阅读说明：本技术 生物碱在制备破骨细胞分化抑制剂中的应用 (Application of alkaloid in preparing osteoclast differentiation inhibitor ) 是由 刘志红 黄丹娥 徐峻 谢黎炜 顾琼 郑双佳 许国焕 韩木兰 于 2020-06-12 设计创作，主要内容包括：本发明公开了生物碱在制备破骨细胞分化抑制剂中的应用。生物碱在制备破骨细胞分化抑制剂中的应用,所述的生物碱是化合物AP-123/40765213或AE-562/43462182,其结构式如式1所示。本发明通过试验发现,化合物AP-123/40765213和/或AE-562/43462182对破骨细胞的分化具有直接而显著地抑制作用,因此可以作为破骨细胞分化抑制剂类药物,用于治疗破骨细胞分化引起的各类疾病,如骨质疏松症。<Image he="306" wi="700" file="DDA0002537622990000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The invention discloses an application of alkaloid in preparing an osteoclast differentiation inhibitor. The application of alkaloid in preparing osteoclast differentiation inhibitor is provided, wherein the alkaloid is compound AP-123/40765213 or AE-562/43462182, and the structural formula is shown in formula 1. According to the invention, experiments show that the compounds AP-123/40765213 and/or AE-562/43462182 have direct and significant inhibitory effects on osteoclast differentiation, so that the compounds can be used as osteoclast differentiation inhibitor medicines and used for treating various diseases caused by osteoclast differentiation, such as osteoporosis.)

1. Application of alkaloid or pharmaceutical salt thereof in preparing osteoclast differentiation inhibitor

The alkaloid is compound AP-123/40765213 or AE-562/43462182, and the structural formula is shown as formula 1:

2. the use according to claim 1, wherein the osteoclast differentiation inhibitor is an anti-osteoporotic agent.

3. The use according to claim 1, wherein the osteoclast differentiation inhibitor is in an oral dosage form or an external dosage form.

4. The use according to claim 1, 2 or 3, wherein the osteoclast differentiation inhibitor comprises the active ingredient compounds AP-123/40765213 and/or AE-562/43462182, or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable pharmaceutical excipients.

Technical Field

The invention belongs to the field of biological medicines, and particularly relates to an application of alkaloid in preparation of an osteoclast differentiation inhibitor.

Background

Osteoporosis is a systemic bone metabolic disease, and with the increase of the life span of human beings and the increase of the aging speed of population, the incidence rate of osteoporosis is increased year by year, so that osteoporosis becomes one of important chronic diseases affecting the quality of life of people and becomes a global public health problem. According to statistics on the annual meeting of the American Society for Bone and Mineral Research (ASBMR) in 2014, osteoporotic fractures are one of the important chronic diseases in the 21 st century, and are expected to occur in 1/2-old women and 1/3-old men worldwide. Therefore, the development of effective anti-osteoporosis drugs has important significance for improving the life quality of human beings, particularly the old, and relieving the social and economic pressure.

Osteoporosis is a metabolic bone disease characterized by decreased bone mass, destruction of bone tissue microstructure, increased bone fragility, and susceptibility to fracture. The action mechanisms of the current anti-osteoporosis drug therapy are mainly divided into two main categories: anti-bone resorption and promoting bone formation. The target or pathway involved in anti-bone resorption mainly comprises OPG/RANKL/RANK signal pathway and cathepsin K, C-src kinase. Relevant targets or pathways for promoting bone formation comprise Wnt/beta-catenin signal pathway, bone morphogenetic protein, Activin signal pathway and parathyroid hormone PTH. At present, the clinical bone absorption resistant drugs mainly comprise bisphosphonates (alendronate sodium, ibandronate sodium and the like), estrogen receptor modulators (raloxifene), RANKL inhibitors (denosumab), cathepsin K inhibitors (Orobatinib) and src kinase inhibitors (Saracatinib). The bone formation promoting drugs mainly include parathyroid hormone analogs (teriparatide PTH1-34), Wnt signaling pathway modulators (AMG785, BHQ880), and the like. Although these drugs can increase the bone density of patients with osteoporosis to some extent, they do not significantly reduce the risk of atypical fractures, and have various side effects, such as the bisphosphonate class of drugs causing necrosis of the mandible and raloxifene causing venous embolism. Therefore, there is a need to develop more effective anti-osteoporosis therapeutic agents with fewer side effects.

The OPG/RANKL/RANK signal system is confirmed to be an important signal path in bone metabolism, and RANKL (receptor activator of NF kB ligand) is combined with RANK on the surface of osteoclast precursor cells or osteoclasts to promote differentiation and activation of the osteoclasts and inhibit apoptosis of the osteoclasts; osteoprotegerin (OPG) prevents binding of RANKL to RANK, acting as a negative regulator. The dynamic balance of RANKL and OPG ratios plays an important controlling role in bone resorption. The research result of the OPG-RANKL-RANK signal system is a milestone in bone molecule biology, not only clarifies the coupling of various signals existing between the OPG-RANKL-RANK signal system and the development and maturation of osteoclasts, but also the coupling imbalance between the bone absorption of osteoclasts and the successful cell osteogenesis, lays a certain foundation for the occurrence mechanism of osteoporosis, and opens up a new idea for the targeted treatment of the osteoporosis. Currently, in the OPG-RANKL-RANK system, America ANIN has developed a monoclonal antibody drug, denosumab, for the prevention of bone-related events in patients with solid tumor bone metastases, and is also being studied for the treatment of osteoporosis. With the analysis of the crystal structure of the OPG-RANKL-RANK, the anti-osteoporosis small-molecule polypeptide drug targeting the OPG-RANKL-RANK becomes a research hotspot. Thus, targeting OPG-RANKL-RANK has been widely demonstrated to be an important strategy for anti-osteoporosis therapy.

The invention content is as follows:

the invention aims to provide application of alkaloid in preparing an osteoclast differentiation inhibitor.

The invention discovers that the compounds AP-123/40765213 and AE-562/43462182 can inhibit the mature differentiation of osteoclast in a dose-dependent manner (figure 1 and figure 2), and the IC50 value is 32nM and 68nM respectively (figure 3). Cytotoxicity test CC50 was 26.85 μ M and 19.93 μ M, respectively (fig. 4). Indicating that the two compounds have good inhibitory activity. Further, by testing the effect of the compounds on the expression of osteoclast-associated genes, it was found that both compounds can significantly reduce gene expression of Ctsk, Nfatc1, tracep, Rank (fig. 5).

Thus, compounds AP-123/40765213 and/or AE-562/43462182, or pharmaceutically acceptable salts thereof, may be used in the preparation of an osteoclast differentiation inhibitor:

the structural formulas of the compounds AP-123/40765213 and AE-562/43462182 are shown in formula 1:

the osteoclast differentiation inhibitor is an anti-osteoporosis drug.

The osteoclast differentiation inhibitor is an oral preparation or an external preparation.

The osteoclast differentiation inhibitor comprises an active ingredient compound AP-123/40765213 and/or AE-562/43462182 and pharmaceutically acceptable pharmaceutic adjuvants.

According to the invention, experiments show that the compounds AP-123/40765213 and/or AE-562/43462182 have direct and significant inhibitory effects on osteoclast differentiation, so that the compounds can be used as osteoclast differentiation inhibitor medicines and used for treating various diseases caused by osteoclast differentiation, such as osteoporosis.

Description of the drawings:

FIG. 1 shows that compound AP-123/40765213 dose-dependently inhibits osteoclast maturation differentiation;

FIG. 2 is a dose-dependent inhibition of osteoclast maturation differentiation by Compound AE-562/43462182;

FIG. 3 is a graph of inhibition curves and IC50 values for compound AP-123/40765213 and AE-562/43462182;

FIG. 4 is a cytotoxicity assay of compound AP-123/40765213 with AE-562/43462182;

FIG. 5 shows the effect of compounds AP-123/40765213 and AE-562/43462182 on the expression of the gene involved in the mature differentiation of osteoclasts.

The specific implementation mode is as follows: