Studies On Enhancing Intratumoral Drug Penetration With Chitosan Grafting Drug Delivery System

Desmoplastic tumors have high degrees of malignancy and are prone to recurrence.The main challenge of chemotherapy is the cancer associated fibroblast barrier(CAFs)derived barnders,which prevent delivered drugs from reaching targeting sites in tumor cells.The drug delivery system holds great potentials of tumor targeting delivery,which could overcome the drug distribution barriers,and achieve precise delivery of tumor targets.Degradable marine biomaterials play an important role in drug delivery system(DDS)research area.However,traditional marine biomaterials have relatively simple functions.In order to meet the desired functions such as improving drug-loading capacity and cells targeting of DDS,it is necessary to perform structure modification process for drug delivery system to achieve the targeted delivery of therapeutic drugs.Chitosan is derived from algae cells,shrimp,crab shell,etc.,and has good biodegradability and biocompatibility.In this study,Chitosan oligosaccharide-g-stearic acid(CSOSA)was prepared by grafting stearic acid on chitosan to encapsulate poorly soluble drags.Angiotensin II type I receptor(ATiR)ligand Telmisartan(Tel)was modified on the prepared CSOSA,and Doxorubicin(DOX)was used as a model drug to construct Tel-modified CSOSA drug delivery(Tel-CSOSA/DOX),which could achieve dual targeting cancer associated fibroblasts and MCF-7 breast cancer cells with high expression of AT iR.Results showed that drugs distribution for Tel-CSOSA/DOX and CSOSA/DOX in tumor tissue after primarily intravenous injection was 35.8%and 0.6%,respectively.Influenced by the pathological structure of dense tumor stroma,the both two drug delivery systems were poorly permeable after the drugs crossed the blood vessel,and the drug distribution patterns were also heterogeneous.After continuous drugs administration,the drug-distributed area of tumor tissue increased to 61.8%in the Tel-CSOSA/DOX group.With the active targeting of cancer associated fibroblasts by ligand Tel,Tel-CSOSA/DOX produces a significant proapoptotic effect for cancer associated fibroblasts by releasing delivered DOX,resulting in significant reduction in secretion of tumor stroma such as hyaluronic acid and collagen.The depth and uniformity of drug distribution was also significantly improved.After CSOSA/DOX treatment,the pathological structure of the tumor and the intratumoral drug distribution were not significantly influenced as that before administration.The percentage of Ki67 positive cells for Tel-CSOSA/DOX,CSOSA/DOX and DOX was 10.5%,57.6%and 61.6%that of the negative control group,respectively,indicating that Tel modification significantly enhanced the in vivo antitumor effect of chitosan based drug delivery system.New drug delivery systems that could eliminate cancer associated fibroblast derived barriers were constructed by utilizing Telmisartan to down-regulate the expression of a-smooth muscle actin(a-SMA)and connective tissue growth factor(CTGF)for reversing the pharmacological effects of cancer associated fibroblast phenotype.In this study,constructed CSOSA was selected as the carrier to encapsulate Telmisartan(Tel)and antitumor drug Doxorubicin(DOX)to prepare drug-loaded drug delivery system(CSOSA/Tel and CSOSA/DOX).Sequentially delivering prepared CSOSA/Tel and CSOSA/DOX to cancer associated fibroblasts and tumor cells to improve anti-tumor efficacy.MCF-7 breast cancer bearing nude mice were adopted as in vivo model to investigate intratumoral drug penetration patterns in vivo.Results showed that the intratumoral drug content increased to 1.93 times that of the control groups after intravenous injection of CSOSA/Tel.The drugs distributed area also increased by 11.9 times compared with the control group,indicating that CSOSA/Tel could significantly increase intxatumoral distribution depth and uniformity of the drug.The expressed a-SMA in tumor tissues after CSOSA/Tel and free Tel treatment decreased by 56.1%and 22.1%,respectively.The tumor stroma such as collagen were also significantly down-regulated,and the distributed structure of hyaluronic acid was also destroyed,contributing to decreased solid stress of 56.5%and 21.2%,respectively.The content of transforming growth factor-?(TGF-?)was decreased by 85.2%and 20.1%,respectively,indicating that the barriers of drug penetration in tumor tissues were eliminated with the reversal phenotype of caner associated fibroblast after CSOSA/Tel administration.In addition,bioactive cytokines that promote tumor cell proliferation are also inhibited.The cytotoxicity results in vitro showed that the combined CSOSA/Tel and CSOSA/DOX at molar ratio of 4:1,2:1 and 1:1 showed good synergistic inhibition of MCF-7 breast cancer cell proliferation.Furthermore,Annexin V/PI study confirmed that CSOSA/Tel,CSOSA/DOX,combined CSOSA/Tel and CSOSA/DOX(Tel:DOX?2:1,mol/mol)could induce 28.5%,50.8%and 81.3%of MCF-7 breast tumor cells apoptosis,indicating that combination of CSOSA/Tel and CSOSA/DOX has better in vitro efficacy when the Tel:DOX molar ratio was 2:1.The anti-tumor efficacy of different drugs on MCF-7 breast tumor bearing nude mice showed that the tumor inhibition rates of DOX and CSOSA/DOX(The dosage of administrated DOX was 3 mg/kg and administered once another day for 3 times)were 28.9%and 34.8%,respectively.After intravenous injection of CSOSA/Tel three times to eliminate the tumor-associated fibroblast barrier,CSOSA/DOX sequential therapy was continued,which induced tumor inhibition rate of 69.5%.At post-intravenous injection of CSOSA/Tel,sequential intravenous injection of CSOSA/Tel and CSOSA/DOX preparations with a Tel:DOX molar ratio of 2:1,the inhibition rate increased to 78.5%,indicating that the elimination of cancer associated fibroblasts derived barriers combined with sequential administration of synergetic drugs has important reference value for the treatment of desmoplastic tumors.