Study On The Preparation And In Vivo Biological Behaviors Of ¹³¹I Labeled Functionalized Graphene Quantum Dots

Due to their excellent physical and chemical properties,graphene quantum dots?GQDs?have been extensively applied in biomedical fields such as drug delivery,bioimaging,and tumor therapy.However,because of the background interference and limited penetration depth of fluorescence in vivo imaging and the detection reults are qualitative or semi-quantitative,most researches about GQDs focus on in vitro study and the in vivo biological behavior,such as metabolism,has not been obtained accurately.Furthermore,the functionization of GQDs is usually considered only to improve biocompatibility or tumor target ability in current studies.However,the two aspects of GQDs must be taken into account together.As a potential target imaging and therapy platform,GQDs should be combined with the diagnostic and therapeutic methods that have been used in clinical applications to develop multimodal theranostics ability.In this thesis,taking advantage of the in vivo imaging and therapy superiority of radionuclides,we labeled GQDs and their functional derivatives with ¹³¹I and studied their biological behaviors.The following results are obtained:GQDs were prepared with a bottom-up method by pyrolysis of citric acid and were characterized detailly.The toxicities of GQDs on HeLa cells and HEK293 cells were evaluated.The results showed that GQDs displayed no obvious toxicity to the two cell lines mentioned above.GQDs were labeled with ¹³¹I by iodogen method,and the labeling conditions were optimized to get a high labeling yield of more that 97%.In vitro partition coefficient and stability tests showed GQDs were hydrophilic and of good stability.Biodistribution of ¹³¹I-GQDs in normal mice and SPECT imaging in HeLa tumor bearing nude mice were studied.It was found that ¹³¹I-GQDs were fast distributed and metabolized,and ¹³¹I-GQDs were mainly metabolized throuth kidney and liver.Because of the enhanced permeability and retention?EPR?effect,¹³¹I-GQDs had a passive targeting effect on tumors.It indicates that GQDs as a drug platform has potential for tumor diagnosis and therapy.In order to improve biocompatibility and targeting ability of GQDs,polyethylene glycol?PEG?and folic acid?FA?were introduced to co-functionalize GQDs.The cytotoxicity and cell fluorescence imaging of GQDs-PEG-FA were tested.The results showed that GQDs-PEG-FA was less toxic than GQDs and could be used as a fluorescent probe for detection of tumor cells overexpressing folic acid receptor.GQDs-PEG-FA was labeled with ¹³¹I and the labeling yield was more that 97%.¹³¹I-GQDs-PEG-FA was hydrophilic and of good in vitro stability.The blood circulation,biodistribution of ¹³¹I-GQDs-PEG-FA in normal mice and SPECT imaging in HeLa tumor bearing nude mice were studied.The results showed that the metabolism of GQDs-PEG-FA was slower than GQDs,which was beneficial to passive targeting of GQDs-PEG-FA at the tumor site via EPR effect.Functionalization of GQDs by PEG-FA lowered the recognition of mononuclear phagocyte system.The SPECT imaging showed that the tumor site can be clearly observed after ¹³¹I-GQDs-PEG-FA administration.It indicates that ¹³¹I-GQDs-PEG-FA can be used as a radioactive probe for detection of tumor cells overexpressing folic acid receptor.DOX was loaded onto ¹³¹I-GQDs-PEG-FA through non-covalent interaction to evaluate the possibility of combination therapy of radionuclide/chemotherapy.The results of in vitro DOX release showed that DOX had higher release rate in the tumor mimic environment compared with the normal physiological environment.The in vitro cytotoxicity of ¹³¹I-GQDs-PEG-FA/DOX was tested,and the results showed that ¹³¹I-GQDs-PEG-FA/DOX had higher cytotoxicity than the radionuclide therapy control group(¹³¹I-GQDs-PEG-FA)and the chemical therapy control group?GQDs-PEG-FA/DOX?.It indicates that ¹³¹I-GQDs-PEG-FA/DOX has good potential for tumor therapy.According to the SPECT imaging of HeLa tumor bearing mice intratumorally injected by ¹³¹I-GQDs-PEG-FA/DOX and the in vitro DOX release results of ¹³¹I-GQDs-PEG-FA/DOX,the tumor therapy experiment was determined by intratumoral injection of ¹³¹I-GQDs-PEG-FA/DOX every three days.¹³¹I-GQDs-PEG-FA/DOX was applied to the combined therapy of HeLa tumor bearing mice and the results showed that either radionuclide therapy or chemical therapy had a certain inhibitory effect on the growth of tumor,and the combination of radionuclide therapy and chemical therapy had synergistic effects for ¹³¹I-GQDs-PEG-FA/DOX had the best results for tumor therapy.Because ¹³¹I is a theranostic nuclide,¹³¹I-GQDs-PEG-FA/DOX has potential to be a theranostic radionuclide/chemotherapy dual-modality radiopharmaceutical.In summary,GQDs and their functional derivatives were labeled by radionuclide(¹³¹I)for the first time in this paper,and their in vivo biological behaviors were studied.The results confirmed the passive targeting effect of GQDs.For the further active targeting effect,a clear SPECT imaging of HeLa tumor can be got by intravenously injction of ¹³¹I-GQDs-PEG-FA and we can propose that¹³¹I-GQDs-PEG-FA are of great potential to be a radioactive probe for detection of tumor cells overexpressing folic acid receptor.¹³¹I-GQDs-PEG-FA/DOX can be applied to the combination therapy of radionuclide/chemotherapy and have the potential to be a theranostic radionuclide/chemotherapy dual-modality radiopharmaceutical.The results of this thesis are expected to extend the application prospect of GQDs in cancer theranostics.