阅读说明：本技术 重楼皂苷ⅱ在制备增敏顺铂疗效药物中的应用 (Application of rhizoma paridis saponin II in preparing medicine with sensitization cisplatin curative effect ) 是由 高文远 满淑丽 吕盼盼 崔静霞 于 2021-09-02 设计创作，主要内容包括：本发明公开了重楼皂苷Ⅱ在制备增敏顺铂疗效药物中的应用,研究表明,在肺癌细胞株H446中,单独使用顺铂仅轻微降低细胞生存率,而顺铂联合重楼皂苷Ⅱ后使生存率大大降低。同时,通过两者多个浓度联合应用测定得到的联合指数CI低于1,表明重楼皂苷II能够增敏顺铂,提高其抗肿瘤疗效。中药重楼来源广泛,从该植物中提取分离出的重楼皂苷Ⅱ,经过比较详细的体外活性试验证明与一线化疗药顺铂合用,能有效地改善顺铂的耐药性,提高顺铂的抗肿瘤活性。(The invention discloses application of paris polyphylla saponin II in preparation of a sensitization cisplatin curative effect medicine, and researches show that in a lung cancer cell strain H446, the single use of cisplatin only slightly reduces the cell survival rate, and the cisplatin and the paris polyphylla saponin II greatly reduce the survival rate. Meanwhile, the combination index CI obtained by the combined application and determination of a plurality of concentrations of the two is lower than 1, which shows that the paris polyphylla saponin II can sensitize cisplatin and improve the anti-tumor curative effect of the cisplatin. The source of the traditional Chinese medicine rhizoma paridis is wide, and the rhizoma paridis saponin II extracted and separated from the plant is proved by a more detailed in vitro activity test to be used together with a first-line chemotherapeutic drug cis-platinum, so that the drug resistance of the cis-platinum can be effectively improved, and the anti-tumor activity of the cis-platinum is improved.)

1. Application of rhizoma paridis saponin II in preparing medicine with sensitization cisplatin therapeutic effect is provided.

Technical Field

The invention belongs to the field of tumor chemotherapy sensitization medicines, and relates to a new application of rhizoma paridis saponin II, namely an application of rhizoma paridis saponin II in preparation of a sensitization cisplatin curative effect medicine.

Background

Cisplatin is an indispensable first-line chemotherapeutic drug for treating lung cancer, has a unique anticancer mechanism and a wide anticancer spectrum, and has been developed into one of the most widely used chemotherapeutic drugs for treating lung cancer in clinic at present. Cisplatin is a small molecular platinum compound, and after the cisplatin is passively transported into cells through a transport channel on the surface of a cell membrane, because the concentration of chloride ions in cytoplasm is lower than that of hydrated ions under normal conditions, the chloride ions are replaced by water to form hydrated cations which can be combined with bases of DNA in cell nuclei to form cross-linking of three forms of DNA-platinum, so that DNA damage is caused, DNA replication and transcription in nuclei are damaged, DNA damage is induced, and then apoptosis is induced. The main problem in cisplatin clinical chemotherapy is general drug resistance. Most cancers initially respond to cisplatin treatment, but almost always recur. The resistance mechanism of cisplatin mainly comprises the reduction of accumulation of cisplatin drug concentration in cytoplasm and the increase of anti-apoptosis effect caused by DNA repair and the like. Therefore, the drug resistance of cisplatin is a problem to be solved urgently in clinical chemotherapy.

The traditional Chinese medicine Paris polyphylla is dried rhizome of Paris polyphylla Smith var.yunnanensis (Franch.) hand-Mazz or Paris polyphylla Smith var.chinensis Hara in the family of liliaceae, has the effects of clearing heat and releasing toxin, relieving swelling and pain and cooling liver and arresting convulsion, is used for treating symptoms such as furuncle swelling, sore throat, venomous snake bite, traumatic injury and pain, cold wind and convulsion and the like, has the functions of stopping bleeding, eliminating phlegm, inhibiting bacteria, relieving pain and calming, preventing pregnancy and killing sperms, resisting cytotoxin and the like through modern pharmacological actions, and has obvious curative effects in clinical application of treating functional uterine bleeding, neurodermatitis, surgical inflammation, inhibiting tumors and the like.

The chemical components of rhizoma paridis mainly include steroid saponins, phytoecdysone and flavonoids. The steroid Saponins are mostly paris polyphylla Saponins (RPS) and have broad-spectrum antitumor activity, and mainly comprise diosgenin and pennogenin, wherein the diosgenin accounts for most of the total Saponins. Paris saponin II belongs to diosgenin, and has multiple anticancer activities. For example, the paris polyphylla saponin II can inhibit the growth of lung cancer of T739 mice and can also inhibit the growth of HepG2 cells, mainly by inducing the cells to generate apoptosis and S-phase retardation. Paris saponin II can also promote apoptosis of ovarian cancer cells to play an anti-cancer role by inhibiting angiogenesis and increasing activity of Bax, cytoplasmic cytochrome C, Caspase-3 and Caspase-9, and can also inhibit NF-kB signal pathway. However, the action mechanism of the paris polyphylla saponin II is complex, the paris polyphylla saponin II has good anti-tumor effect and wide anti-tumor activity, but reports on whether the combined action of the paris polyphylla saponin II and the conventional chemotherapeutic cisplatin can improve the sensitivity of the cisplatin are not found.

Disclosure of Invention

The invention aims to overcome the defects of the prior art and provides the application of the paris polyphylla saponin II in preparing the sensitization cisplatin curative effect medicine.

The technical scheme of the invention is summarized as follows:

application of rhizoma paridis saponin II in preparing medicine with sensitization cisplatin therapeutic effect is provided.

The invention has the advantages that:

research shows that in lung cancer cell strain H446, the single use of cisplatin only slightly reduces the cell survival rate, and the combination of cisplatin and paris saponin II greatly reduces the survival rate. Meanwhile, the combination index CI obtained by the combined application and determination of a plurality of concentrations of the two is lower than 1, which shows that the paris polyphylla saponin II can sensitize cisplatin and improve the anti-tumor curative effect of the cisplatin.

The source of the traditional Chinese medicine rhizoma paridis is wide, and the rhizoma paridis saponin II extracted and separated from the plant is proved by a more detailed in vitro activity test to be used together with a first-line chemotherapeutic drug cis-platinum, so that the drug resistance of the cis-platinum can be effectively improved, and the anti-tumor activity of the cis-platinum is improved.

Drawings

FIG. 1 shows the results of the anti-tumor activity experiments of the single and combined treatment groups of Paris saponin II and cisplatin. Wherein A is the in vitro anti-tumor activity of the rhizoma paridis saponin II; b is the antitumor activity of the single or combined cisplatin group.

Detailed Description

Rhizoma paridis saponin II with molecular formula of C₅₁H₈₂O₂₀The molecular weight is 1015.18, white crystal powder can be dissolved in organic solvents such as methanol and DMSO, and can also be easily dissolved in water; is derived from rhizome of Paris yunnanensis Franch of Liliaceae. Purchased from grand philips biotechnology limited, CAS No.: 50773-42-7, structural formula:

the present invention will be further illustrated by the following specific examples.

Method for testing antitumor activity

1. In vitro antitumor Activity test

1.1 cell lines and culture

The small cell lung cancer cell line H446 cell line is purchased from Changsha Youze biotechnology limited, and is an adherent cell, and is cultured in a culture medium (purchased from Solebao company, and contains 100U/mL penicillin and 100 mu g/mL streptomycin) containing 10% inactivated fetal bovine serum and RPMI1640, and the temperature is 37 ℃ and the content of 5% CO₂Culturing in incubator under saturated humidity condition, and subculturing for 3-4 days.

1.2 determination of antitumor Activity of drugs

(1) The paris polyphylla saponin II has the following anti-tumor effect: the H446 cells in logarithmic growth phase are digested with pancreatin and then prepared into the cells with the concentration of 3 multiplied by 10⁵one/mL cell suspension was plated in 96-well cell culture plates at 200. mu.L/well. At 37 deg.C, 5% CO₂And culturing under saturated humidity condition for 24h, adding rhizoma paridis saponin II solution (with DMSO as solvent) and corresponding solvent (1 μ LDMSO) at final concentrations of 0.25, 0.5, 2, and 5 μmol/L as control, and setting the tested drugs in 5 dose groups with 8 parallel wells. Then, after further culturing the cells at 37 ℃ for 24 hours, 100. mu.L of MTT (0.5 mg/mL) freshly prepared in PBS (pH 7.4) was added to each well. The culture was continued for 4h and the medium was gently aspirated off. Adding 100 mu L DMSO into each well, dissolving MTT formazan grains, oscillating and mixing uniformly by using a micro oscillator, selecting a 570nm position in an enzyme labeling instrument to measure the light absorption value OD, repeating the experiment for three times, and taking the average value. Using the tumor cells treated by the solvent control as a control group, setting the cell survival rate of the control group as 100%, and calculating the cell survival rate of the rest groups according to OD values, wherein the formula is as follows: cell viability was determined as OD value in experimental group/OD × 100% in control group. The results are shown in FIG. 1A.

(2) The combination of the rhizoma paridis saponin II and cisplatin has the following synergistic anti-tumor effect: human lung cancer cell line H446 (3X 10) in log phase growth⁵one/mL) were plated into 96 well cell culture plates at 200 μ L per well. At 37 deg.C, 5% CO₂Culturing under saturated humidity condition for 24 hr, and adding final concentrateThe control was a solution of polyphyllin II in degrees (0.5. mu. mol/L, 1. mu. mol/L), and cisplatin solutions (in which the solvent was DMSO) and the corresponding solvent (1. mu. LDMSO) added at different final concentrations (50. mu. mol/L, 100. mu. mol/L, 200. mu. mol/L, 300. mu. mol/L and 500. mu. mol/L). 8 parallel wells were set for each concentration, and after further culturing the cells at 37 ℃ for 24 hours, 100. mu.L of MTT (methyl thiazolyl tetrazolium) freshly prepared in PBS (pH 7.4) was added to each well, followed by further culturing for 4 hours, and then 100. mu.L of DMSO was added to each well to dissolve the MTT formazan particles, followed by shaking and mixing with a micro-shaker, measuring the absorbance OD at 570nm with a microplate reader, repeating the experiment three times, and taking the average value. The results are shown in FIG. 1B, which is a graph showing the effect of low concentrations of parinusaponin II on cell viability following treatment with cisplatin in combination. As can be seen from FIG. 1B, the survival rates of the cisplatin groups at low concentrations (0.5. mu. mol/L) of 100. mu. mol/L, 200. mu. mol/L and 300. mu. mol/L were respectively reduced from (96.41. + -. 0.03) to (56.24. + -. 0.43), from (67.55. + -. 0.32) to (43.64. + -. 0.56) and from (56.65. + -. 0.43) to (26.65. + -. 0.73), the cell survival rates were almost doubled and the co-therapy coefficient was less than 1 (using CompuSyn software), indicating a synergistic effect of the two combinations. Therefore, the low concentration of the 0.5 mu mol/L paris polyphylla saponin II can obviously improve the cisplatin chemical treatment effect of tumor cells and increase the cisplatin sensitivity.

Results of antitumor Activity test with Secondary antibody

See figure 1 and table 1.

TABLE 1 combination index of Paris Saponin II and cisplatin combination

Three conclusions

The experiment proves that the paris polyphylla saponin II has higher anti-lung cancer activity and can be used as a cisplatin chemical treatment sensitizer to be applied to anti-tumor.