| Objective:This study was aimed to explore the feasibility of99mTc-RGD-BN for breasttumor imaging by evaluating correlated pharmacokinetics and biodistribution in nudemouse models bearing human breast carcinoma xenografts, followed by a pilot cinicaltrial involving recruited healthy volunteers, and further confirm the diagnostic valuesthrough the study of99mTc-RGD-BN SPECT/CT imaging in breast tumor patients.Methods:The novel radioactive small molecular probe--99mTc-RGD-BN was prepared byHYNIC-RGD-BN lyophilized kits in a solution with Na99mTcO-4agents. Weperformed quality control for this purification process via Radioactive thin-layerchromatography (RTLC). The biodistribution and99mTc-RGD-BN SPECT/CTimaging in nude mouse models bearing human breast carcinoma were assessed forfurther usage in tumor imaging.In our pilot clinical trial,6healthy volunteers received a mean740-MBq dose of99mTc-RGD-BN administered through bolus injection in the arm. The safety andfeasibility were estimated through evaluations of pharmacokinetics, biodistribution atdifferent time points on the radiation dose of the volunteers. We also recuited10breast tumor patients for99mTc-RGD-BN SPECT/CT imaging. According to thefindings in pathological analysis, the correlation between radioactive uptake andexpression of integrin αvβ3or/and GRPR was assessed by calculating the ratios oftumor and non-tumor (T/N) and the percentage of positive cells in integrin αvβ3or/and GRPR staining. Then, the diagnostic valutes of99mTc-RGD-BN SPECT/CTimaging for breast tumor was evaluated.Results:The labeling yields was over95%by RTLC analysis. In the T47D nude mousexenograft model with human breast cancer, the99mTc-RGD-BN biodistributionshowed a low uptake and rapid clearance in blood stream (from (0.91±0.21)%ID/g of 1h to (0.07±0.01)%ID/g of8h). The kidneys maintained a high uptake value of(14.54±0.89)%ID/g at8h post-injection, which revealed a renal clearance of99mTc-RGD-BN. Other organs showed moderate or low uptake ratios. While thetumors had relatively high uptake values, as (4.17±1.12)%ID/g,(3.76±0.67)%ID/g,(2.71±0.64)%ID/g and (1.42±0.35)%ID/g at1,2,4and8h, respectively. The T/Nratios in99mTc-RGD-BN imaging of the T47D nude mouse showed that a highest ratioof tumor to blood, tumor to heart and tomor to muscle at4h after injection(17.26±4.06、4.53±1.41and5.17±0.73, respectively).In MDA-MB-435nude mouse xenograft model of human breast cancer, afterinjection of99mTc-RGD-BN, the biodistribution showed a low uptake and rapidclearance in blood (from (0.88±0.22)%ID/g of1h to (0.08±0.01)%ID/g of8h). Thekidneys maintained a high uptake value of (14.55±0.91)%ID/g at8h after injectionrevealling the renal clearance of99mTc-RGD-BN. The other organs showed moderateor low uptakes. The tumors had relatively high uptake values of (4.55±1.18)%ID/g,(3.95±0.69)%ID/g,(2.86±0.65)%ID/g and (1.56±0.32)%ID/g at1,2,4and8h,respectively.The T/N ratios in99mTc-RGD-BN imaging of the MDA-MB-435nudemouse showed that the highest ratio of tumor to blood, tumor to heart and tomor tomusle apeared at4h after injection (18.56±3.98、5.18±1.37and5.53±0.70,respectively).The pharmacokinetics of6healthy volunteers who accepted99mTc-RGD-BNinjection quickly removed the radioactive tracer in the blood. The radioactive uptakerate of the blood (43.05±6.24)%at10minutes, and the blood clearance extended byabout85%in60minutes. The excreted urine radioactivity showed a peak within inthe first24hours, about (73.56±2.04)%of total injection dose excretion in urine.After injection of99mTc-RGD-BN, at different time points, SPECT body imagingof in vivo biodistribution showed that a higher intake of radioactive in human bodyliver, kidneys and gastrointestinal. The basal body radiation absorbed dose for theradioactive distribution was2.17×10-3mSv/MBq, while the radiation absorbed dosein kidney was2.55×10-2mSv/MBq.In our study, no significant clinical abnormalities or abnormal biochemicaloutputs were recorded. Accepting99mTc-RGD-BN SPECT/CT imaging, sevenmalignant cases were diagnosed, of the10cases further confirmed by pathological examinations, including5invasive ductal carcinoma cases,1ductal carcinoma in situand1invasive lobular carcinoma. During the study,7malignant lesions showedaccumulation of abnormal radioactivities with T/N ratios ranging from2.56to4.43.The rest3were diagnosed as benign cases with2fibroadenoma and1adenosis. Noaccumulation was found in the3benign cases with T/N ratios ranging from1.01to1.10. The diagnostic outcomes of99mTc-RGD-BN SPECT/CT imaging were coincidedwith the ultrasound and mammography.Further, immunohistochemical staining results showed that integrin αvβ3and/orGRPR was/were expressed in all malignant cases (1case expressed αvβ3,1caseexpressed GRPR,4cases expressed both αvβ3and GRPR). In two imaging positivecases, an axillary lymph metastastic legion and a bone metastastic lesion showedpositve expressions of the two molecules. The T/N ratio and integrin αvβ3and/orGRPR correlation factor (r=0.958) had shown significantly positive connections withthe percentage of positive cells in tumor regions.Conclusion:New radioactive molecular probe99mTc-RGD-BN has been lyophilized for kitpreparation, and the tagging process is simple and fast to ensure that the labeling rates.The novel integrin αvβ3and GRPR as targets of novel radioactive molecular probes,in a nude mouse model of human breast cancer or normal human volunteers, hadshowed ideal pharmacokinetic and biodistributional characteristics, as well as a goodradiation dosimetry safety. Throught this study,99mTc-RGD-BN had been shown acertain diagnostic value in breast cancer cases, especially in the detection of distantmetastases compared to the traditional inspection methods in patients with breastplaceholder. Positive correlation between99mTc-RGD-BN imaging T/N ratios ofintegrin αvβ3and GRPR expression confirms that99mTc-RGD-BN as a new type ofradioactive molecules for in vivo imaging. |
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