Preparation method and application of alkyl-modified artesunate nano prodrug

Abstract

The invention relates to a preparation method and application of an alkyl-modified artesunate nano prodrug, and belongs to the technical field of antitumor drugs. Although artesunate has antitumor activity, the artesunate is low in water solubility, short in half-life period and poor in metabolic stability, so that clinical application is limited. According to the invention, artesunate is combined with octadecylamine, octadecanol or octadecanethiol, and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI) and 4-dimethylaminopyridine (DMAP) are used as catalysts, so that the artesunate nano prodrug is successfully synthesized. According to the artesunate nano prodrug, the anti-tumor activity of artesunate is remarkably improved, and the artesunate nano prodrug has a remarkable inhibition effect on tumor cells such as A549, MCF-7 and Hela.

Description

Technical Field

[0001] The present invention belongs to the technical field of anti-tumor nanomedicines, and particularly relates to a method for simply preparing a long alkyl chain-modified artesunate nano-prodrug and its application. Background Art

[0002] Artesunate, a derivative of artemisinin containing a sesquiterpenoid structure, has been widely used in the clinical treatment of malaria, exhibiting multiple pharmacological effects, including immunomodulation, anti-inflammatory, antibacterial sensitization, and antitumor effects. Li Bin et al. reported that artemisinin and its derivatives exert their antitumor effects primarily by inducing apoptosis and inhibiting tumor cell proliferation (Chinese Journal of Clinical Pharmacology and Therapeutics, 2010). However, artesunate's low solubility in water and short half-life limit its clinical application, especially when high doses are required. Furthermore, artesunate's poor metabolic stability in vivo results in low bioavailability, further limiting its therapeutic efficacy. To address these issues, significant research has been conducted. Ismail et al. developed amphiphilic artesunate cationic liposomes using diploid technology and self-assembly methods, demonstrating the advantages of higher artesunate content, longer half-life, and stronger antimalarial properties. (Artesunate-based Nanomedicines: Synthesis, Characterization, and Antimalarial Studies [D]. Southeast University, 2019). Meng Haijing et al. modified artesunate with polyethylene glycol monomethyl ether to obtain a product with improved water solubility and biocompatibility. Although prodrug strategies have the potential to regulate drug properties at the molecular level, these modifications often only address a single issue and struggle to balance multiple requirements, such as solubility, stability, and targeting.

[0003] Based on the aforementioned state of the art, the present invention creatively applies an octadecyl modification strategy to the structural modification of artesunate, addressing the challenges of selective modification and activity preservation through systematic molecular design. The resulting novel prodrug not only improves the physicochemical properties of the original drug but also possesses the unique ability to self-assemble into a nano-drug delivery system, opening up new avenues for the clinical application of artesunate. This technological innovation is expected to significantly enhance the therapeutic efficacy of artesunate and also provide new ideas and methods for the modification of other poorly soluble drugs. Summary of the Invention

[0004] Artesunate is an important antimalarial drug with a wide range of biological activities. Structural modification of artesunate is of great significance to further improve its bioavailability and efficacy. The present invention provides a method for preparing and using an octadecyl chain-modified artesunate nanoparticle. The nanoparticle is prepared by the following steps:

[0005] Artesunate and an octadecyl derivative are placed in a round-bottom flask, followed by the addition of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride and 4-dimethylaminopyridine as catalysts. Under normal temperature conditions, the reaction mixture is dissolved in anhydrous dichloromethane and stirred for 24 hours. After the reaction is completed, water is added to quench the reaction, followed by extraction with dichloromethane. The organic phase is washed with saturated brine and dried over anhydrous sodium sulfate. Thereafter, the crude product is concentrated by rotary evaporation. Finally, the product is purified by silica gel column chromatography (petroleum ether: ethyl acetate) to obtain a colorless oily product.

[0006] In the step, the molar ratio of artesunate, 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, 4-dimethylaminopyridine and octadecyl derivative is 1:0.1-1.5:0.1-0.5:1.2-2.

[0007] The column chromatography method in the step uses 200-300 mesh silica gel, and the elution liquid is petroleum ether:ethyl acetate = 1:1. BRIEF DESCRIPTION OF THE DRAWINGS

[0008] Figure 1 This is the mass spectrum of artesunate nanoprodrug 1a.

[0009] Figure 2 This is the mass spectrum of artesunate nanoprodrug 1b.

[0010] Figure 3 This is the mass spectrum of artesunate nanoprodrug 1c.

[0011] Figure 4 Schematic diagram of the particle size of artesunate nanoprodrug 1a,

[0012] Figure 5 Schematic diagram of the particle size of artesunate nanoprodrug 1b,

[0013] Figure 6 Schematic diagram of the particle size of artesunate nanoprodrug 1c. Specific implementation plan

[0014] In this study, artesunate was reacted with octadecylamine, octadecanol, and octadecanethiol using 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI) and 4-dimethylaminopyridine (DMAP) as catalysts.

[0015] Example 1

[0016] Artesunate (0.1 g, 0.26 mmol), octadecylamine (0.10 g, 0.39 mmol), EDCI (4.9 mg, 0.026 mmol) and DMAP (0.036 g, 0.294 mmol) were taken in a dry round bottom flask, dissolved in 5 mL dry dichloromethane and stirred at room temperature for 24 h. After completion of the reaction, quenched with DI water, extracted with dichloromethane, washed the combined organic phase with saturated NaCl solution, dried over anhydrous Na2S04and concentrated under reduced pressure to get the crude product. The crude product was purified by silica gel column chromatography, monitored by TLC and finally artesunate octadecylamine la was obtained as colorless oil in 95% yield.

[0017] Example 2

[0018] Artesunate (0.1 g, 0.26 mmol), octadecylamine (0.10 g, 0.39 mmol), EDCI (4.9 mg, 0.026 mmol) and DMAP (0.036 g, 0.294 mmol) were taken in a dry round bottom flask, dissolved in 5 mL dry dichloromethane and stirred at room temperature for 24 h. After completion of the reaction, quenched with DI water, extracted with dichloromethane, washed the combined organic phase with saturated NaCl solution, dried over anhydrous Na2S04and concentrated under reduced pressure to get the crude product. The crude product was purified by silica gel column chromatography, monitored by TLC and finally artesunate octadecylamine la was obtained as colorless oil in 95% yield.

[0019] Example 3

[0020] Artesunate (0.1 g, 0.26 mmol), octadecylamine (0.10 g, 0.39 mmol), EDCI (4.9 mg, 0.026 mmol) and DMAP (0.036 g, 0.294 mmol) were taken in a dry round bottom flask, dissolved in 5 mL dry dichloromethane and stirred at room temperature for 24 h. After completion of the reaction, quenched with DI water, extracted with dichloromethane, washed the combined organic phase with saturated NaCl solution, dried over anhydrous Na2S04and concentrated under reduced pressure to get the crude product. The crude product was purified by silica gel column chromatography, monitored by TLC and finally artesunate octadecylamine la was obtained as colorless oil in 95% yield.

[0021] Example 4

[0022] Well grown cells were prepared as single cell suspension and the cell density was adjusted to 5 x 10 3Cells were seeded in 96-well plates at a density of 100 μL per 100 μL of cell suspension per well. Three to six replicate wells were set up for each group, and a blank control group was established. Cells were pre-cultured in a 37°C, 5% CO2 incubator until adherent. Then, gradient concentrations of artesunate prodrug solutions were added for intervention and cultured for 24-72 hours. After incubation, 10 μL of CCK-8 reagent was added to each well, gently mixed, and incubated at 37°C in the dark for 1-4 hours. The absorbance was then measured using a microplate reader at a wavelength of 450 nm (reference wavelength 600-650 nm). Cell viability or inhibition rate was calculated using the following formula: Cell viability (%) = (OD value of experimental group - OD value of blank group) / (OD value of control group - OD value of blank group) × 100; inhibition rate (%) = (OD value of control group - OD value of experimental group) / (OD value of control group - OD value of blank group) × 100. According to the experimental data in Table 1, the IC values ​​of artesunate prodrug on lung cancer cells A549, breast cancer cells MCF-7 and cervical cancer cells Hela are 50 It can be seen from the above values ​​that artesunate prodrug can significantly inhibit the proliferation of target cells.

[0023] Table 1 IC values ​​of artesunate prodrugs on A549, MCF-7 and Hela cells at 48 h 50 value

[0024]

[0025] In summary, the alkyl-modified artesunate nanoprodrug provided by the present invention has good inhibitory activity against tumor cells and good anti-cancer activity, and has certain application value and prospects in the development of corresponding anti-cancer drugs.

[0026] It should be understood that the application of the present invention is not limited to the above examples. For those skilled in the art, improvements or changes can be made based on the above description. All these improvements and changes should fall within the scope of protection of the claims attached to the present invention.

Claims

1. An alkyl-modified artesunate nano-prodrug, and the application of the prodrug in the field of anti-tumor. The artesunate nano-prodrug has the following structural formula: wherein X is NH, O or S.

2. The artesunate nanoprodrug according to claim 1, characterized in that: The structure of the artesunate nanoprodrug is shown in 1a, 1b or 1c:

3. The use according to claim 1, characterized in that The prodrug can be applied in the anti-tumor field.

4. The use according to claim 3, characterized in that The tumors include, but are not limited to, one or more of lung cancer, namely A549, breast cancer, namely MCF-7, and cervical cancer, namely Hela.

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