Design And Antitumor Application Of Nano-drug Delivery System Based On Lipid Coated Calcium Carbonate

Calcium carbonate nanoparticles have been widely used in biomedicine because of their biodegradability,in vivo stability,ease of preparation,particles in very small size range,ease of surface modification and p H sensitivity.A large number of drug delivery systems based on calcium carbonate nanoparticles have been developed.These delivery systems can play a role in the diagnosis,treatment and treatment integration of tumors by loading various types of drugs.However,the disadvantages of calcium carbonate nanoparticles including easy aggregation and unstable in aqueous solution seriously limit their use in drug delivery.Lipid materials have the unique advantages of high cellular affinity and biocompatibility.The design of lipid-coated calcium carbonate nanoparticles could be an effective approach to solve these challenges.Thereby,in this paper,three different lipid coated calcium carbonate nanoparticles were prepared and evaluated.This paper consists of three parts,which respectively introduce three different kinds of lipid coated calcium carbonate nanoparticles.In the first part,to evaluate the feasibility of preparation methods,DOX was used as the model drug and through a reverse microemulsions method,lipid coated calcium carbonate nanoparticles loaded with Doxorubicin(DOX)were prepared.Then,miR-375 was loaded by electrostatic adsorption,and obtained the final lipid-coated calcium carbonate nanoparticles co-loaded with DOX and miR-375(LCC-DOX/miR-375 NPs).TEM analysis confirmed that LCC-DOX NPs were spherical in shape with a dark calcium carbonate core.The size and zeta potential were nearly 60 nm and 27.74 m V,respectively.Release study of DOX showed that the LCC-DOX NPs could remain stable in normal tissue p H and accelerate the release of DOX in the acidic tumor microenvironment.Both the in vitro and in vivo results demonstrated that LCC-DOX/miR-375 NPs showed many excellent advantages to deliver DOX and miR-375 simultaneously including highly efficient cellular uptake ability,increased intracellular DOX accumulation,and accelerated DOX release in weakly acid condition.More importantly,LCC-DOX/miR-375 NPs could enhance the anti-tumor effect by reversing the multi-drug resistance of DOX in hepatocellular carcinoma cell lines.In the second part,considering that the adsorption of miR-375 on the surface of nanoparticles might affect its stability,miR-375 was further loaded into the core of the calcium carbonate nanoparticles.At the same time,sorafenib was used to replace the previous DOX.Finally,lipid coated calcium carbonate nanoparticles with a size of 50nm and loaded with sorafenib and miR-375(miR-375/Sf-LCC NPs)were prepared.The NPs show high loading efficiency and accelerated drug release in weakly acid condition.Furthermore,chemotherapeutic drugs including sorafenib have been shown to induce autophagy in liver cancer,and Autophagy acts as a double-edged sword in cancer,evolution and therapy by both promoting cancer survival and contributing to cancer cell death.Autophagy removes newly mutated cells and damaged mitochondria in the early stage of cancer.However,upon tumor establishment,autophagy induces survival in stressful environment,decreases therapeutic effects and cell death.Because miR-375 could reduce the level of autophagy in liver cancer,in this part,we expected to explore synergistic effect from the perspective of autophagy.In vitro and in vivo results showed that miR-375/Sf-LCC NPs could exert the synergistic effect of sorafenib and miR-375 through autophagy inhibition in liver cancer.In the third part,in order to further expand the use of lipid-coated calcium carbonate nanoparticles,Selenium(Se)was doped into the calcium carbonate NPs to prepare selenium-doped calcium carbonate NPs.Then,cisplatin was loaded into these nanoparticles to prepare the final Pt/Se@Ca CO₃ NPs.By testing the cytotoxicity of different cisplatin-selenium ratios,the best combination index(CI)was obtained.Pt/Se@Ca CO₃ NPs were prepared according to the best synergy ratio.Then,the intracellular uptake and in vivo distribution of cisplatin and selenium were analyzed.In addition,the in vitro and in vivo efficacy of Pt/Se@Ca CO₃ NPs were evaluated.The results proved that Pt/Se@Ca CO₃ NPs could deliver cisplatin and Se to tumor at the predetermined optimized synergistic ratio,consequently,resulting in synergistic efficacy both in vitro and in vivo.Moreover,with the addition of Se,the side effects of cisplatin were significantly reduced.In this paper,three different lipid coated calcium carbonate nanoparticles were designed,prepared and evaluated.At the same time,the synergistic mechanism of loading drugs was studied in detail.In summary,this study provides a new idea and a new approach for the rational design of lipid coated calcium carbonate and provides possibility for the clinical transformation of calcium carbonate nanoparticles in tumor therapy.