| Cutaneous melanoma is a highly malignant tumor with an increasing morbidity and fatality rates in recent years.Commonly used oral,injection administration and chemotherapy have poor performance in medium and long terms treatment,because oral or injection means higher exposure concentration and systemic toxicity when the drug concentration reaches the treatment window,and strong side effects of chemotherapy further reduce patients’ compliance.On the other hand,drug resistance is also a problem that affects treatment.In this research,resveratrol,an effective ingredient extracted from Polygonum cuspidatum,has significant inhibitory effect on melanoma,skin cancer,colon cancer and other tumor cells.In addition,fluorouracil,a chemical widely used in clinical antitumor,also has a high inhibitory effect on melanoma.Resveratrol has good inhibition effect on melanoma cells and good lipid solubility,but its low water solubility makes its bioavailability low.Fluorouracil has shown strong efficacy in the treatment of skin cancer by preventing the synthesis of DNA,but its short half-life makes it have problems such as fast metabolism and poor bioavailability in clinical application.The combined administration of 5-Fu and resveratrol promotes the antitumor activity of 5-Fu,which can be used to treat cutaneous melanoma through a variety of synergistic ways,such as mediating melanoma cell apoptosis,blocking cell cycle s phase and anti-proliferation.The low solubility of drugs not only seriously reduces oral bioavailability,but also leads to poor intravenous administration,high adverse reaction rate,and serious systemic toxicity problems caused by over-administration.These shortcomings together lead to medication fatigue and poor treatment compliance.By means of transdermal delivery of dissolving microneedle,two kinds of drugs with difficult transdermal delivery can be delivered to the dermis to achieve efficient and precise targeted delivery without damaging the skin.In this study,the analytical methods of fluorouracil and resveratrol were established.The maximum absorption wavelengths of resveratrol and fluorouracil were 307 nm and266 nm,and the standard curves of the two drugs were established.The results showed that the absorbance of resveratrol and fluorouracil had a good linear relationship in the concentration range of 2-10 μg/m L and 1-15 μg/m L.RSDs of daytime,intra-day and recovery tests were all less than 4%,which met the methodological requirements.Preprescription studies of resveratrol and fluorouracil were completed.In this study,antisolvent method was used to obtain resveratrol nanosuspension.Orthogonal experiments were designed to optimize the process with particle size and dispersion index as the results,and the optimal process was drug concentration(4 mg/m L),and stabilizer concentration(1.5 mg/m L).The optimum size of nanocrystals was 140±6nm and the polydispersion index was 0.093±0.026.In this study,spray-drying and freezedrying methods were used to solidified resveratrol nanosuspension.Scanning electron microscopy and X-ray diffraction scanning were used to test the morphology and crystal shape of the dry powder,and dissolution tests were conducted in vitro.The results showed that the spray dried powder was sphere-like and amorphous,with a particle size of 2-5μm.In the dissolution test,the solubility and dissolution rate of the powder were 30 times higher than that of the bulk drug,and 15 times higher than that of the bulk drug.The inclusion complex of fluorouracil @ hydroxypropyl β-cyclodextrin was prepared by aqueous solution inclusion method.The inclusion complex powder was obtained by freeze-drying,which improved its insolubility and short half-life in neutral solution.The inclusion complexes were verified by Fourier infrared,nuclear magnetic hydrogen spectrum and X-ray diffraction scanning.The characteristic peaks of fluorouracil in the inclusion complexes were completely disappeared,and only the characteristic peaks of hydroxypropyl β-cyclodextrin were shown.The test results showed that the inclusion complex of fluorouracil @ hydroxypropyl β-cyclodextrin was prepared successfully,and the inclusion rate was 59.14±1.29%.In the design of dissolving microneedle,polyvinylpyrrolidone and polyvinyl alcohol were selected as the composite matrix,and the soluble matrix with good mechanical strength was obtained by casting and demoulding.The maximum force that microneedles could bear was 0.83 N.Laser confocal fluorescence microscopy showed that resveratrol nanocrystals and fluorouracil inclusion complexes were evenly distributed in dissolving microneedle.Mechanical tests and in vivo and in vitro puncture tests showed that dissolving microneedle had good mechanical properties and insertion properties.Camera observation and skin surface water loss instrument were used to test the recovery state of skin after microneedle administration.In the test,skin basically recovered its original appearance after 360 s after administration,and skin surface water loss was the same as normal skin after 4h.In Franz diffusion cell test,dissolving microneedle released84.13±5.19% resveratrol and 90.65±18.23% fluorouracil at 12 h,and 81.24±5.23%resveratrol and 72.48±7.82% fluorouracil in vivo.In biocompatibility experiments,the dissolving microneedle codelivery system of resveratrol nanocrystals and fluorouracil inclusion complexes prepared in this study showed high biosafety.Cytotoxicity test showed that the survival rate of cells was above85%,and no hemolysis occurred.In conclusion,dissolving microneedle codelivery of resveratrol nanocrystals and fluorouracil inclusion complexes has good pharmaceutical and biological properties,and can achieve accurate transdermal drug delivery at the target site,which has a good clinical application prospect in the treatment of cutaneous melanom. |
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