Radiolabeling and Preclinical Evaluation of Therapeutic Efficacy of ²²⁵Ac-ch806 in Glioblastoma and Colorectal Cancer Xenograft Models

Abstract

The epidermal growth factor receptor (EGFR) protein is highly expressed in a range of malignancies. Although therapeutic interventions directed toward EGFR have yielded therapeutic responses in cancer patients, side effects are common because of normal-tissue expression of wild-type EGFR. We developed a novel tumor-specific anti-EGFR chimeric antibody ch806 labeled with ²²⁵Ac and evaluated its in vitro properties and therapeutic efficacy in murine models of glioblastoma and colorectal cancer. Methods: ²²⁵Ac-ch806 was prepared using different chelators, yielding [²²⁵Ac]Ac-macropa-tzPEG₃Sq-ch806 and [²²⁵Ac]Ac-DOTA-dhPzPEG₄-ch806. Radiochemical yield, purity, apparent specific activity, and serum stability of ²²⁵Ac-ch806 were quantified. In vitro cell killing effect was examined. The biodistribution and therapeutic efficacy of ²²⁵Ac-ch806 were investigated in mice with U87MG.de2-7 and DiFi tumors. Pharmacodynamic analysis of tumors after therapy was performed, including DNA double-strand break immunofluorescence of γH2AX, as well as immunohistochemistry for proliferation, cell cycle arrest, and apoptosis. Results: [²²⁵Ac]Ac-macropa-tzPEG₃Sq-ch806 surpassed [²²⁵Ac]Ac-DOTA-dhPzPEG₄-ch806 in radiochemical yield, purity, apparent specific activity, and serum stability. [²²⁵Ac]Ac-macropa-tzPEG₃Sq-ch806 was therefore used for both in vitro and in vivo studies. It displayed a significant, specific, and dose-dependent in vitro cell-killing effect in U87MG.de2-7 cells. ²²⁵Ac-ch806 also displayed high tumor uptake and minimal uptake in normal tissues. ²²⁵Ac-ch806 significantly inhibited tumor growth and prolonged survival in both U87MG.de2-7 and DiFi models. Enhanced γH2AX staining was observed in ²²⁵Ac-ch806–treated tumors compared with controls. Reduced Ki-67 expression was evident in all ²²⁵Ac-ch806–treated tumors. Increased expression of p21 and cleaved caspase 3 was shown in U87MG.de2-7 and DiFi tumors treated with ²²⁵Ac-ch806. Conclusion: In glioblastoma and colorectal tumor models, ²²⁵Ac-ch806 significantly inhibited tumor growth via induction of double-strand breaks, thereby constraining cancer cell proliferation while inducing cell cycle arrest and apoptosis. These findings underscore the potential clinical applicability of ²²⁵Ac-ch806 as a potential therapy for EGFR-expressing solid tumors.