A Conversation Between Kelsey Martin and Johannes Czernin.

Abstract

1091 Objectives: Triple negative breast cancers (TNBCs) have a higher risk of earlier metastasis and visceral recurrence comparing to other molecular subtypes, and are associated with dismal prognosis. However, there are no specific targets for TNBCs and chemotherapy remains the only available systemic treatment. Hence, cell surface proteins that are specifically expressed in TNBC malignant cells and not expressed in normal breast tissue would be ideal biomarkers, and are potential therapeutic targets for various strategies. Recent publications showed Nectin-4, a membranous protein, was overexpressed in a variety of tumors including TNBCs. Our study aimed to evaluate the feasibility of radioimmunoimaing with 99mTc-labeled Nectin-4 monoclonal antibody (mAb) for breast cancer.Methods: Nectin-4 mAb was labeled with 99mTc by SHNH. The radiolabeling yield and radiochemical purity of 99mTc-Nectin-4 mAb were measured by instant thin layer chromatography (ITLC). Nectin-4 overexpression and underexpression breast cancer cell lines, MDA-MB-468 (TNBC cell line) and MCF-7, were used in this study. Cell uptake assays was performed to evaluate the specificity of binding to Nectin-4 in vitro. The SPECT images and biodistribution study was acquired in mice bearing tumors, and the tumor/muscle (T/M) and tumor/blood (T/B) ratio were calculated.Results: 99mTc-HYNIC-Nectin-4 mAb was successfully synthesized with high radiolabelling yield (65.50 %-79.20%) and radiochemical purity (95.03 ± 2.40%). Western blot and immunohistochemistry showed that the expression level of Nectin-4 in MDA-MB-468 was higher than that in MCF-7. 99mTc-HYNIC-Nectin-4 mAb could be accumulated by MDA-MB-468 cells specifically in vitro with the highest uptake of 4.27 ± 0.09% at 8 h. The SPECT imaging illustrated the MDA-MB-468 tumor had obviously higher 99mTc-HYNIC-Nectin-4 mAb uptake in comparison to MCF-7 tumor,and the MDA-MB-468 blocking group had relatively lower activity accumulation. The results of biodistribution study demonstrated that the MDA-MB-468 tumors had significantly higher uptake of radiolabeled probe with 15.32 ± 1.04% ID/g (36h) comparing to MCF-7 tumors (3.02 ± 0.20% ID/g, p<0.001), which could be inhibited by excess Nectin-4 mAb (4.33 ± 0.48% ID/g, p<0.001). In addition, the mice bearing MDA-MB-468 tumor exhibited superior T/M and T/B ratio (13.19 ± 4.44 and 1.37 ± 0.09) comparing to the MCF-7 group (4.16 ± 1.18 and 0.47 ± 0.05) and MDA-MB-468 blocking group (4.09 ± 0.66 and 0.38 ± 0.03), respectively.Conclusion:99mTc-labeled Nectin-4 mAb can bind to Nectin-4 with high specifically and affinity in vitro and in vivo. Its excellent Nectin-4 targeting provides the potential for multiple anti-tumor strategies translation, including radioimmunological therapy, antibody-drug conjugates (ADC), and antibody/bispecific antibody in various Nectin-4 overexpressing cancers including TNBCs. Data are expressed as mean ± SD (n = 4). Acknowledgements: This work was supported by the National Natural Science Foundation of China (No. 81630049).