Innovation Delivered

SPECT imaging evaluation of ¹¹¹indium-chelated cetuximab for diagnosing EGFR-positive tumor in an HCT-15-induced colorectal xenograft

Bin-Bin Shih^a, Yi-Fang Chang^b, Chun-Chia Cheng^b, Hao-Jhih Yang^c, Kang-Wei Chang^c, Ai-Sheng Ho^a, Hua-Ching Lin^d, Chun Yeh^a, Chun-Chao Chang^e,f

^a Division of Gastroenterology, Cheng Hsin General Hospital, Taipei, Taiwan, ROC

^b Hematology and Oncology, Mackay Memorial Hospital, Taipei, Taiwan, ROC

^c Institute of Nuclear Energy Research, Atomic Energy Council, Taoyuan, Taiwan, ROC

^d Division of Proctology, Cheng Hsin General Hospital, Taipei, Taiwan, ROC

^e Division of Gastroenterology and Hepatology, Department of Internal Medicine, Taipei Medical University Hospital, Taipei, Taiwan, ROC

^f Division of Gastroenterology and Hepatology, Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan, ROC

http://dx.doi.org/10.1016/j.jcma.2017.02.010

Summary

Colorectal cancer (CRC) occurs with high incidence worldwide, but is usually diagnosed in late stage with metastasis by the conventional methods. Epidermal growth factor receptor (EGFR) is overexpressed in 97% of CRC cells, serving a promising diagnostic candidate. In the present study, Cetuximab, an anti-EGFR monoclonal antibody was conjugated with an isotope chelator, diethylene triamine penta acetic acid (DTPA), labeled with ¹¹¹indium (¹¹¹In) and injected to tumor bearing mice. Biological distrubution was investigated by SPECT/CT imaging.

Results revealed that ¹¹¹In-Cetuximab accumulated in the both small (50mm³) and large (250mm³) tumors, whereas the ratio of tumor to muscle in the large tumor was 7.5-fold. The biodistribution data indicated that the ¹¹¹In-cetuximab bound to tumor specifically that was higher than that in other organs. Consequently, ¹¹¹In-cetuximab is suggested to be suitable for early diagnosis and prognostic monitor of EGFR-positive CRC in further clinical practice.

Results from nanoSPECT/CT

• The tumor of the ¹¹¹In-Cetuximab group was apparently observed both in 24h and 48h and higher than that in the ¹¹¹In group.
• ¹¹¹In-Cetuximab majorly accumulated in liver and tumor, otherwise, ¹¹¹In accumulated only in the kidney.
• The tumor to muscle ratio of ¹¹¹In-Cetuximab was measured 7.5-fold, which was higher than that of ¹¹¹In group measured as 3.1-fold, indicating that ¹¹¹In-cetuximab specifically bound to EGFR-positive tumors as a reliable diagnosing agent.
• The result also indicated that ¹¹¹In labeled with Cetuximab through chelator DTPA was easily excreted out the mice better than free ¹¹¹In, suggesting that this labeling method may not lead to accumulation of ¹¹¹In metal in mice.

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.

Nuclear imaging-guided PD-L1blockade therapy increases effectiveness of cancer immunotherapy

Hannan Gao¹, Yue Wu ¹, Jiyun Shi², Xin Zhang¹, Tianyu Liu¹, Biao Hu¹, Bing Jia¹, Yakun Wan³, Zhaofei Liu¹, Fan Wang^1,2,4

¹Medical Isotopes Research Center and Department of Radiation Medicine, State Key Laboratory of Natural and Biomimetic Drugs, School of Basic Medical Sciences, Peking University, Beijing, China

²Key Laboratory of Protein and Peptide Pharmaceuticals, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China

³Shanghai Novamab Biopharmaceuticals Co., Ltd, Shanghai, China

⁴Bioland Laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory), Guangzhou, China

doi:10.1136/jitc-2020-001156

Summary

The interaction between programmed death receptor-1 (PD-1) and its ligand (PD-L1) inhibits the function of effector T cells and the priming of naive T cells, leading to impaired antitumor immunity. Therefore the blockade of PD-1/PD-L1 signaling pathway has been a breakthrough in cancer therapy, but the response rate in solid tumors is only 20-30%. In this study, a new radiolabeled nanobody-based imaging probe 99mTc-MY1523 targeting PD-L1 was developed for the enhanced therapeutic efficacy of PD-L1 blockade immunotherapy.

Results show that the new probe has high binding affinity and specificity to PD-L1. SPECT/CT imaging revealed fast blood clearance, renal-route excretion and satisfactory tumor uptake.

As the timing for PD-L1 blockade therapy is crucial due to dynamic and heterogeneous expression of PD-L1 in tumors, it was essential to prove that SPECT/CT imaging is able to detect changes in the PD-L1 expression. Therefore PD-L1 expression was increased by interferon-γ (IFN-γ) treatment of tumor bearing mice and PD-L1 expression was determined by SPECT/CT imaging and verified by the flow cytometry. It proves that SPECT/CT imaging of 99mTc-MY1523 can be used to monitor PD-L1 expression in tumors in a real time, dynamic and quantitative manner.

The PD-L1 blockade therapy initiated during the therapeutic time window determined by ⁹⁹mTc-MY1523 SPECT/CT imaging significantly enhanced the therapeutic efficacy: the tumor growth was dramatically suppressed, and the survival time of mice was evidently prolonged.

Results from nanoScan SPECT/CT

Three types of tumor cells (4T1, A20 or MC-38) were inoculated subcutaneously into the right flank of BALB/c or C57/BL6 mice, respectively. Four days later they were injected i.t. with PBS or IFN-γ for 5 days, and then were subjected to SPECT/CT imaging.

Mice were injected intraveneously with 18 MBq ⁹⁹mTc-MY1523 and imaged at 2 hours p.i. (n=4) using the nanoScan SPECT/CT system with the following parameters: pinhole SPECT (peak: 140 keV, 20% width; frame time: 25 s), helical CT (50 kVp, 0.67 mA, rotation 210°, exposure time: 300 ms). SPECT and CT images were merged using the Nucline software V.2.0 (Mediso Ltd.). The regions of interest were drawn for the determination of tumor sizes (mm3) and radioactivity (Bq), then the tumor uptake was calculated as percentage injected dose per volume (%ID/cc).

• Results show increased tumor uptake of 99mTc-MY1523 compared to the corresponding control group in all animal models:

When imaging results showed the upregulated PD-L1 expression in tumors after IFN-γ intervention on day 8 and 12 after tumor cell inoculation, the mice were subjected to PD-L1 blockade therapy: they were ip. injected with 200μg αPD-L1 antibody twice with 4 days interval, while using PBS, IFN-γ and αPD-L1 antibody without IFN-γ intervention as controls. Tumor sizes were measured twice a week and calculated as volumes (mm³)=length×width×height/2.

• As shown on the figure below, although IFN-γ intervention expedited the tumor growth, the imaging-guided therapy dramatically improved the therapeutic efficacy. The tumor growth was significantly suppressed, and three of five tumors completely disappeared. Compared to control groups, the survival time of mice in the treated group was also remarkably prolonged.