| 【Background】Gastric cancer, with its high incidence and mortality, has developed into a serious healthy problem in the world. Accurate screening and diagnosing of precancerous diseases, precancerous lesions and early gastric cancer is crucial to reduce the mortality rate of gastric cancer. However, the early symptoms of gastric cancer is not typical, the existing technologies are difficult to achieve accurate diagnosis of small cancerous lesions. Therefore, find and establish a method for early diagnostic of gastric cancer with high specificity and sensitivity is urgently needed to reduce mortality, especially before the change of the anatomical structure. MG7 monoclonal antibody, a specific gastric cancerassociated antibody was primary identified by D Fan et al, is distinguished only in the presence of gastric cancer lesions with high sensitivity and specificity, is a promising marker for early diagnostic of gastric cancer. Molecular imaging is an interdisciplinary of molecular biology, biochemistry, imaging studies and other disciplines. Molecular imaging makes it possible to monitor the pathological processes of cancer in real-time at the cellular and molecular levels and has a great value of clinical use.【Purpose】Use MG7 monoclonal antibody as a target molecule to prepare a radioactive probe, evaluate the physicochemical properties and imaging capabilities of the probe at the cellular and molecular levels and establish a diagnostic imaging method of gastric cancer based on MG7 monoclonal antibody.【Methods】1. Preparation and qualification of 68Ga-NOTA-MG7 probe68Ga-NOTA-MG7 probe was prepared by an indirect labeling method using NOTA as a bridge. Labeling efficiency was measured using paper chromatography. Then the stability, cytotoxicity and partition coefficient were tested.2. In vitro and in vivo studies of 68Ga-NOTA-MG7 probeFor immunofluorescence staining study, the MG7 antibody were incubated with different gastric cancer cell lines and immortalized gastric epithelial cell line to observe the distribution and targeting efficacy. For the cell uptake and efflux study, the probe was incubated with BGC-823 gastric cancer cells, and cell lysates were collected at different time points, and then measured the radioactivity. For the PET/CT imaging and Cherenkov imaging, 68Ga-NOTA-MG7 or 68Ga-NOTA-MG7/MG7 was injected into each mouse under isoflurane anesthesia through tail vein. The images were acquired at 30, 60 and 90 min. For each nano PET/CT and Cherenkov scan, regions of interest(ROIs) were drawn over the tumor, liver and kidney on the decay-corrected whole-body coronal images to observe the distribution of probe.3. Biodistribution study of 68Ga-NOTA-MG7 probe12 tumor bearing nude mice were divided into A, B, C, D groups. For A, B, C groups, mice were intravenously injected with 68Ga-NOTA-MG7 probe(7.4 MBq / 200 μl), for D group, mice were injected with 68Ga-NOTA-MG7 / MG7 mixture(68Ga-NOTA-MG7 injection volume 7.4 MBq / 200 μl, MG7 injection volume: 20 mg / kg). For A, B, C groups, mice were sacrificed at 30 min, 60 min, 90 min respectively. For D group, mice were sacrificed in 60 min. Major organs and tumors were obtained and the radioactivity was measured by a γ-counter.【Results】1. Preparation and qualification of 68Ga-NOTA-MG7 probe68Ga-NOTA-MG7 probe was prepared by chemical coupling methods. The radiolabeling efficiency was approximately 99% without purification. Serum stability experiment confirmed that after 120 min of incubation, more than 97% of 68Ga-NOTAMG7 remained intact, indicating it has a good stability. Octanol-water partition coefficient(log P) of 68Ga-NOTA-MG7 was-2.42±0.11, indicating that 68Ga-NOTA-MG7 was highly hydrophilic. MTT cytotoxicity test showed that compared with 18F-FDG, the probe showed no significant cytotoxicity in the experimental dose.2. In vitro and in vivo studies of 68Ga-NOTA-MG7 probeImmunofluorescence staining study confirmed that MG7 antibody has a specific binding ability to gastric cancer cells. Cellular uptake and efflux experiments showed that the probe can achieve maximum uptake at 60 min. PET / CT imaging confirmed that Predominant uptake of the probe can be observed in tumor, liver and kidneys. For tumor, the maximum uptake was at 60min(2.53± 0.28 % ID / g). The blocking experiments confirmed that when the 68Ga-NOTA-MG7 probe co-injected with MG7 antibody, tumor uptake was significantly reduced. Cherenkov imaging showed a specific uptake of the probe in tumor, and the results were consistent with PET/CT imaging.3. Biodistribution study of 68Ga-NOTA-MG7 probeBiodistribution study confirmed that after intravenous injection of 68Ga-NOTA-MG7 probe, the maximum tumor uptake was observed at 60min(2.72 ± 0.40% ID / g). The blocking experiment showed that tumor uptake was significantly reduced when the probe co-injected with MG7 antibody. Except for the tumor tissue, Predominant uptake of 68GaNOTA-MG7 was also visualized in the liver and kidneys.【Conclusion】Gastric cancer specific probe 68Ga-NOTA-MG7 was successfully prepared with good physical and chemical stability and low toxicity. The probe has a great in vivo imaging capability. Further modified for the probe, is expected to achieve the diagnosis and treatment goal simultaneously. The furture of the probe is very promising. |
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