PET imaging of colorectal and breast cancer by targeting EphB4 receptor with ⁶⁴Cu-labeled hAb47 and hAb131 antibodies

Objectives Overexpression of EphB4 has been discovered in various cancer cells, and is being evaluated as a therapeutic target. We developed series of ⁶⁴Cu-labeled antibodies for positron emission tomography (PET) imaging of tumor EphB4 expression.

Methods Anti-EphB4 antibodies (hAb47 and hAb131) were conjugated with the ⁶⁴Cu-chelator DOTA through lysine, cysteine, or oligosaccharide on the antibody, respectively. The EphB4 binding activity of these probes was evaluated through the bead-based binding assay with EphB4-alkaline phosphatase (AP). The resulting PET probes were further evaluated in both HT29 and MDA-MB-231 xenografts.

Results All three conjugation methods retained the majority of EphB4 binding activity of the antibodies. Although DOTA-Sug-hAb47 demonstrated highest receptor binding activity based on EphB4 binding assay, the corresponding PET probe was trapped in liver quickly in vivo. In HT29 xenografts, both ⁶⁴Cu-DOTA-Lys-hAb47 and ⁶⁴Cu-DOTA-Cys-hAb47 demonstrated prominent tumor accumulation, which reached maximum at 48 h post injection (18.13 ± 1.73 %ID/g and 11.81 ± 2.05 %ID/g, respectively). In contrast, ⁶⁴Cu-DOTA-Lys-hIgG had low tumor accumulation, which demonstrated the target specificity of EphB4-antibody based probes. Moreover, ⁶⁴Cu-DOTA-Lys-hAb131 (29.48 ± 2.60 %ID/g) demonstrated significantly higher HT29 tumor accumulation than ⁶⁴Cu-DOTA-Lys-hAb47. ⁶⁴Cu-DOTA-Lys-hAb131 was also found to specifically accumulate in MDA-MB-231 tumor model.

Conclusions We have demonstrated that EphB4 could serve as a valid target for colorectal and breast cancer imaging. The success of this approach would be valuable to evaluate disease course and therapeutic efficacy at the earliest stages of anti-EphB4 treatment.