Biodegradable Polymers-based Cancer Nanomedcines:Design And Evaluation

Cancer is the leading disease that threatens human health.Chemotherapy is still the basic method of clinical malignant tumor therapy.However,the commonly used chemotherapy drugs have great limitations in successful clinical use,such as non-specific interaction,low drug efficiency and severe toxicity.In order to minimize the side effects and improve the therapeutic efficacy of chemotherapy,we designed and prepared biodegradable polymers based anti-cancer drug delivery systems.The responsive drug release behavior,antitumor efficiency,effect on reducing toxicity,and drug combination was evaluated.The main contents and conclusions of this dissertation are summarized below:1)A novel enzyme-sensitive polymer-doxorubicin conjugate was designed to delivery chemotherapeutic drug in a tumor-specific behavior and selectively activated in tumor tissue.Doxorubicin(DOX)was conjugated to carboxyl-terminated 4-arm poly(ethylene glycol)through a tetrapeptide linker,alanine-alanine-asparagine-leucine(AANL),which was one of the substrates of legumain,an asparaginyl endopeptidase that was found highly expressed in human tumor tissues.In vitro release showed that the existence of legumain could improve the release of DOX greatly,but less than 4%of DOX was released in the absence of legumain protease.For the MTT assay of free DOX and 4-arm PEG-AANL-DOX on legumain high-expressed MDA-MB-435 cell lines and legumain weak-expressed 4T1 cell lines,the IC50 ratio of free drug against drug conjugate showed ten times reduction,indicating that the existence of the enzyme-responsive tetrapeptide linker improved the cytotoxic significantly.This enzyme-responsive prodrug provides a useful strategy for anti-tumor drug delivery.2)In the cancer chemotherapy,the development of multi drug resistant(MDR)of cancer cells during treatment is the main cause of chemotherapy failure,and the over-expression of P-glycoprotein(P-gp)has been recognized to be the most important cause of MDR in cancer.Podophyllotoxin(PPT)is a kind of lignan anti-tubulin agent extracted from natural plant.PPT shows the ability of microtubule assembly inhibition and P-gp expression of MDR cancer cell inhibition at the same time.However,because of the poor aqueous solubility and high toxicity,PPT cannot be used in clinical cancer therapy.In order to enhance the efficiency and reduce side effect of PPT,a polypeptide based PPT conjugate PLG-g-mPEG-PPT was developed and used for the treatment of multi drug resistant breast cancer.The PLG-g-mPEG-PPT was prepared by conjugating PPT to poly(L-glutamic acid)-g-methoxy poly(ethylene glycol)(PLG-g-mPEG)via ester bonds.The PPT conjugates self-assembled into nanoparticles with average sizes about 100 nm in aqueous solution.Western blotting assay showed that the PLG-g-mPEG-PPT could effectively inhibit the expression of P-gp in the multiple drug resistant MCF-7/ADR cells.In vitro cytotoxicity assay indicated that the resistance index(RI)values of PLG-g-mPEG-PPT on different drug-resistant cancer cell lines exhibited 57-270 folds reduction than of traditional microtubule inhibitor chemotherapeutic drug PTX or DTX.Hemolysis assay demonstrated that the conjugation greatly decreased the hemolytic activity of free PPT.Maximum tolerated dose(MTD)of PLG-g-mPEG-PPT increased greatly(13.3 folds)as compared to that of free PPT.In vivo study showed that the PLG-g-mPEG-PPT conjugate remarkably enhanced the antitumor efficacy against MCF-7/ADR xenograft tumors with an tumor suppression rate(TSR)of 82.5%,displayed significantly improved anticancer efficacy.3)The synthesis of polymer-drug conjugate capable of convenient preparation and controlled release of therapeutic agents is still an urgent requirement in drug delivery field.Herein,we develop a novel anti-cancer conjugate for on-demand delivery of Podophyllotoxin(PPT)with redox sensitive behaviors.A simple polymer,poly(ethylene glycol)-b-poly(L-Aspartic acid)(mPEG-b-P(Asp-EI)),was synthesized and PPT-DTPA was directly conjugated to the carboxyl groups of mPEG-b-P(Asp-EI)to obtain the disulfide-containing polymer-PPT conjugate(P(SS-PPT)).The P(SS-PPT)micelles showed rapid drug release under tumor-relevant reductive conditions as designed.Western blotting assay showed that the P(SS-PPT)could effectively inhibit the expression of P-gp in the multiple drug resistant MCF-7/ADR cells.In vitro cytotoxicity study showed that the P(SS-PPT)micelles exhibited significantly enhanced cytotoxicity against a variety of MDR tumor cells and the RI values decreased greatly.4)We synthesized the PLG-g-mPEG-PPT and P(SS-PPT)conjugates and developed them as nanovehicles for the co-delivery and drug-combination of doxorubicin(DOX)and Podophyllotoxin(PPT).PPT can inhibit the level of P-gp expression in MDR cancer cell and DOX is one of the main substrate of P-gp,combination of PPT and DOX can enhance the therapy efficiency of DOX in MDR cancer cell.In vitro cytotoxicity assays against MCF-7/ADR cells demonstrated that the DOX and PPT co-delivered nanoparticles(Co-NPs)exhibited synergistic effect in inducing cancer cell apoptosis and the CI(combination index)value was about 0.3.Observations in Flow cytometry(FCM)and Laser confocal Microscopy(LSCM)demonstrated that the co-delivery system improved the uptake level of DOX in MCF-7/ADR cells.The above results would be expected to provide important evidence for designing simple and effective nanocarriers for the delivery of anti-cancer chemotherapeutics,and also provide some references and new strategies for biodegradable polymers-based chemotherapy.