Efficient bioconjugation of 5-fluoro-5-deoxy-ribose (FDR) to RGD peptides for positron emission tomography (PET) imaging of αvβ3 integrin receptor

Abstract

The utility of 5-fluoro-5-deoxyribose (FDR) as an efficient bioconjugation agent for radiolabelling of the RGD peptides c(RGDfK) and c(RGDfC) is demonstrated. The bioconjugation is significantly superior to that achieved with 2-fluoro-2-deoxyglucose (FDG) and benefits from the location of the fluorine at C-5, and that ribose is a 5-membered ring sugar rather than a 6-membered ring. Both features favour ring opening to the aldehydic form of the sugar to promote smooth oxime ligation with aminooxy ether functionalised peptides. [¹⁸F]FDR was prepared in this study by synthesis from fluoride-18 using an automated synthesis protocol adapting that used routinely for [¹⁸F]FDG. c(RGDfK) was functionalised with an aminooxyacetyl group (Aoa) via its lysine terminus, while c(RGDfC) was functionalised with an aminooxyhexylmaleimide (Ahm) through a cysteine–maleimide conjugation. Bioconjugation of [¹⁸F]FDR to c(RGDfC)-Ahm proved to be more efficient than c(RGDfK)-Aoa (92% versus 65%). The unlabelled (¹⁹F) bioconjugates c(RGDfK)-Aoa-FDR and c(RGDfC)-Ahm-FDR were prepared and their in vitro affinity to purified integrin α_vβ₃ was determined. c(RGDfK)-Aoa-FDR showed the greater affinity. Purified “hot” bioconjugates c(RGDfK)-Aoa-[¹⁸F]FDR and c(RGDfC)-Ahm-[¹⁸F]FDR were assayed by incubation with MCF7, LNCaP and PC3 cell lines. In both cases the conjugated RGD peptides showed selectivity for PC3 cells, which express α_vβ₃ integrin, with the c(RGDfK)-Aoa-[¹⁸F]FDR demonstrating better binding, consistent with its higher in vitro affinity. The study demonstrates that [¹⁸F]FDR is an efficient bioconjugation ligand for RGD bioactive peptides.