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Quantitative radioimmunoPET imaging of EphA2 in tumor-bearing mice
Weibo Cai1, Alireza Ebrahimnejad, Kai Chen
1Department of Radiology and Bio-X Program, The Molecular Imaging Program at Stanford, Stanford University School of Medicine, 1201 Welch Road, Stanford, CA 94305-5484, USA.
Purpose:
EphA2 receptor tyrosine kinase is significantly overexpressed in a wide variety of cancer types. High EphA2 expression has been correlated with increased metastatic potential and poor patient survival. Although many recent reports have focused on blocking the EphA2 signaling pathway in cancer, the in vivo imaging of EphA2 has not yet been investigated.
Methods:
We labeled 1C1, a humanized monoclonal antibody against both human and murine EphA2, with (64)Cu through the chelating agent 1,4,7,10-tetraazacyclododecane N,N',N'',N'''-tetraacetic acid (DOTA) and carried out positron emission tomography (PET) imaging of eight tumor models with different EphA2 expression levels. Western blotting of tumor tissue lysate was performed to correlate the EphA2 expression level with (64)Cu-DOTA-1C1 uptake in the tumors. Immunofluorescence staining and biodistribution studies were also carried out to validate the in vivo results.
Results:
The radiolabeling yield was 88.9 +/- 9.5% (n = 7) and the specific activity of (64)Cu-DOTA-1C1 was 1.32 +/- 0.14 GBq/mg of 1C1 mAb. The antibody retained antigen-binding affinity/specificity after DOTA conjugation as measured by FACS analysis. The uptake of (64)Cu-DOTA-1C1 in CT-26 tumors was as high as 25.1 +/- 2.5 %ID/g (n = 3) at 18 h post injection. (64)Cu-DOTA-IgG, an isotype-matched control, exhibited minimal non-specific uptake in all eight tumor models. In vivo EphA2 specificity of (64)Cu-DOTA-1C1 was confirmed by successful blocking of CT-26 tumor uptake by unlabeled 1C1. Most importantly, the tumor uptake value obtained from PET imaging had excellent linear correlation with the relative tumor tissue EphA2 expression level measured by Western blot, where r (2) equals 0.90 and 0.92 at 18 h and 42 h post injection, respectively.
Conclusion:
The tumor uptake of (64)Cu-DOTA-1C1 measured by microPET imaging reflects tumor EphA2 expression level in vivo. This is, to our knowledge, the first report of quantitative radioimmunoPET imaging of EphA2 in living subjects. Future clinical investigation of (64)Cu-DOTA-1C1 is warranted.
Insights
This study developed a novel method for in vivo imaging of EphA2 receptor tyrosine kinase, a protein overexpressed in many cancers. The developed imaging agent, (64)Cu-DOTA-1C1, accurately reflects EphA2 expression levels in tumors, paving the way for future clinical applications.
Area of Science:
- Oncology
- Radiochemistry
- Molecular Imaging
Background:
- EphA2 receptor tyrosine kinase is significantly overexpressed in various cancers, correlating with increased metastasis and reduced patient survival.
- Targeting EphA2 is a promising strategy, but in vivo imaging of EphA2 expression has remained unexplored.
Purpose of the Study:
- To develop and validate a novel positron emission tomography (PET) imaging agent for quantitative in vivo visualization of EphA2 expression.
- To assess the correlation between the uptake of the imaging agent and EphA2 levels in preclinical cancer models.
Main Methods:
- A humanized monoclonal antibody, 1C1, targeting EphA2 was labeled with Copper-64 (64Cu) using the chelator DOTA.
- Positron emission tomography (PET) imaging was performed on eight tumor models with varying EphA2 expression.
- Western blotting, immunofluorescence staining, and biodistribution studies were used to validate imaging results.
Main Results:
- The radiolabeled antibody, (64)Cu-DOTA-1C1, demonstrated high tumor uptake in EphA2-expressing tumors (25.1 ± 2.5 %ID/g at 18 h).
- The imaging agent showed high specificity, with minimal uptake in control IgG and successful blocking studies.
- PET imaging results showed an excellent linear correlation (r² = 0.90-0.92) with EphA2 expression levels determined by Western blot.
Conclusions:
- Quantitative radioimmunoPET imaging of EphA2 is feasible in vivo using (64)Cu-DOTA-1C1.
- The developed imaging agent accurately reflects tumor EphA2 expression levels.
- This study represents the first report of quantitative in vivo EphA2 imaging, warranting further clinical investigation.
