Addressing Multi-Drug Resistance In Cancer With Bioactive Material And Its Therapeutic Applications

Multi-drug resistance in tumor is one of the main reasons for clinical chemotherapy failures.Therefore,it is crucial to reverse and overcome chemoresistance for reducing cancer mortality.In addition,compared to traditional biomedical materials,bioactive materials could be designed with desired bioactivities to regulate the biological functions,which has been demonstrated with broad prospects for a series of clinical applications.In this dissertation,we designed and fabricated a series of bioactive materials with specific biofunctions to reverse chemoresistance of tumor and achieve efficient tumor inhibition effects.The overall work could be categorized in the following three research areas:1.Regulating molecular mechanisms in drug resistance tumor(Chapters 2-3):reactivating cisplatin in cisplatin-resistant tumor cells;reversing relative broadspectrum chemoresistance by metabolism inhibition.2.External field assisted precise drug delivery(Chapters 3-4): designing and synthesizing ultrasound-responsive bioactive materials to achieve effective deep tumor penetration;fabricating plasmonic pyramid arrays to achieve transient intracellular delivery of biomolecules by photothermal effects.3.Biomolecules-based chemotherapy-free cancer therapy(Chapter 6): enzymatic reaction triggered peptide assemblies to interfere with microtubule self-assembly to inhibit drug-resistant tumor growth.The main research results obtained in this dissertation are listed as follows.(1)Design and synthesis bioactive nanogel to reactivate cisplatin in drug resistance cancer cell through regulating related molecular mechanismsFor enhancing drug activity and apoptotic factor in cisplatin-resistant cells,we developed a multifunctional nanogel(Valproate-D-Nanogel)to reactivate cisplatin and enhance early apoptosis.Valproate-D-Nanogel was prepared by copolymerization of carboxymethyl chitosan with diallyl disulfide and then grafted with Valproate to reverse chemoresistance in cisplatin-resistant human lung adenocarcinoma cells.Valproate-DNanogel could reactivate cisplatin by high level of G2/M arrest up to 3.2-fold enhancement induced by Valproate.Concurrently,intracellular ROS-P53 crosstalk enhances early apoptosis,which was resulted from upregulation of ROS(up to 8-fold increase)and P53(up to 18-fold increase).Enhanced G2/M arrest and ROS-P53 crosstalk could work together to reverse cisplatin resistance with a high resistance reversal index(50.22).Animal study revealed that an increased in vivo tumor suppression(up to 15-fold compared to free cisplatin)and a reduction in systemic toxicity were observed after treatment with Valproate-D-Nanogel.(2)Relative broad-spectrum chemoresistance reversing through mutually reinforcing metabolic inhibition and autophagyIn order to inhibit tumor rapid proliferation and abnormal intracellular metabolism,we developed a strategy to enhance tumor killing effects through mutually reinforcing metabolic inhibition and autophagy.First,the mitochondrial targeting molecule triphenylphosphine and the metabolic inhibitor lonidamine were grafted onto polylysine as prodrug.After self-assembly of polylysine prodrug,ferrocene and glucose oxidase were immobilized to provide additional chemotherapeutic functions(FG/TNanoprodrug).Effective mitochondrial targeting and metabolic inhibition were observed in drug-resistant cancer cells.In addition,owing to inhibited metabolism,more glucose was catalyzed to produce excess ROS via glucose oxidase and ferrocene in FG/T-Nanoprodrug.Meanwhile,such overproduced ROS increased mitochondrial permeability to facilitate the release of cytochrome C from mitochondria for high levels of autophagy.Overall,FG/T-Nanoprodrug induced enhanced metabolic inhibition(up to 3.7-fold compared to free lonidamine)and autophagy(up to 125.3-fold compared to free lonidamine),demonstrating effective therapeutic effects towards drug-resistant tumor proliferation both in vitro and in vivo.(3)Ultrasound regulated stiffness tunable nanogel for deep tumor penetration and chemoresistance reversalIn addition to endogenous stimuli-responsive nanomedicine,exogenous(ultrasound)responsiveness was additionally embedded into nanomedicine for targeted and precise drug delivery into deep tumor site.Polylysine and Pluronic F127 was first co-assembled to form nanocomplex,which was further cross-linked by genipin to endow stable structure,ultrasound-modulated stiffness,and endosomal escape capability.Subsequently,the ICAM-1 antibody was grafted onto the nanogel(Gen PLPFT)for active cancer targeting.Upon external ultrasound stimulation,F127 in Gen PLPFT was shed to induce expansion of the nanogel with increased size from 329 nm to 516 nm and decreased Young’s modulus from 336.78 Kpa to 3.93 Kpa.Through intravenous injection,the relatively rigid Gen PLPFT was able to achieve high levels of accumulation at the tumor site through active targeting and long-term stable blood circulation.Under ultrasound stimulation at tumor site,Gen PLPFT became softer with enhanced deformability to achieve deep tumor penetration.Three-dimensional cell mass and in vivo studies have shown that Gen PLPFT could penetrate into deeper regions of xenograft tumors with enhanced therapeutic effects against drug-resistant tumor and reduced cytotoxicity.(4)Enhanced intracellular delivery of biomacromolecules(mi RNA)using plasmonic pyramidal arrays by photothermal effects:To increase the intracellular delivery efficacy of biomolecules,plasmonic pyramidal arrays were fabricated to induce photothermal effects for immediate and efficient intracellular mi RNA delivery.After short-term laser stimulation(8 μs @ 640nm),a transient high temperature was generated on the plasmonic pyramids to disrupt the cell membrane for facilitating intracellular delivery of biomolecules.Meanwhile,after removal of the laser irradiation,the temperature of plasmonic pyramidal arrays rapidly decreased to 37 °C to avoid cell damage.Experimental data revealed that mi RNA-185 was successfully delivered into drug-resistant esophageal cancer cells with optimized delivery efficiency(64%)and cell viability(>95%).Furthermore,high killing effect(61.9%)on drug-resistant esophageal cancer cells could be achieved after synergistic treatment with mi R-185 and Taxol during laser irradiation.(5)Enzyme induced peptide assemblies with intrinsic apoptosis against drugresistant melanoma in the absence of chemotherapyTo avoid severe side effect and systematic toxicity for patients with drug-resistant(advanced)cancer,biomolecules based cancer therapy with low/no toxicity would be of great value in clinic.In this project,we designed and synthesized a tyrosinaseinduced tripeptide(Phe-Phe-Tyr,FFY)assemblies to induce apoptosis against drugresistant melanoma.Catalyzed by overexpressed tyrosinase in melanoma cells,FFY is first oxidized to a melanin-like FFY dimer with a diquinone structure(m FFY),which further self-assembled into m FFY assemblies.Then,m FFY assemblies could inhibit microtubule self-polymerization and induce severe G2/M arrest(up to 13.9 %).Subsequently,mitochondrial dysfunction was also aroused,leading to overproduction of cleaved caspase 3(3.1-fold higher than control)and cleaved PARP(6.3-fold higher),achieving a high level of resistance reversal in the absence of chemotherapeutic agents.In vivo studies demonstrated that the volume of resistant melanoma tumor was reduced by 87.4% compared to control groups after treatment with enzyme induced peptide assemblies.