Development and Preclinical Evaluation of Dual-Target 68Ga/177Lu-Labeled FAP-Bisphosphonate Radioligands with Enhanced Osteoma Retention

Abstract

Bone metastases and associated skeletal-related events (SREs) severely compromise the survival and quality of life of patients with advanced cancer. Although bisphosphonates enable efficient bone targeting, their radiolabeled analogues exhibit limited uptake in osteolytic lesions due to the low hydroxyapatite content. Fibroblast activation protein, highly expressed in cancer-associated fibroblasts within the tumor microenvironment, provides tumor specificity and may enhance tracer accumulation in osteolytic bone metastases. To overcome this limitation, we developed two heterodimeric [⁶⁸Ga]Ga/[¹⁷⁷Lu]Lu-DOTA-FAPI-bisphosphonate radioligands ([⁶⁸Ga]Ga/[¹⁷⁷Lu]Lu-DFP-1/2) by integrating a FAP-targeting moiety with a bisphosphonate backbone. Both compounds were synthesized with high radiochemical purity (>95%) and excellent in vitro stability. In A549-FAP xenograft models, dynamic PET/CT imaging revealed comparable tumor uptake of [⁶⁸Ga]Ga-DFP-1/2 and [⁶⁸Ga]Ga-FAPI-04 up to 2 h postinjection, with [⁶⁸Ga]Ga-DFP-2 demonstrating faster clearance from nontarget organs. In tibial A549-FAP osteogenic models, [⁶⁸Ga]Ga-DFP-2 showed significantly higher uptake (4.88 ± 1.01% ID/mL) than mono-FAP or monobisphosphonate tracers (p < 0.05). In osteolytic A549 models, [⁶⁸Ga]Ga-DFP-2 uptake (3.16 ± 0.82% ID/mL) also exceeded that of monobisphosphonate tracers (p < 0.01). Biodistribution studies confirmed prolonged [¹⁷⁷Lu]Lu-DFP-2 retention (26.94 ± 4.40% ID/g at 168 h). Collectively, these findings demonstrate that [⁶⁸Ga]Ga/[¹⁷⁷Lu]Lu-DFP-2 possesses favorable dual bone- and tumor-targeting characteristics and holds strong translational promise as a novel theranostic agent for the imaging and treatment of lung cancer bone metastases.