Application Of Functional Metal-based Nanomaterials In Cancer Diagnosis And Treatment

Cancer is one of the major public problems threatening global health.The diagnosis and treatment of cancer has become an urgent scientific problem to be solved.Nanotechnology,as an emerging technological means,has broad application prospects in many fields such as biomedicine.Nanomaterials are an important part of nanotechnology.Among the developed nanomaterials,metal-based nanoparticles（MNPs）with outstanding catalytic capacity,good electrical and thermal conductivity and excellent optical properties could be considered as candidates for constructing intelligent platforms for cancer diagnosis and treatment.This paper makes full use of the advantages of MNPs to bulid multifunctional nanoplatforms for early diagnosis and treatment of cancer,aiming to provide new ideas and research methods for designing multifunctional novel diagnostic and therapeutic reagents.Part One:ATP induced alteration in the peroxidase-like properties of hollow Prussian blue nanocubes:a platform for alkaline phosphatase detectionObjective:Quantitative determination of alkaline phosphatase（ALP）in an alkaline environment was achieved using the enhanced peroxidase-like activity of hollow mesoporous Prussian blue nanocubes（h PBNCs）by 5’-adenosine triphosphate（ATP）.This method provided timely and accurate diagnosis and assessment of tumor occurrence and development.Methods:Hollow mesoporous Prussian blue nanocubes（h PBNCs）were synthesized on the basis of mesoporous Prussian blue nanoparticles（MPBs）via a hydrothermal method followed by a PVP protected hydrochloric acid-etching process.With the help of ATP,h PBNCs showed enhanced peroxidase activity over a broadened p H range.The role of ATP in enhancing the catalytic activity of h PBNCs was demonstrated in detail by means of UV-vis spectrophotometer,Electron paramagnetic resonance spectrometer（ESR）,fluorescence spectrophotometer and electrochemical workstation.On the basis of the different influences of ATP,ADP and AMP on the catalytic activity of h PBNCs,a sensitive colorimetric assay for the determination of ALP was established by employing colourless TMB as the catalytic substrate.Results:The experimental results showed that ATP changed the catalytic performance of h PBNCs through the following five aspects:（1）ATP can significantly improve the stability of h PBNCs in alkaline solution;（2）The active intermediate involved in the catalytic reaction promoted by ATP was singlet oxygen（¹O₂）instead of·OH;（3）ATP can stabilize the oxidized TMB and keep it stable for a long time;（4）The addition of ATP improved the affinity between the substrate and the catalyst,and increased the catalytic sites for the decomposition of H₂O₂;（5）ATP improved the conductivity of h PBNCs.The combined effects significantly enhanced the catalytic activity of h PBNCs in the p H range of 2～12.In addition,the ATP-induced enhancement of the catalytic activity of h PBNCs was considered to be the result of the adsorption of ATP onto the h PBNCs,and the enhancement of the enzymatic activity efficiency was closely related to the structure of ATP.By utilizing the h PBNCs-ATP based colorimetric assay,the recoveries of ALP in serum samples were in the range of97.68%to 105.40%with the relative standard deviations（RSD,n=3）from2.13%to 3.22%.Our method can be used for the qualitative analysis of ALP in complex biological samples with good recovery and precision.Conclusions:In this study,ATP can act as a boosting agent to improve the peroxidase-like activity of h PBNCs over a broad p H range.On the basis of the different influences of ATP,ADP and AMP on the catalytic activity of h PBNCs,a sensitive colorimetric assay for the detection of ALP was developed.Our work would shed light on further exploration of modulators for improving the catalytic performance of artificial enzymes and extending their applications in the diagnosis of cancer.Part Two:MOF-derived porous ZnO-Co₃O₄ nanocages as peroxidase mimics for colorimetric detection of copper（II）ions in serumObjective:A novel turn-on colorimetric sensor for selective detection of Cu²⁺in serum samples was established based on porous bimetallic transition metal oxide nanocages（ZnO-Co₃O₄ NCs）.Our findings were expected to shed light on early cancer diagnosis.Methods:Porous bimetallic transition metal oxide nanocages（ZnO-Co₃O₄NCs）were prepared by direct calcination using ZnCo-zeolitic-imidazolate-framework（ZnCo-ZIF）as precursors.The obtained ZnO-Co₃O₄ NCs exhibited excellent peroxidase activity.Introduction of cysteine（Cys）significantly inhibited the catalytic activity of ZnO-Co₃O₄ NCs,which can be recovered upon addition of Cu²⁺via the strong binding ability between Cu²⁺and Cys.Based on the ZnO-Co₃O₄ NCs/Cys system,selective detection of Cu²⁺both in buffered solution and serum samples was successfully achieved.Results:A novel turn-on colorimetric sensor for detection of Cu²⁺was established based on ZnO-Co₃O₄ NCs as peroxidase mimics.The linear range for Cu²⁺determination was 2 to 100 n M with a detection limit of 1.08 n M.Due to the high sensitivity and selectivity,the proposed assay was also successfully applied for quantitative measurement of Cu²⁺in serum without pretreatment.Conclusions:On the basis of high peroxidase-like activity of ZnO-Co₃O₄NCs,a novel“turn-on”sensing platform for Cu²⁺detection was established.The sensor realized the rapid and accurate detection of metal ions in biological samples,providing a convenient technical solution for the rapid detection of potential clinical cancer.Part Three:Mitochondria-targeting multifunctional nanoplatform for cascade phototherapy and hypoxia-activated chemotherapyObjective:Considering the hypoxic microenvironment of tumor,a mitochondrial targeted nanoplatform was designed for cancer treatment by integrating photodynamic therapy（PDT）,photothermal therapy（PTT）and hypoxia activated chemotherapy.The consumption of O₂ in the PDT process indeed aggravated tumor hypoxia,which greatly facilitated the therapeutic effect of the released hypoxia-activated prodrugs,leading to a synergistic anticancer effect of hypoxia activated chemotherapy,PTT and PDT.Methods:Hollow copper sulfide nanoparticles coated with polydopamine（HCu S@PDA NPs）with high near-infrared（NIR）photothermal conversion efficiency were used as the photothermal nanoagents in our study.Importantly,the hollow and mesoporous structure endows HCu S NPs with high drug loading capacity,making them as NIR responsive TH302 carriers for chemotherapy/photothermal synergistic therapy.Subsequently,PDA was employed to encapsulate the HCu S NPs core.The PDA coating can not only act as a smart photothermal sensitive gatekeeper,but also serve as a substrate for the conjugation with a NIR excitable PDT agent and a mitochondria-targeting ligand.The structure and properties of HCu S@PDA-Ce6/TPP NPs were characterized by scanning electron microscopy（SEM）,transmission electron microscopy（TEM）,X-ray diffractometer（XRD）,Fourier transform infrared spectroscopy（FTIR）and N₂ adsorption and desorption experiments.High performance liquid chromatography（HPLC）was used to monitor the loading and release of TH302.B16F10 cells were choiced for cytotoxicity and cellular uptake studies.B16F10 tumor-bearing C57BL/6 mice were selected to evaluate the in vivo antitumor effect of HCu S-TH302@PDA-Ce6/TPP NPs.The tissue morphology was observed by hematoxylin-eosin staining（H&E staining）.Results:The successful synthesis of HCu S-TH302@PDA-Ce6/TPP NPs was characterized by SEM,TEM,XRD,FTIR and other means.Drug release experiments proved that the platform exhibited p H/near-infrared light-sensitive release.Cellular uptake experiments demonstrated that HCu S-TH302@PDA-Ce6/TPP NPs were preferentially enriched in mitochondria under the action of TPP.DCFH-DA and Hypoxyprobe-1^TM were used as the indicators to prove that nanoplatform-mediated photodynamic therapy（PDT）-triggered the generation of intracellular reactive oxygen species（ROS）and intensified hypoxia.MTT experiments indicated that HCu S-TH302@PDA-Ce6/TPP NPs exhibited the highest cytotoxicity upon a combination of 660 and 808 nm laser irradiation,demonstrating that this combined PTT,oxygen-consumed PDT and the successive bioreductive chemotherapy had the optimum therapeutic efficacy.In addition,the nanoplatform also exhibited enhanced therapeutic effects in vivo.Conclusions:A novel mitochondrial-targeted therapeutic system based on HCu S-TH302@PDA-Ce6/TPP NPs was proposed for tumor-specific synergistic hypoxia-activated chemotherapy/PDT/PTT.Both in vitro and in vivo studies demonstrated the greatly improved anticancer activity compared to conventional hypoxia-associated chemotherapy.Our work would promote the clinical application of the combination of HAPs plus phototherapy in tumor treatment.Part Four:Artesunate-based multifunctional nanoplatform for phototherapy/chemodynamic therapy synergistic anticancer therapyObjective:The therapeutic effect of PDT is severely limited by the tumor hypoxic microenvironment and the induced cellular resistance to the PDT treatment.By combination of artesunate（Art）with PDT,a multifunctional nanoplatform based on Z-scheme heterostructured Bi₂S₃@Bi NRs was extablished.The nanoplatform with the ability simultaneously produced O₂ and ROS to alleviate the insufficient O₂ supply in tumor area,which realized synergistic CDT,PTT and PDT with negligible systemic toxicityMethods:Bi₂S₃ NRs were synthesized by a solvothermal method.Bi layer was further grown on Bi₂S₃ NRs through hydrazine treatment of Bi₂S₃ NRs to form Bi₂S₃@Bi NRs.The self-coating polydopamine/ammonium bicarbonate（PDA/ABC）layer on the surface of the Bi₂S₃@Bi NRs served as a p H sensitive substrate for loading Art.Art,a class of endoperoxide-containing sesquiterpene extracted from the Chinese herb Artemisia annua,can be activated by intracellular Fe²⁺ions to generate ROS and thus achieve efficient H₂O₂-independent CDT treatment.Importantly,Art analogues have been reported to inhibit the GPX4 expression,which can greatly enhance the PDT efficiency of Bi₂S₃@Bi NRs.In addition,the cancer-targeting efficiency of the nanoplatform can be increased by HA modification.The structure and properties of Bi₂S₃@Bi@PDA-HA NRs were characterized by TEM,XRD,FTIR and X-ray photoelectron spectroscopy（XPS）.The loading and release of Art were monitored by HPLC.Cytotoxicity and cellular uptake studies were performed using 4T1 cells.The antitumor effect of Bi₂S₃@Bi@PDA-HA/Art NRs in vivo was evaluated in tumor-bearing BALB/c mice,and the tissue morphology was observed by H&E staining.Results:The successful synthesis of nanoplatform（Bi₂S₃@Bi@PDA-HA NRs）was characterized by TEM,XRD,FTIR,XPS.In vitro experiments demonstrated that the nanoplatform exhibited excellent the photothermal performance and possessed the ability to produce effective O₂ and ROS under NIR irradiation.Drug release experiments demonstrated that the platform showed p H/near-infrared light-sensitive release.Cellular uptake experiments demonstrated that the layer of HA endowed the nanoplatform to be internalized by CD44-positive cells through endocytosis.Using DCFH-DA and[Ru（dpp）₃]Cl₂（RDPP）as indicators,we demonstrated the excellent O₂ and ROS supplying ability of Bi₂S₃@Bi@PDA-HA NRs upon NIR irradiation.Glutathione peroxidase 4（GPX4）level was determined by enzyme-linked immunosorbent assay（ELISA）.The result showed that the combination of Art and PDT could effectively inhibit the expression of GPX4 in 4T1 cells and enhance oxidative damage.MTT experiments showed that Bi₂S₃@Bi@PDA-HA/Art NRs exhibited high cytotoxicity in both hypoxic and normoxic conditions,demonstrating the Bi₂S₃@Bi@PDA-HA NRs could rapidly generate O₂ to compensate the O₂ consumption during the PDT process.Notably,the platform also exhibited significant antitumor efficacy in vivo.Conclusions:A multifunctional therapeutic platform based on Bi₂S₃@Bi@PDA-HA/Art NRs was proposed for synergistic PDT/PTT/CDT.Bi₂S₃@Bi NRs with Z-scheme heterostructured NRs enabled the nanoplatform with excellent phothothermal performance and the ability to simultaneously produce O₂ and ROS production for phototherapy of hypoxic tumor.Art encapsulated in the nanoplatform can induce ferroptosis by consumption of extracellular GSH and decreased GPX4 levels to achieve efficient H₂O₂-independent CDT treatment,which in turn significantly enhanced the PDT efficiency of Bi₂S₃@Bi NRs.The excellent anti-tumor activity and good biocompatibility endowed the multifunctional nanoplatforms with great potential in clinical applications.