Tumor Targeting Nano CT Contrast Agent With Chemotherapy Drugs

Objective: With the development of medical care and the improvement of human living standards,many diseases that once threatened people are no longer a problem.But in recent years,tumor diseases still pose a serious threat to human health.The selectivity of traditional antitumor drugs is poor,the so-called kill one thousand self harm eight hundreds will be generated.The cancer cells are not only killed,but the normal cells are also damaged,which often lead to serious systemic toxicities and side effects.Therefore,it is urgent to find an intelligent drug carrier which is highly selective and can realize the integration of tumor diagnosis and treatment.Computed tomography(CT)is one of the non-invasive imaging diagnostic techniques widely used in clinical practice,and the contrast of soft tissue should be enhanced by using CT contrast agent.To improve the imaging performance of the contrast agent and reduce their side effects,clinical academics and contrast agent developer have explored continuously for several decades.A large amount of researches focused on the developments from iodine substituted small molecules in clinical application into nano-sized iodinated CT contrast agents.In addition,to increase their contrast,some metal and inorganic nanoparticles with high X-ray absorption performance were introduced into the researches of X-ray CT contrast agent.For example,gold nanoparticles have been researched extensively due to their high atomic number and their high X-ray attenuation effect.In this work,the multifunctional iodine-containing nanoparticles with Au doping were prepared for using as X-ray CT contrast agents and antitumor drug carriers to achieve accurate diagnosis and treatment of tumor cells.Content : The multifunctional iodine-containing nanoparticles P(MATIB-co-MBA-co-GMA)-FA-Au NP were successfully prepared by multiple steps,including the synthesis of iodine-containing monomer compounds,the preparation of the crosslinked iodine-containing nanoparticles,the surface modification with amino groups,the conjugation of folic acid(FA)molecules,and the gold nanoparticles depositing in situ.The morphologies and structures of the nanoparticles were characterized by using transmission electron microscopy(TEM),ultraviolet-visible spectrophotometer(UV-vis)and fourier infrared spectrometer(FT-IR).Their sizes and zeta potentials were measured in water using a laser scattering spectrometer.Their performances used as CT contrast agent were evaluated by investigating their X-ray attenuation effects.Their performances used as drug carriers were evaluated via investigating their drug loading and releasing.The cellular uptakes and cytotoxicities of the nanoparticles were investigated in vitro.Methods: In this study,the iodine-containing polymer nanoparticles P(MATIB-co-MBA-co-GMA)-FA-Au NP were synthesized via precipitation polymerization.First of all,the crosslinked polymer nanoparticles P(MATIB-co-MBA)were prepared with 2-methacryl(3-amide-2,4,6-triiodobenzoic acid)(MATIB)as monomers and N,N-methylenebis(acrylamide)(MBA)as crosslinker.Glycidyl methacrylate(GMA)as comonomers were added in the meantime to introduce epoxy groupson the surface of the nanoparticles.In the second stage,the surfaces of P(MATIB-co-MBA-co-GMA)nanoparticles were introduced amino groups by the ring-opening reaction with EDA.Subsequently,FA as tumor target molecules were modified onto the surface of the nanoparticles via the formation of amide linkages between the amino groups on the surface of nanoparticles and the carboxylic acid groups of FA to increase theactive uptake of nanoparticles.Finally,the gold nanoparticles were deposited on the generated nanoparticles P(MATIB-co-MBA-co-GMA)-FA in situ.Thus multifunctionalized iodine-containing nanoparticles with Au doping were prepared to use as X-ray CT contrast agent and anti-cancer drug carriers with tumor targeting for tumor diagnosis and treatment.Next,the comprehensive characterizations of the polymer nanoparticles obtained from each step were given.Firstly,TEM was used to characterize their surface morphologies,particle sizes and particle size distributions.The FT-IR spectra and UV-vis spectra were used to characterize their structures.The particle sizes in solution and the zeta potentials of the nanoparticles were characterized by a laser scattering spectrometer.The content of I and Au in the nanoparticles were measured by inductively coupled plasma mass spectrometry(ICP-MS).Their performances used as drug carriers were evaluated by investigating the loading and releasing of the antitumor drug doxorubicin hydrochloride(DOX).The targeted cellular uptake sand drug delivery propertity of the P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles were investigated using breast cancer cells(MCF-7)in vitro by confocal fluorescence microscope and flow cytometryassay.The cytotoxicity of DOX loaded P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles were investigated by MTT assay.Results: From the results of TEM,FT-IR spectra,UV-vis spectra,size and size distribution,and zeta potential,the iodine-containing nanoparticles P(MATIB-co-MBA-co-GMA)-FA-Au NP were prepared successfully.From the observation of TEM images,the nanoparticles from each step had regular spherical shape,uniform size and good dispersity.The average size of P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles is about 135 nm.Compared to that of P(MATIB-co-MBA-co-GMA)nanoparticles,the zeta potential of the P(MATIB-co-MBA-co-GMA)-EDA nanoparticles changed to positive charging from the results of particle analyser,which indicated that the EDA was successfully modified on the surface of P(MATIB-co-MBA-co-GMA)nanoparticles.The strong peak at 280 nm which attributed to the characteristic absorbance of FA molecule was shifted to 304 nm in the UV–vis spectrum of P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles,suggested that FA molecules was successfully connected to the P(MATIB-co-MBA-co-GMA)-EDA nanoparticles.The results of in vitro X-ray CT imaging showed that the X-ray attenuation effect of the nanoparticles doping with Au NP was significantly enhanced.The experimental results of drug loading showed that the DOX loading capacity of the P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles could reach 51.3%,and the corresponding encapsulation rate was 34.2%.The DOX release rate in p H 6.0 phosphate buffer solution(PBS)was significantly higher than that in p H 7.4 buffer solution.The results showed that the DOX release from the nanoparticles was dependent on p H values.The results from laser confocal fluorescence microscopy and flow cytometry analyzer showed that the fluorescence intensity of the tumor cells treated with DOX-loaded P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles was obviously higher than that of the tumor cells treated with DOX loaded P(MATIB-co-MBA-co-GMA)-Au NP nanoparticles,indicating the targeted delivery of DOX by the P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles.The results of cytotoxicity experiment showed that the P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles showed no obvious toxicity at less than 100 μg/m L.Compared to the nanoparticles without FA modification,the nanoparticles with FA modification could carry more anti-tumor drugs into tumor cells and kill tumor cells more effectively.Conclusion : The P(MATIB-co-MBA-co-GMA)-FA-AuNP nanoparticles with uniform size and good stability were prepared.The drug-loaded nanoparticles exhibited the targeted drug delivery in tumor cells,p H-dependent controlled release and low cytotoxicity.In vitro drug delivery study indicated that the FA-conjugated nanoparticles could deliver DOX into MCF-7 cells more efficiently than the nanoparticles without functionalization of FA.And the DOX-loaded P(MATIB-co-MBA-co-GMA)-FA-Au NP nanoparticles showed more efficient for killing tumor cells than that of P(MATIB-co-MBA-co-GMA)-Au NP nanoparticles.The results of X-ray CT imaging showed that the X-ray attenuation effect of the nanoparticles doping with Au NP was significantly enhanced.The multifunctional iodine-containing nanoparticles with Au doping have potential to be using as X-ray CT contrast agents and antitumor drug carriers to achieve accurate diagnosis and treatment of tumor cells.