IMPACT OF DIFFERENT [Tc(N)PNP]-FRAMEWORKS ON THE PHARMACOKINETICS OF THE SMALL RGDFK PEPTIDE

Introduction The [ 99mTc][Tc(N)(PNP)]-system, where PNP is a bisphosphinoamine, is an interesting platform for the development of tumor ‘receptor-specific' agents. Here, we compared the reactivity and impact of three [Tc(N)(PNP)]-frameworks on the stability, receptor targeting properties, biodistribution, and metabolism of the corresponding [ 99mTc][Tc(N)(PNP)]-tagged RGDfK peptide to determine the best-performing agent and to select the framework useful for the preparation of [99mTc][Tc(N)(PNP)]-housing molecular targeting agents. Methods RGDfK pentapeptide was conjugated to Cys and labeled with the [Tc(N)(PNP)]- frameworks (see figure). Radioconjugates were evaluated for their lipophilicity, stability, in vitro and in vivo targeting properties, and performance. Results All compounds were equally easy to synthesize and purify (RCY ≥ 95%). Remarkably, the use of PNP3OH allows to label the peptide at room temperature without significantly reducing the labeling efficiency or stability of the final compound. The main influences of the synthon on the radioconjugate were observed in the in vitro cell binding and in vivo performances. In healthy and xenograft animal models, different pharmacokinetics and tumor accumulation were observed as a function of lipophilicity and sterical hindrance of the synthon. By considering the overall data, the ws[ 99mTc][Tc(N)(PNP3OH)]− and [99mTc][Tc(N)(PNP3)]− synthons perform better in terms of efficiency and biological profile than the [99mTc][Tc(N)(PNP43)]− one. Conclusions In this study, we assessed and compared the effects of the chemical-physical properties of three different [99mTc][Tc(N)(PNP)]-synthons on the synthesis and biological behavior of a small [99mTc][Tc(N)(PNP)]- labeled peptide (i.e. RGDfK) to identify the best performing synthon useful in preparation of a target specific compound. All compounds are equally easy to synthesize and purify. Indeed, the good labeling properties at room temperature of ws[ 99mTc][Tc(N)(PNP3OH)]−framework can be exploited to extend this platform to 99mTc labeling of temperature-sensitive biomolecules. Variation in the nature of the PNP has a profound impact on the overall chemical and physical properties of the radioconjugate. This has a great influence on their biological properties (in vitro cell binding and in vivo biodistribution and pharmacokinetics). Basically, ws[ 99mTc][Tc(N)(PNP3OH)]− and [99mTc][Tc(N)(PNP3)]−tagged RGDfK are more performing, thus more suitable for further radiopharmaceutical applications (1). Funding: The authors acknowledge Associazione Italiana per la Ricerca sul Cancro (AIRC) for financial support (AIRC, IG 2020 ID 24528) and Bracco Imaging SpA. References [1] Bolzati C, Salvarese N, Spolaore B, Vittadini A, Forrer D, Brunello S, Ghiani S, Maiocchi A. Water-Soluble [Tc(N)(PNP)] Moiety for Room-Temperature 99mTc Labeling of Sensitive Target Vectors. Mol Pharmaceutics (2022) doi: 10.1021/acs.molpharmaceut.1c00816.