Early detection of ovarian carcinoma cells

Feasibility of Imaging EpCAM Expression in Ovarian Cancer Using Radiolabeled DARPin Ec1

Anzhelika Vorobyeva^1,2,† , Elena Konovalova^3,†, Tianqi Xu¹, Alexey Schulga^2,3, Mohamed Altai¹, Javad Garousi¹, Sara S. Rinne⁴, Anna Orlova^2,4,5, Vladimir Tolmachev^1,2 and Sergey Deyev^2,3,6,7 

¹ Department of Immunology, Genetics and Pathology, Uppsala University, 751 85 Uppsala, Sweden

² Research Centrum for Oncotheranostics, Research School of Chemistry and Applied Biomedical Sciences, Tomsk Polytechnic University, Tomsk, Russia

³ Molecular Immunology Laboratory, Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia

⁴ Department of Medicinal Chemistry, Uppsala University, Uppsala, Sweden

⁵ Science for Life Laboratory, Uppsala University, Uppsala, Sweden

⁶ Bio-Nanophotonic Lab, Institute of Engineering Physics for Biomedicine (PhysBio), National Research Nuclear University ‘MEPhI’, Moscow, Russia

⁷ Center of Biomedical Engineering, Sechenov University, Moscow, Russia

^† A.V. and E.K. contributed equally

https://doi.org/10.3390/ijms21093310

Summary

Up to 85% of ovarian cancer patients are diagnosed only at advanced stages, when cancer has already spread through the body. The present study aimed to find a more efficient way for diagnosis and treatment using mouse xenograft models.

As epithelial cell adhesion molecule (EpCAM) is overexpressed in 55%–75% of ovarian carcinomas (OC), it might be a promising target. Designed ankyrin repeats protein (DARPin) Ec1 binds to EpCAM with subnanomolar affinity. In the present study, DARPin Ec1 was labeled with ¹²⁵I using N-succinimidyl-para-iodobenzoate (PIB) and injected to mice bearing SKOV-3 or OVCAR-3 xenografts. In vitro experiments showed highly specific binding to ovarian carcinoma cells, moreover, slow internalization, which is essential for in vivo imaging a few hours after injection. In vivo biodistribution analyses of SPECT/CT images suggest that EpCAM on ovarian cancer xenografts is sufficiently accessible to permit DARPin-mediated delivery of cytotoxic payload.

Results from nanoScan SPECT/CT

For establishment of xenografts, 107 of SKOV-3 and OVCAR-3 cells or 5x106 Ramos cells (EpCAM-negative lymphoma xenografts served as specificity control) in 100µl of media were subcutaneously injected in the right hind leg of female Balb/c nu/nu mice. The experiments in mice bearing SKOV-3 and Ramos xenografts were performed 2–3 weeks after implantation. The experiments in mice bearing OVCAR-3 xenografts were performed 7 weeks after implantation.

Mice were injected with ¹²⁵I-PIB-Ec1 (20µg, 1.2MBq for SKOV-3, and 6µg, 2.8MBq for OVCAR-3), SPECT/CT images were acquired 6h pi time later for 20min. with nanoScan SPECT/CT.

• In vitro studies revealed specific binding to SKOV-3 and OVCAR-3 cells; rapid binding and slow dissociation and internalization
• SPECT/CT imaging demonstrated that radiolabeled ¹²⁵I-PIB-Ec1 provided clear visualization of both EpCAM-expressing xenografts. In vivo biodistribution is characterized by high tumor-to-organ ratio, the only organ with noticeable activity were kidneys.