CN114588152A - Use of PU.1 inhibitor DB2313 - Google Patents
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- NUVPJXUYFGWDGB-UHFFFAOYSA-N 2-[4-[[2-fluoro-3-[[4-[6-(N'-propan-2-ylcarbamimidoyl)-1H-benzimidazol-2-yl]phenoxy]methyl]phenyl]methoxy]phenyl]-N'-propan-2-yl-3H-benzimidazole-5-carboximidamide Chemical compound CC(C)NC(=N)C1=CC2=C(C=C1)N=C(N2)C1=CC=C(OCC2=CC=CC(COC3=CC=C(C=C3)C3=NC4=C(N3)C=C(C=C4)C(=N)NC(C)C)=C2F)C=C1 NUVPJXUYFGWDGB-UHFFFAOYSA-N 0.000 title claims abstract description 44
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
- A61K31/4164—1,3-Diazoles
- A61K31/4184—1,3-Diazoles condensed with carbocyclic rings, e.g. benzimidazoles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
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- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Life Sciences & Earth Sciences (AREA)
- General Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Organic Chemistry (AREA)
- Rheumatology (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Epidemiology (AREA)
- Pain & Pain Management (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Physical Education & Sports Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention discloses an application of a PU.1 inhibitor DB2313, and discloses a pharmaceutical preparation prepared by using DB2313 as an active ingredient, wherein the pharmaceutical preparation can simultaneously realize immune function regulation and inflammatory symptom improvement. The research of the invention proves that the compound DB2313 can effectively improve the toe swelling degree and the arthritis index of a mouse with collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA), restore the immune balance, reduce the phenomena of synovial membrane hyperplasia, inflammatory cell infiltration and cartilage and bone destruction, reduce the blood flow signal of knee joints, improve the inflammatory symptoms of disease models, and can be used as a potential therapeutic drug for rheumatoid arthritis.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to application of a PU.1 inhibitor DB2313 which can be used as a potential treatment medicine for rheumatoid arthritis.
Background
Rheumatoid Arthritis (RA) is a chronic inflammatory and destructive disease of the joints characterized by synovial hyperplasia and infiltration of different cell types into the synovial membrane of the joints. Currently, there is no cure method for RA, but clinical manifestations can be improved to a certain extent by non-steroidal anti-inflammatory drugs, disease-improving antirheumatic drugs, biological agents and other drugs, but most of the drugs cause serious adverse reactions after long-term application. Therefore, searching for a new key mechanism for RA attack provides experimental basis for the research and development of novel therapeutic drugs with good curative effect and less adverse reactions. PU.1 is a member of the E26 transformation-specific transcription factor family and is essential for the differentiation and function of various bone marrow cells. The important role of pu.1 in immune cells prompted us to explore its role in autoimmune diseases, especially RA. The research result of DB2313 provides a new thought and a potential therapeutic target for the pathogenesis of RA.
Disclosure of Invention
The invention aims to provide application of a PU.1 inhibitor DB 2313. The research of the invention proves that the compound DB2313 can effectively improve the toe swelling degree and the arthritis index of a mouse with collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA), restore the immune balance, reduce the phenomena of synovial membrane hyperplasia, inflammatory cell infiltration and cartilage and bone destruction, reduce the blood flow signal of knee joints, improve the inflammatory symptoms of disease models, and can be used as a potential therapeutic drug for rheumatoid arthritis.
The application of the PU.1 inhibitor DB2313 is to prepare a medicinal preparation by taking DB2313 as an active ingredient, and the medicinal preparation can realize immune function regulation and inflammatory symptom improvement at the same time and can be used as a treatment medicament for rheumatoid arthritis.
The medicine comprises one or more of but not limited to serum products, vaccines, diagnostic products, chemical medicines and biochemical medicines.
CAS number of small molecule inhibitor DB 2313: 2170606-74-1, the structure is as follows:
the present invention uses DB2313 in the treatment of collagen-induced arthritis (CIA) and collagen anti-induced arthritis (CAIA) mice to determine the potential of DB2313 as a potential drug for the treatment of RA and to explore its mechanism of action.
In order to develop research on rheumatoid joint treatment medicines, the invention establishes a CIA model and a CAIA model. After the DB2313 is used for treating, the toe swelling degree and the arthritis index of a CIA mouse and a CAIA mouse can be effectively improved, the immune balance is restored, synovial membrane hyperplasia, inflammatory cell infiltration, cartilage and bone destruction phenomena are reduced, the blood flow signal of the knee joint is reduced, and the inflammation of a disease model is improved, so that the inhibition effect on the immunity and the inflammation of the CIA mouse is shown, the medicine is a treatment medicine capable of treating both symptoms and root causes, and has a good development prospect.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this specification, are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification, illustrate exemplary embodiments of the invention and together with the description serve to explain the invention and not to limit the invention.
FIGS. 1-5 show the effect of DB2313 on mouse CIA; wherein FIG. 1 is a general observation of the swelling of the joints of groups of mice 21 days after DB2313 treatment, showing that the swelling morphology of the anterior and posterior paws of CIA mice is improved by DB2313 treatment; fig. 2 is an inflammation curve plotted against the arthritis index, joint swelling volume. Shows that after the treatment of DB2313, the swelling volume of joints and the arthritis index of the CIA mice are reduced; FIG. 3 is a graph showing Doppler ultrasound results for groups of mice showing reduced knee blood flow signals in DB2313 compared to vehicle group, suggesting that synovial pannus formation is inhibited; FIG. 4 shows the HE staining results of knee joints of mice in each group, wherein synovial hyperplasia of knee joints and inflammatory cell infiltration are reduced in the DB2313 group compared with the vehicle group; fig. 5 shows Trap staining results of knee joints of mice in each group, and shows that knee joint osteoclastogenesis is reduced in the DB2313 group compared with the vehicle group, indicating that the effect on bone destruction is reduced.
FIGS. 6-10 show the effect of DB2313 on mouse CAIA; wherein FIG. 6 is a general observation of the swelling of the joints of various groups of mice 8 days after DB2313 treatment, showing that the swelling morphology of the anterior and posterior paws of CAIA mice is improved by DB2313 treatment; FIG. 7 is an inflammation curve plotted against the arthritis index, the degree of swelling of the joints, showing a decrease in the number of joint swelling and the arthritis index in CAIA mice treated with DB 2313; FIG. 8 is a graph showing Doppler ultrasound results for groups of mice showing a reduction in knee blood flow signals in DB2313 compared to vehicle group, suggesting that synovium pannus formation is inhibited; FIG. 9 shows the HE staining results of knee joints of mice in each group, wherein synovial hyperplasia of knee joints and inflammatory cell infiltration are reduced in the DB2313 group compared with the vehicle group; FIG. 10 shows the results of safranin fast green staining of knee joints of mice in each group, and the destruction of knee joint cartilage was reduced in DB2313 group compared to vehicle group.
Detailed Description
It is to be understood that the following detailed description is exemplary and is intended to provide further explanation of the invention as claimed. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the invention. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise.
As introduced in the background art, the current treatment of rheumatoid arthritis often requires a combination of therapeutic drugs of different mechanisms to achieve effective symptom relief, and the development of a single drug capable of improving both immune function and inflammatory symptoms can significantly reduce the medication burden of patients. In order to solve the technical problems, the invention provides application of a PU.1 inhibitor DB2313 in preparing an anti-rheumatoid arthritis product.
In order to make the technical solutions of the present invention more clearly understood by those skilled in the art, the technical solutions of the present invention will be described in detail below with reference to specific embodiments.
Materials and methods
1. Material
1.1 Experimental animals
SPF-grade DBA/1 mice (male, 7 weeks old, 18-20g, certification number: 2017-. SPF grade C57BL/6J mice (male, 7 weeks old, 18-20g) were purchased from Shanghai Spiker laboratory animals, LLC. All animals were housed in the SPF animal testing center, clinical pharmacology institute of university of medical, Anhui. All animal experiments were approved by the ethical review committee on animal experiments at the institute of clinical pharmacology, university of ammicro medical science.
2. Method of producing a composite material
2.1 DB2313 processing the CIA model
Chicken type II collagen was dissolved well in 5mL of acetic acid (0.1M) and then emulsified with complete Freund's adjuvant. On day 0, mice were injected intradermally with 0.15ml of the emulsion. On day 21, mice were re-stimulated by intradermal injection of 0.1mL CII emulsion. On day 29, the success rate of the CIA model was 95%, excluding the failed model. The mice were divided into three groups. After joint swelling occurred on day 29, mice were treated by intraperitoneal injection at 0.3mg/3 day. CIA model mice were treated with equal volume of vehicle (PBS). On day 59, mice were euthanized and blood and ankle joints were collected for follow-up studies.
2.2 DB2313 processing the CAIA model
On day 0, 4mg of 5-clone collagen antibody mixture was intraperitoneally injected, and on day 3, 40. mu.g of LPS was intraperitoneally injected. Mice were treated intravenously at 0.3 mg/day at d7, d9, d11, d13, d15, respectively. The control group was given an equal amount of physiological saline, and the number of joint swelling and the arthritis index were observed and recorded after daily injection. On day 18, mice were euthanized with an excess of anesthetic, and blood and ankle joints were collected for follow-up studies.
2.3 volume/number of joint swelling
And (3) CIA: the swelling volume of the joints of the mice in each group was recorded at d29, d32, d35, d38, d41, d44, d47 and d50 after molding.
CAIA: the number of swelling in the joints of each group of mice was recorded daily after molding. The scoring criteria were: the amount of swelling of the joints of each group of mice was recorded daily after molding. The mouse paw includes 5 finger joints and one ankle or wrist joint, so the maximum swollen joint count per paw is 6 and the maximum swollen joint count per mouse is 24.
2.4 arthritis index
And (3) CIA: the arthritis index of each group of mice was scored after molding d29, d32, d35, d38, d41, d44, d47, d 50.
CAIA: groups of mice were scored daily for arthritis index after molding.
The scoring criteria were on various scales from 0-4: 0 minute, local red swelling phenomenon does not occur; 1 minute, swelling of finger joints appears; slight swelling of ankle or wrist occurs in 2 minutes; 3 minutes, the whole paw has severe swelling phenomenon; for 4 minutes, the paw is stiff or deformed.
2.5, detecting the intensity of blood flow signals at the knee joint by adopting an ultrasonic Doppler mode, and judging the formation of pannus in the knee joint according to the area occupied by the Doppler color image.
2.6 the mouse is placed on an X-ray operation platform after anaesthetizing, the knee joint position of the mouse is close to the plane of the operation board, the X-ray software is opened, the parameters and the position are adjusted, and the picture is shot and stored.
2.7HE staining: mice were euthanized with an excess of anesthetic. After fixation and decalcification, the knee joint was dehydrated with gradient alcohol, embedded in paraffin, and sliced to 5 μm. The slices are dewaxed and hydrated by gradient alcohol, stained by hematoxylin and eosin, dehydrated by gradient alcohol and sealed by neutral glue. Histopathological changes were observed and recorded using an optical microscope.
2.8 Safranin O staining: the slices after dewaxing are soaked in safranin O staining solution for 3min, washed with distilled water for 1min, soaked in fast green staining solution for 2min, washed with distilled water for 1min, differentiated with 1% glacial acetic acid for 1min, dehydrated with 95% ethanol and decolorized with xylene. Finally, the slice is sealed by neutral glue and the morphological change of the slice is observed.
2.9 Trap staining: the substrate solution was prepared according to the instructions and placed in a 37 ℃ water bath for 10 min. Then, cells were mixed with the substrate solution, incubated for 1 hour, and counterstained with hematoxylin for 2-3 minutes. Then, the picture was obtained after fixing with neutral glue. The positive staining cytoplasm is deep red, and the nucleus is bluish purple.
Second, experimental results
1. Effect of DB2313 on CIA mice
The study first evaluated the overall therapeutic efficacy of DB2313, and the swelling morphology of the anterior and posterior paws of CIA mice was also improved by DB2313 treatment (fig. 1), with a reduction in the joint swelling volume and arthritis index (fig. 2). In addition, the formation of pannus of joints was also inhibited in the DB2313 group compared to the vector group (FIG. 3). The H & E staining results showed that DB2313 reduced synovial hyperplasia, inflammatory cell infiltration in the CIA model (fig. 4), and the TRAP staining results showed that DB2313 reduced osteoclastogenesis in the CIA model (fig. 5).
2. Effect of DB2313 on CAIA mice
The study first evaluated the overall therapeutic efficacy of DB2313, and the swelling morphology of the CAIA mice forepaws and hindpaws was also improved by DB2313 treatment (fig. 6), with a reduction in the number of joint swelling and arthritis index (fig. 7). In addition, the synovial pannus formation in joints was also inhibited in the DB2313 group compared to the vehicle group (FIG. 8). H & E staining results showed that DB2313 reduced synovial hyperplasia, inflammatory cell infiltration in the CIA model (fig. 9). The results of Safranin O staining showed that DB2313 reduced cartilage destruction in the CIA model (fig. 10).
Claims (2)
- Use of the pu.1 inhibitor DB2313, characterized by:DB2313 is used as an active ingredient for preparing a pharmaceutical preparation which can simultaneously realize the regulation of immune function and the improvement of inflammatory symptoms.
- 2. Use according to claim 1, characterized in that:the medicinal preparation is used as a medicament for treating rheumatoid arthritis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210391056.9A CN114588152A (en) | 2022-04-14 | 2022-04-14 | Use of PU.1 inhibitor DB2313 |
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| CN202210391056.9A CN114588152A (en) | 2022-04-14 | 2022-04-14 | Use of PU.1 inhibitor DB2313 |
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| CN114588152A true CN114588152A (en) | 2022-06-07 |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017223260A1 (en) * | 2016-06-23 | 2017-12-28 | Albert Einstein College Of Medicine, Inc. | Pu.1 inhibitors |
| CN109200055A (en) * | 2017-07-08 | 2019-01-15 | 上海中医药大学附属龙华医院 | A kind of drug and application thereof for treating rheumatoid arthritis |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017223260A1 (en) * | 2016-06-23 | 2017-12-28 | Albert Einstein College Of Medicine, Inc. | Pu.1 inhibitors |
| CN109200055A (en) * | 2017-07-08 | 2019-01-15 | 上海中医药大学附属龙华医院 | A kind of drug and application thereof for treating rheumatoid arthritis |
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