Polymeric Nanomedicine For Cancer Therapy: Design And Evaluation

Traditional small-molecule chemotherapeutic agents widely used in clinic have many problems:poor targeting,short half-life,low utilization rate and high side effects.Compared with traditional small-molecule chemotherapeutic agents,Nanomedicines based on polymeric carrier have the following advantages in the treatment of cancer:improved drug solubility,extended circulation time in vivo,passive targeting ability through EPR effects,low side effects and so on.According to the specific characteristics of tumor microenvironment,in this manuscript,we designed simple and appropriate carriers based on biodegradable materials for the delivery of small-molecule chemotherapy drugs in vivo.Meanwhile,based on the impact of drugs on the tumor microenvironment,we studied the effect of combined treatment between different drugs to overcome the problems in cancer treatment.The main research contents and conclusions are summarized as follows:(1)A pH and redox dual-sensitive biodegradable polysaccharide,succinic acid-decorated dextran-g-phenylalanine ethyl ester-g-cysteine ethyl ester(Dex-SA-L-Phe-L-Cys),was synthesized to load doxorubicin hydrochloride(DOX·HCl).The DOX-loaded nanoparticles,which were prepared in aqueous solution and free of organic solvents,could spontaneously self-assemble into uniform sizes.When loading DOX·HCl,mercapto Dex-SA-L-Phe-L-Cys was oxidized into a crosslinked disulfide linkage to form pH and redox dual-sensitive nanoparticles(DOX-S-NPs).The amphiphilic polymer loaded DOX·HCl into the core through electrostatic and hydrophobic interactions,meanwhile the crosslinked disulfide bond could stabilize the drug loaded nanoparticles.Furthermore,the cellular uptake of the DOX-S-NPs was comparable with that of free DOX·HCl,determined by confocal laser scanning microscopy(CLSM)and flow cytometry.Cytotoxicity assay in vitro showed that the DOX-S-NPs and free DOX·HCl were similar in inhibiting the proliferation of cancer cells.DOX-S-NPs displayed similar antitumor efficiency compared with free DOX·HCl,but lower toxicity by body weight.These dual-sensitive DOX-S-NPs provide a useful strategy for anti-tumor therapy.(2)In order to solve the problems of short half-life and poor solubility of drugs in vivo,we developed simple and controllable polymer-drug conjugates.An amphiphilic anionic copolymer,poly(L-glutamic acid)-g-methoxy poly(ethylene glycol)(PLG-g-mPEG),was synthesized.Then gemcitabine(GEM)or dihydroartemisinin(DHA)was bonded to PLG-g-mPEG by amide or esterification reaction,respectively,and the well-soluble polymer-drug conjugates GEM-NPs and DHA-NPs were prepared.DHA and DHA-NPs,which possess an intramolecular endoperoxide bridge,could be activated by heme or ferrous iron to produce reactive oxygen species(ROS),as characterized by flow cytometry and confocal laser scanning microscopy.The excess ROS generation could excite expression of heme oxygenase-1(HO-1)and suppress cytidine deaminase(CDA)expression.Moreover,more than 90%GEM was catalyzed into an inactive metabolite 2?-deoxy-2?,2?-difluorouridine(dFdU)by stromal and cellular CDA.Under low CDA expression,the inactivation of GEM is decreased in turn to exert excellent therapeutic efficiency.Furthermore,cytotoxicity assay in vitro showed that DHA+GEM and GEM-NPs+DHA-NPs had synergistic effect,with molar ratio of DHA and GEM at 10.Cell apoptosis detection,performed by flow cytometry(FCM)analysis,indicated that the combination treatment group produced markedly more pronounced apoptosis than monotherapy.The study of the combination treatment provides a reliable explanation for the principle of the combination.(3)Hypoxia-activated prodrugs(HAPs)have the potential to selectively kill hypoxic cells and convert tumor hypoxia from a problem to a selective treatment advantage.However,HAPs were unsuccessful in most clinical trials owing to inadequate hypoxia within the treated tumors,as implied by a further sub-study of a phase III clinical trial.Here,a novel strategy for the combination of HAPs plus vascular disrupting agent(VDA)nanomedicine for efficacious solid tumor therapy was developed.An effective VDA nanomedicine of poly(L-glutamic acid)-g-methoxy poly(ethylene glycol)/combretastatin A4(CA4-NPs)was prepared.The immunohistochemical CD31 staining assay,photoacoustic(PA)imaging and hypoxia probe immunofluorescence staining assay confirmed that the CA4-NPs could selectively cut off the supply source of oxygen and potentiate hypoxia levels within solid tumors,provided a strong rationale for studying the therapeutic efficacy of TPZ plus CA4-NPs.In vitro MTT assay highlighted that TPZ required low oxygen tension for complete activation to exert chemotherapeutic activity.After treatment with the combination of TPZ plus CA4-NPs,the complete tumor reduction was observed in metastatic breast 4T1 xenograft mice(initial tumor volume was 180mm~3),and the significant tumor shrinkage and antimetastatic effects were observed in challenging large tumors with initial volume of 500 mm~3.The report here highlights the potential of using a combination of HAPs plus VDA nanomedicine in solid tumor therapy.(4)A novel hypoxia-responsive nano-carrier,poly(L-glutamic acid)-g-methoxy poly(ethylene glycol)/aminoazobenzene(PLG-g-mPEG/AAB),was synthesized to load irinotecan(CPT-11)through hydrophobic interaction of?-?stack and prepared a hypoxia-responsive nanomedicine CPT-11-NPs.CPT-11-NPs had stable nanostructure under physiological conditions,but it could hydrolyze under the hypoxia condition(NADPH+nitroreductase).CLSM and flow cytometry assays indicated that the cellular uptake of the CPT-11-NPs was comparable with that of free CPT-11,and the hypoxia had no effect on the uptake of nanoparticles.Under the normoxic or hypoxic incubation,cytotoxicity assay in vitro showed that CPT-11-NPs and free CPT-11 were similar in inhibiting the proliferation of cancer cells.Meanwhile,during in vivo anti-tumor study,CPT-11-NPs exhibited lower toxicity by body weight than free CPT-11.CA4-NPs could induce hypoxia within tumors,which maybe induce the hydrolysis of hypoxia-responsive nanomedicine CPT-11-NPs to release CPT-11.In metastatic 4T1 breast tumor model,the combination treatment strategy of CA4-NPs and CPT-11-NPs displayed more tumor inhibition than the monotherapy.The above results would be expected to provide important evidence for designing simple and effective nano-carriers for the delivery of chemotherapeutic agents,and also provide some references and new strategies for nanomedicine-based chemotherapy.