Study On In-situ Deformation "Giant" Nanosystem In The Treatment Of Tumor Hunger

Hunger therapy is considered as a "green" emerging therapy with little side effects,which inhibits tumor growth by blocking the supply of tumor nutrients and energy.It has many advantages over other therapies,but there are still many challenges in the treatment of tumors.Firstly,starvation therapy hinders the supply of tumor nutrition and energy,and easily causes metastasis.Secondly,small-sized nanocarriers are easily cleared from the space by lymphatic drainage and extravasation,which results in a short retention time of the nanocarriers in the tumor.Larger-sized nanoparticles are confined to the periphery of tumor tissue,making the drug unable to reach an effective site of action,which greatly reduces the effect of starvation treatment.Thirdly,the efficiency of drug activation is limited by the obstacles crossing cell membranes and escaping from the endosome.These limitations severely limit the development and application of hunger therapy.In order to solve the obstacles to cross cell membranes and escape from the endosome and meet its different needs during circulation,infiltration and accumulation,this project designed the GOx@ZIF-OVA nanosystem.In this nanosystem,glucose oxidase(GOx)is selected as a therapeutic drug.GOx can consume glucose both inside and outside the cell and produce toxic hydrogen peroxide(H2O2),which avoids the limitation of drug cross-cell membrane and escaping from the inner body.Moreover,even in the periphery of tumor tissue,tumor cells can be killed layer by layer.Zeolite imidazolate framework-8(ZIF-8)as a carrier can effectively protect the catalytic activity of the GOx.Surface modified ovalbumin(OVA)enables GOx@ZIF-OVA to spontaneously self-assemble into a giant aggregate "giant" in an acidic environment,which not only meets its different needs in the process of circulation,infiltration and accumulation,but also inhibits metastasis,thus significantly enhancing the therapeutic effect of cancer.This subject was studied from the following three parts.(1)Construction and characterization of in-situ deformed "giant"nanosystem GOx@ZIF nanoparticles were synthesized by a mild one-pot biomimetic mineralization method;then OVA was coated on the surface of GOx@ZIF by physical methods to obtain GOx@ZIF-OVA nanosystem.The structure of the nano-system was characterized by Fourier transform infrared spectroscopy(FT-IR).The particle size potential,morphological characteristics and stability of the system were investigated using a transmission electron microscope(TEM)and a nano-particle size analyzer.Results:GOx@ZIF-OVA nanosystem has strong dispersion and stability at pH 7.4,particle size of 142.2±9.1 nm,and potential of-20.1±2.12 mV.At pH 5.0,GOx@ZIF-OVA nanosystem can be spontaneously assembled into "giant" with particle size of 5779.4±598.3 nm,which can greatly increase the accumulation and retention time of the nanosystem in the tumor site.(2)Study on the antitumor activity of in-situ deformed "giant"nanosystem in vitro Taking mouse breast cancer cells 4T1 as the research object,the antitumor ability of GOx@ZIF-OVA in vitro was studied.The effect of GOx@ZIF-OVA on the proliferation of 4T1 breast cancer cells was examined by the MTT method.The production of H2O2 was examined using a fluorescence microscope.The uptake of cells was measured using a fluorescence microscope.Scratch experiment was used to investigate the effect of GOx@ZIF-OVA on metastasis inhibition of 4T1 breast cancer cells.By constructing 3D tumor cell spheroids,the ability of nanoparticles to inhibit tumor cell growth inside and outside the cell was verified.Results:Compared with other groups,the GOx@ZIF-OVA group has a strong uptake capacity,can produce a large amount of H2O2,and inhibit the metastasis of tumor cells,effectively inhibiting the growth of tumor cells.In addition,3D tumor cell spheroids experiment showed that GOx@ZIF-OVA nanoparticles can kill tumor cells layer by layer,both inside and outside the cell,without being restricted by deep penetration tumor tissues and across cell membranes.(3)Study on the distribution,antitumor activity and anti-metastasis of in-situ deformed "giant" nanosystem in vivo Tumor-bearing BALB/c mice model were constructed,and the distribution and accumulation of the delivery system in the mice were observed using a live imaging fluorescence imager.The antitumor activity and toxic side effects of the nanosystem were examined by measuring changes in tumor volume,body weight,blood glucose and H&E staining pathological tissue sections in mice.The distribution of CD3+CD8+T cells in tumor tissues of tumor-bearing mice was examined by immunofluorescence analysis.Sections were used to verify the effect of GOx@ZIF-OVA on metastasis inhibition.Results:In vivo fluorescence experiment showed that GOx@ZIF-OVA group had stronger fluorescence at 48 h in the tumor,which significantly increased the accumulation of the preparation in the tumor site.In vivo antitumor experiment showed that the GOx@ZIF-OVA group not only significantly improved the tumor suppressive effect,but also had low side effects and high safety.Immunofluorescence experiment and H&E staining experiment showed that GOx@ZIF-OVA group can activate the immune response in vivo,enhance the defense ability in vivo,and effectively inhibit lung metastasis.This project has successfully constructed a pH-stimulated responsive deformable "giant" nanosystem for tumor-specific self-assembly and synergistic cancer treatment.GOx@ZIF-OVA has low hemolysis and high biocompatibility.Not only can it respond to pH in situ self-assembly into large aggregates "giants"to meet its needs for circulation,infiltration and accumulation,but also increase the infiltration of T cells and trigger an effective antitumor immune response and inhibit metastasis.More importantly,in vitro and in vivo studies have shown that GOx@ZIF-OVA can also significantly inhibit cancer without the limitation of crossing cell membranes and escaping from the endosome.