycystic kidney disease is a drug that reduces the volume and cystic index of polycystic kidneys.

5. The application of the neuropeptide Y1 receptor antagonist according to claim 1 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The aforementioned hereditary polycystic kidney disease treatment drug is a pharmaceutical composition consisting of a neuropeptide Y1 receptor antagonist as the sole active monomer component, or a neuropeptide Y1 receptor antagonist used in combination with other drugs.

6. The application of the neuropeptide Y1 receptor antagonist according to claim 5 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The drug also includes pharmaceutically acceptable carriers or excipients.

7. The application of the neuropeptide Y1 receptor antagonist according to claim 1 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The neuropeptide Y1 receptor antagonist mentioned is BIBO3304.

8. The application of the neuropeptide Y1 receptor antagonist according to claim 7 in the preparation of a therapeutic drug for hereditary polycystic kidney disease, characterized in that, The concentration of BIBO3304 in the hereditary polycystic kidney disease treatment drug is 100-250 μM.

Images