Research On A Novel Drug Delivery System For Lung Targeting Of Plumbagin Based On Nanostructured Lipid Carriers

Nanostructured lipid carriers（NLCs）are a new type of good carrier for pulmonary drug administration,which has a passive targeting effect on the lung.Compared with traditional pulmonary preparations,NLCs has the advantages of improving drug stability,increasing bioavailability of insoluble drugs,or achieving sustained or controlled release in the lungs.NLCs related research has become one of the new hot spots in the research of novel pulmonary drug delivery system,such as modifying the surface of NLCs to achieve active targeting of lungs,which is expected to further improve the drug therapeutic index and reduce the adverse reactions of drugs.D-mannose with the structure of 3,4-OH can be recognized by alveolar surface mannose receptors（MRs）.Modification of NLCs with mannose can increase pulmonary targeting and make the lungs concentrated.Plumbagin（PLB）is a natural naphthoquinone with anti-tumor,anti-pulmonary fibrosis,anti-inflammatory and other pharmacological activities.However,its poor water solubility and low bioavailability after oral administration make it difficult for the ordinary dosage form to achieve the purpose of clinical treatment of pulmonary fibrosis.Based on this,the mannose-modified plumbagin nanostructured lipid carriers（PLB-Man-NLCs）was prepared,and its quality was evaluated.The in vitro pharmacodynamics,in vivo tissue distribution and pharmacokinetics were preliminatively investigated,in order to improve the retention of PLB in the lung and achieve the purpose of treating pulmonary fibrosis.In this paper,the encapsulation efficiency and drug loading were determined by ultrafiltration centrifugal method,and the Plumbagin nanostructured lipid carrier（PLB-NLCs）was prepared by ethanol injection method.The encapsulation efficiency and particle size were used as evaluation indexes for single factor screening,and Box-Behnken response surface method was used to optimize the formulation of PLB-NLCs.Then,Octamine-mannose（ODA-Man）was synthesized by Schiff’s base reaction principle and added into the optimal PLB-NLCs prescription to obtain PLB-Man-NLCs.The quality evaluation results showed that PLB-NLCs and PLB-Man-NLCs were spherical or quasi-spherical with complete and round surfaces under transmission electron microscopy.Particle size was（205.82±2.16）nm and（175.66±3.29）nm,PDI was 0.195±0.005and 0.230±0.011,Zeta potential was（-20.62±1.59）m V and（-42.81±2.44）m V,respectively.The encapsulation Encapsulation efficiency were（85.63±0.92）%and（85.85±0.90）%,and the drug loading was（5.68±0.06）%and（5.17±0.05）%,respectively.In vitro release experiments,with p H 7.4 and p H 5.0 as the release medium,PLB suspension only released about 37%of PLB at 4 h,while NLCs released about 90%of PLB,indicating that NLCs formulation can significantly improve the solubility of PLB.The release curve of PLB-Man-NLCs in vitro were fitted,and the Weibull equation was the best.Fibroblasts（NIH3T3）were used as model cells in vitro for cytotoxicity and proliferation inhibition experiments.The results of cytotoxicity assay showed that low concentrations of PLB and PLB-Man-NLCs showed no cytotoxicity to NIH3T3 cells.High concentrations of PLB and PLB-Man-NLCs both decreased the survival rate of NIH3T3 cells and showed some cytotoxicity,but the cytotoxicity of PLB-Man-NLCs group was significantly lower than that of PLB group.The results of cell proliferation inhibition experiment showed that PLB-Man-NLCs could inhibit the proliferation of NIH3T3 cells induced by TGF-β₁compared with the model group,and the inhibition effect was concentration dependent.The results indicated that PLB-Man-NLCs had the potential to weaken the degree of lung fiber.In vivo imaging was performed to investigate the distribution of NLCs in mouse tissues.The results showed that NLCs began to accumulate in the lung tissues 1 h after entering the systemic circulation and could remain for 24 h.The fluorescence intensity of Mannose-modified nanostructured lipid carriers（Man-NLCs）in the isolated lung tissue after dissection was significantly higher than that of the NLCs group,and the fluorescence intensity of Man-NLCs group in the liver tissue was reduced,indicating that mannose-modified nanoparticles enhanced lung targeting.Pharmacokinetic experiments on rats showed that C_maxof PLB-NLCs group and PLB-Man-NLCs group were 5.07 times and 7.43 times of PLB group,AUC_0-Twere 2.21 times and 3.98 times,AUC_0-∞were 1.94 times and 3.05 times,respectively.The results showed that NLCs could increase the plasma concentration of PLB in rats.In this study,mannose-modified plumbagin nanostructured lipid carrier was successfully prepared,which effectively enhanced the inhibitory effect of PLB on the proliferation of TGF-β₁induced NIH3T3 cells and reduced the cytotoxicity of PLB.At the same time,the blood concentration and lung targeting of PLB were improved,which laid a good foundation for the subsequent development of PLB anti-pulmonary fibrosis treatment preparations...