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Objective: Development of companion diagnostics for targeted radionuclide therapy is critical, especially for full-size antibodies with prolonged circulation times. Engineering antibodies to modify their in-vivo pharmacokinetics, such as incorporating neonatal Fc receptor (FcRn)-binding mutations, can potentially enable earlier imaging timing and improved patient stratification. This study aimed to evaluate the impact of FcRn-binding mutations on the in-vitro binding characteristics and in-vivo biodistribution and imaging performance of a CD44v6-targeting full-size antibody, UU-40, labeled with different radionuclides, and to assess its potential as a companion diagnostic.

Methods: The study involved engineering UU-40 with LALA and IAHA mutations, evaluating specific binding, internalization, and affinity using in-vitro cell assays. Biodistribution and imaging studies [PET and single-photon emission computed tomography (SPECT)] were conducted in mice carrying human tumor xenografts in a dual-nuclide setting.

Results: The FcRn mutations (LALA/IAHA) did not affect antibody specificity or affinity, which was target-specific and affinity remained in the subnanomolar range. Biodistribution studies demonstrated that the residualizing radiometal label (¹⁷⁷Lu) resulted in higher liver and spleen uptake compared with the nonresidualizing ¹²⁵I-label, leading to reduced tumor-to-organ ratios. Tumor uptake was higher in A431 xenografts, with peak accumulation at 24 h postinjection. SPECT and PET imaging confirmed superior contrast at later time points (∼24 h) with ¹²⁵I-UU-40_LALA/IAHA, while earlier imaging with ⁶⁸Ga was hindered by increased nonspecific accumulation.

Conclusion: FcRn-binding mutations in full-size antibodies significantly alter their in-vivo pharmacokinetics without affecting binding affinity or specificity. Introducing these mutations enables earlier imaging time points, enhancing the potential for companion diagnostics in clinical settings.