Construction Of Magnetic Nanoparticles And Its Application In MRI Imaging And Drug/Gene Delivery

Cancer has become the number one disease that seriously endangers human health,and the mortality rate is extremely high.The three commonly used treatments for malignant tumors are surgery,chemotherapy,and radiotherapy.However,surgical treatments are of high risk,great trauma,and complicated procedures.Chemotherapy and radiotherapy can efficiently kill cancer cells,but they can cause serious damage to normal cells,greatly reducing the patient's life quality.At the same time,there are two major defects in a single treatment method:Firstly,it is impossible to detect or diagnose the disease in the early stage;Secondly,is that the therapeutic effect on the lesion location after treatment cannot be evaluated in real time.Therefore,the integration of diagnosis and treatment has great promise for the future.In recent years,the development of nanotechnology has provided new strategy for the realization of theranostics.The nano-theranostic treatment system integrates anti-cancer drugs and imaging agents into nanoparticles.Diagnosis and real-time detection of malignant tumors can be evaluated by magnetic resonance imaging?MRI?,because MRI is a powerful tool for diffusion-weighted imaging of soft tissue.However,there are still challenges in the nano-theranostic system.The MRI is of low resolution,lack of targeting,and the drug carriers can not enable rapid and controlled drug release in target cells.Gene therapy also has these problems.The MRI contrast agent based on superparamagnetic iron oxide nanoparticles?SPIONs?can be modified by hydrophilic polymers,exhibiting good water solubility,excellent biocompatibility,low toxicity,and long circulation time in human body.At the same time,SPIONs are highly sensitive to external magnetic fields and can shorten transverse relaxation time?T2?.Therefore,SPIONs are a class of efficient contrast agents.The receptors of folic acid on many tumor cell membranes as active targets.The drug carriers contain the reduction-responsive disulfide bond and the disulfide bond can be chemically cleaved under GSH or DTT conditions.Therefore,the drug can be released quickly and efficiently in the targeting cell.In summary,the SPIONs based drug carrier with a reduction responsibility can achieve integration of diagnosis and treatment.On the other hand,polyethyleneimine?PEI?as a non-viral vector of the condensing gene can be linked to SPIONs to be a gene carrier for theranostic system.Positively charged PEI compresses negatively charged DNA through electrostatic interactions,and it can protect the DNA from being degraded in the human body,.At the same time,PEI is linked by cross-linking agent with disulfide bond.Disulfide bond can be chemically cleaved and release DNA in the presence of GSH or DTT.Therefore,this strategy in our experiment will provide a new idea for the integration of gene diagnosis and therapy in a nano-sized magnetic gene carrier with redox responsibility.In this paper,we will study the reduction-responsive magnetic durg/gene carriers,and their cell-level treatment and imaging.1.MNPs-DOX drug deliveryIn this dissertation,a multi-functional polymer and SPIONs were used to assemble magnetically integrated nanoparticles by exchanging ligands.It has functions such as blood circulation stability,targeting,controlled drug release,and MRI imaging.N,N-bis?acryloyl?cystamine?BACy?as a cross-linking agent,doxorubicin?DOX?,dopamine?DA?,folate-polyethylene glycol amine?FA-PEG-NH₂?during the preparation process Using methoxy polyethylene glycol amine?mPEG-NH₂?as a comonomer,a multifunctional copolymer polyamidoamine?PAMAM?was synthesized by Michael addition reaction.Then,PAMAM directly assembled to the surface of SPIONs by ligand exchange with SPIONs to form reduced response-type targeted magnetic nanoparticles?MNPs-DOX?loaded with anticancer drugs.As a hydrophilic part,PEG can improve the stability and dispersibility of nanoparticles in aqueous solution,and the cross-linked structure can improve the cycle stability of nanoparticles.In an in vitro drug release experiment of MNPs-DOX,the structure showed that in an environment containing glutathione?GSH?,drug carriers accumulated up to 60%of the drug release within 48 hours.In the absence of GSH,ie in PBS?pH 7.4?,the cumulative release of drug carriers is only 20%.MNPs-DOX has a high drug-loading efficiency of 857?g DOX/mg iron,and MNPs-DOX transverse relaxation R₂ is as high as 126 mM^-1·S^-1,which can be used for MRI imaging and superior to clinically applied products.In cell experiments,MNPs have low toxicity to HUVEC cells?normal cells?,and MNPs-DOX has a highly effective therapeutic effect on HeLa cells?tumor cells?.Confocal experiments of HeLa cells showed that MNPs-DOX can enter into the cell.The above experimental results show that the prepared MNPs-DOX has a promising clinically-used and highly efficient integrated drug carrier for diagnosis and treatment.2.rPEI-DMA@SPIONs gene deliveryIn this paper,cationic polymers and superparamagnetic iron oxide nanoparticles?SPIONs?form gene carriers through exchange ligands,and they can be used as MRI contrast agents to achieve the goal of integrating genetic diagnosis and treatment.Multifunctional copolymer rPEI-DMA was synthesized by Michael addition reaction and N,N-bis?acryloyl?cystamine was used as cross-linker to branch low molecular weight?MW=600?polyethylenimine?PEI?Using dopamine methacrylamide?DMA?as a comonomer,the multifunctional copolymer rPEI-DMA was synthesized by Michael addition reaction.rPEI-DMA and SPIONs were exchanged by ligands to form a polycationic magnetic nanoparticle rPEI-DMA@SPIONs with a reduction-responsive type with a high gene-loading efficiency of 9.77 mg DNA/mg iron.At the same time,the R₂ relaxation value of rPEI-DMA@SPIONs is 81mM^-1·S^-1,which can become a MRI contrast agent.Dynamic light scattering experimental results show that rPEI-DMA@SPIONs can rapidly degrade and release DNA in the environment containing GSH.The results of gel electrophoresis experiments and ethidium bromide fluorescence experiments showed that when the rPEI-DMA@SPIONs-DNA mass ratio?rPEI-DMA@SPIONs/DNA?was 1,the DNA was all compressed,demonstrating excellent rPEI-DMA@SPIONs.DNA carrying capacity;At the same time,the potential of rPEI-DMA@SPIONs and DNA changes with the change of the mass ratio.When the mass ratio is 1,the charge reversal from positive to negative is favorable for endocytosis.The results of cell experiments showed that rPEI-DMA@SPIONs has a low toxicity and a strong gene transfection effect.In summary,rPEI-DMA@SPIONs is expected to become an excellent gene carrier.