Multifunctional Nanoplatform With Cell Structure Destruction Ability For Antimicrobial/Antitumor Applications

At present,bacterial infections and malignant tumors have become a major threat to human health.Clinically,chemotherapy based on a variety of antibiotics and active anticancer drugs is the main treatment for bacterial infections and tumor growth inhibition.However,the massive and long-term use of antibiotics can lead to the development of strong bacterial resistance,and even the generation of“Superbacteria”,which makes the situation of research on the treatment of bacterial infections more serious.At the same time,the process of tumor treatment is also accompanied by the development of tumor cells’tolerance to anti-cancer drugs,which brings great challenges for the actual clinical personalized and precise treatment.In recent years,in order to overcome the problems existing in traditional clinical treatment methods,a variety of new treatment methods have been developed,including photothermal therapy,photodynamic therapy,chemodynamic therapy,acoustic dynamic therapy,gas therapy and so on.The mechanism effect of the above treatment can be summarized as the degeneration or structural change of the material in the cell caused by heat,reactive oxygen species or physical and mechanical action.Although these novel therapies act on a relatively large number of cellular pathways and have a lower chance of bacterial/tumor resistance through mutation,there are some limitations to these therapies.For photothermal therapy,the treatment process will induce cells to produce stress response,such as the expression of heat shock protein,so as to improve the heat resistance of cells,greatly reducing the treatment efficiency.In addition,the high temperature produced by photothermal therapy will cause thermal damage to normal tissues,and reducing the treatment temperature will not achieve satisfactory therapeutic effect.Therefore,how to balance the killing efficiency and side effects on normal tissues is still worth discussing.Treatment strategies based on the generation of reactive oxygen species,including photodynamic therapy,chemodynamic therapy,and acoustic dynamic therapy,are affected by factors such as oxygen content,hydrogen peroxide content,and GSH level in the microenvironment of lesions.In addition,the accumulation of reactive oxygen species can also cause stress reactions such as autophagy,which makes cells produce“new drug resistance”and limits the efficiency of treatment.It is an important way to solve the problem of bacterial/tumor drug resistance to explore new therapeutic methods or combine existing therapeutic methods to make their advantages and disadvantages complementary.The basic structure of cells,including cell membrane,cytoplasm,nucleus（including the cell wall in bacteria）and other organelles,is the basis for maintaining normal physiological activities of cells.The destruction of cell structure means the loss of normal physiological function of cells and cell death.Among them,the structure of cell membrane plays a fundamental role in the life process of bacteria/tumors by stabilizing the intracellular environment,controlling the entry and exit of substances,regulating cell recognition and signal transmission.Bacterial cell wall can inhibit mechanical and osmotic damage,prevent invasion of macromolecules and assist cell movement and growth.Therefore,this paper innovatively designed three multifunctional nanoplatforms aimed at the destruction of cell structure,and discussed the structural damage effects of reactive oxygen species,carbon monoxide gas,mechanical stress and rigid force on bacterial/tumor cells,especially on cell membrane.In vitro and in vivo models were used to verify the good killing effect of these three multifunctional nanoplatforms on multidrug resistant bacteria and malignant tumors,which has potential clinical research significance.The main research work of this paper includes the following three parts:1.Multifunctional SGQDs-CORM@HA nanosheet cascade activation of photodynamic/gas synergistic destruction of drug-resistant Staphylococcus aureus cell structureIn this study,a multifunctional SGQDs-CORM@HA（SCH）nanosheet was designed to treat wound infections caused by methicillin-resistant Staphylococcus aureus（MRSA）by producing reactive oxygen species and carbon monoxide（CO）gases that can severely damage bacterial structures under light conditions.The multifunctional SCH nanosheets are composed of photodynamic graphene quantum dots（SGQDs）,covalently coupled with CO gas precursor molecules（CORM-401）,and adsorbed hyaluronic acid by electrostatic adsorption.After interaction between SCH and MRSA,the surface modified hyaluronic acid was degraded by bacteria-specific hyaluronidase,and the exposed monolayer graphene nanosheet structure could be embedded into bacteria.Under light conditions,the SCH inside the bacteria can efficiently produce singlet oxygen,and further induce the release of CO from the CORM-401 molecule on the nanosheet.On the one hand,singlet oxygen produced causes oxidative damage to bacteria,and more importantly,its cascade activation releases CO gas,which causes mechanical damage to bacterial structures.The photodynamic/gas damage of the nanosheet on cell membrane and cell wall was investigated by using a fluorescent liposome model,and the dual destruction effect of singlet oxygen and gas on cell structure was proved.Finally,the effectiveness of SCH nanosheets in the treatment of wound infection caused by MRSA was verified by mouse model,which provides a new idea for the development of new antibiotics and the optimization of treatment regimen.2.Mechanical effect triggered by transformable nano-antibiotics are used to disrupt the membrane structure of drug resistant Gram-negative bacteriaIn this study,a transformable nano-antibiotic（TNA）consisting of four independent functional domains,including a hydrophobic core of pyrene molecules,was designed to induce the formation of micellar nanoparticles from TNA molecules.Phenylalanine dipeptide,induced polypeptide byβ-folding assembly;Lysine tripeptide（K3）,as an outer membrane lipopolysaccharide（LPS）recognition unit;Sulfanilamide（Sul）derivatized structure as an outer membrane barrel protein（Bam A）recognition unit.In the presence of recognition units LPS and Bam A,TNA molecules bind LPS and Bam A and are gradually converted into rigid nanofibers.After interaction with Acinetobacter baumannii,TNA nanoparticles bind to LPS and Bam A on the outer membrane of Acinetobacter baumannii and deform into rigid nanofibers.Mechanistic studies have shown that rigid TNA nanofibers can penetrate the outer membrane and cell wall of bacteria due to strong mechanical stress.Further proteomic analysis revealed that the disruption of extracellular membrane structure induced by TNA nanofibers could seriously affect the electron transport and intrinsic biosynthetic and metabolic pathways required for bacterial survival.In vitro cell and in vivo bacterial models have demonstrated that TNA also has a significant killing effect on multidrug resistant Acinetobacter baumannii and other Gram-negative bacteria,including polymyxin resistance.The deformation-disruption strategy used in this study has important research implications in the fight against infections caused by multidrug resistant Gram-negative bacteria.3.Tumor microenvironment-responsive HMn O2/ICG-CF4KY^P is used for tumor cell membrane destruction and immunoactivation therapyIn this study,we designed a hollow manganese dioxide nanocorporeal with tumor microenvironment in response to degradation,and it was loaded with enzyme responsive polypeptide molecule ICG-CF₄KY^p to construct the nanocomposite HMn O₂/ICG-CF₄KY^p（HMIC）.In tumor cells,the hollow manganese dioxide coat is degraded by acidic conditions in lysosomes and high GSH in tumor cells,triggering the release of the drug molecule ICG-CF₄KY^p.Subsequently,the phosphatate bond in ICG-CF₄KY^p molecule is cleaved by the high level of alkaline phosphatase in tumor cells,and the ICG-CF₄KY^ppolypeptide formed by removing the water-soluble phosphate group can self-assemble into rigid nanofibers.The rigid ICG-CF₄KY^p nanofibers generated in situ in tumor cells will affect the integrity of cell membrane structure,and at the same time,the reactive oxygen species produced by photosensitizer ICG in the fiber under the condition of light can further damage the cell membrane,causing damage to tumor cells.The damage to cell membrane caused by nanofibers can activate the immunogenic death pathway of tumor cells,accelerate the release of intracellular antigens,induce a strong immune response,and achieve a synergistic mechanical/photodynamic immunoactivation therapeutic effect.The method used in this study has important reference significance for the clinical research of malignancy related diseases.In summary,this paper designed three multifunctional nanoplatforms aimed at destroying the structure of bacterial/tumor cells.The structural damage effects of carbon monoxide gas,mechanical stress,rigid force and reactive oxygen species on bacterial/tumor cells,especially on cell membrane,were discussed.In vitro and in vivo models were used to verify that these three multifunctional nanoplatforms have good killing effects on multidrug resistant bacteria and malignant tumors.The feasibility of the strategy used in this study for structural disruption of cells was confirmed by in-depth investigation of the antimicrobial/antitumor mechanisms of three functional nanoplatforms.Finally,in vivo animal models were used to verify the effectiveness of the proposed treatment strategies,which provides a research basis for the design and development of novel mechanical therapy and multiple collaborative therapy strategies.