Site-specific labeling of scVEGF with Fluorine-18 for positron emission tomography imaging

Objectives Vascular endothelial growth factor is the most important mediator of angiogenesis. Single-chain (sc)-VEGF protein containing an N-terminal Cys-tag designed for site-specific modification with a variety of imaging and therapeutic moieties. Site-specific labeling of scVEGF with thiol-reactive synthon, N-[2-(4-18F-fluorobenzamido) ethyl] maleimide (18F-FBEM) for VEFGR PET imaging will provide a new tracer which have great potential for clinical translation.

Methods 18F-FBEM-scVEGF was synthesized by site-specific conjugation of N-[2-(4-fluorobenzamide) ethyl] maleimide to a thiol group in Cys-tag of scVEGF at room temperature. The functional activity after labeling was tested by immunofluoescence staining, cellular uptake and efflux. The tumor targeting and in vivo properties were evaluated by biodistribution and microPET studies in tumor-bearing mice were performed.

Results The radiolabeling yield and specific activity of 18F-FBEM-scVEGF was 20.6 ± 15.1% and 2.12 ± 0.43 MBq/µg, respectively. Noninvasive microPET and direct tissue sampling experiments demonstrated that 18F-FBEM-scVEGF had VEGFRs specific tumor uptake in subcutaneous MDA-MB-435 human melanoma model, subcutaneous U87MG human glioma model,, as well as subcutaneous 4T1 murine breast cancer model. The optimal tumor uptake was achieved at 2 hours p.i., which can be partially, but significantly blocked by co-injection of non-labeled scVEGF protein. Overall, 18F-FBEM-scVEGF showed significant high and VEGFRs specific tumor uptake.

Conclusions The scVEGF was site-specifically labeled with 18F via 18F-FBEM synthon and the tracer 18F-FBEM-scVEGF exhibited high-receptor binding affinity and tumor targeting efficacy. 18F-FBEM-scVEGF may have the potential for clinical translation as PET imaging probe.

Research Support This work was supported by the Intramural Research Program (IRP) of the National Institute of Biomedical Imaging and Bioengineering (NIBIB), National Institute of Health (NIH). H.W. holds an Imaging Science Training Fellowship, which is jointly supported by the Radiology and Imaging Sciences Department, NIH Clinical Center and the Intramural Research Program, NIBIB, NIH. We acknowledged the NIH/CC cyclotron facility for isotope production