In Vitro and In Vivo Comparison of 3,2-HOPO Versus Deferoxamine-Based Chelation of Zirconium-89 to the Antimesothelin Antibody Anetumab

Abstract

Introduction: [²²⁷Th]Th-3,2-HOPO-MSLN-mAb, a mesothelin (MSLN)-targeted thorium-227 therapeutic conjugate, is currently in phase I clinical trial; however, direct PET imaging using this conjugate is technically challenging. Thus, using the same MSLN antibody, we synthesized 3,2-HOPO and deferoxamine (DFO)-based zirconium-89 antibody conjugates, [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb and [⁸⁹Zr]Zr-DFO-MSLN-mAb, respectively, and compared them in vitro and in vivo. Methods: [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb and [⁸⁹Zr]Zr-DFO-MSLN-mAb were evaluated in vitro to determine binding affinity and immunoreactivity in HT29-MSLN and PDX (NCI-Meso16, NCI-Meso21) cells. For both the zirconium-89 conjugates, in vivo studies (biodistribution/imaging) were performed at days 1, 3, and 6, from which tissue uptake was determined. Results: Both the conjugates demonstrated a low nanomolar binding affinity for MSLN and >95% immunoreactivity. In all the three tumor types, biodistribution of [⁸⁹Zr]Zr-DFO-MSLN-mAb resulted in higher tumor uptake(15.88-28-33%ID/g) at all time points compared with [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb(7-13.07%ID/g). [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb femur uptake was always higher than [⁸⁹Zr]Zr-DFO-MSLN-mAb, and imaging results concurred with the biodistribution studies. Conclusions: Even though the conjugates exhibited a high binding affinity for MSLN, [⁸⁹Zr]Zr-DFO-MSLN-mAb showed a higher tumor and lower femur uptake than [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb. Nevertheless, [⁸⁹Zr]Zr-3,2-HOPO-MSLN-mAb could be used to study organ distribution and lesion uptake with the caveat of detecting MSLN-positive bone lesions. Clinical trial (NCT03507452).

Keywords: 32-HOPO; DFO; PET imaging; antibody conjugate; mesothelin; zirconium-89.