Synthesis, Purification, and Tumor Cell Uptake of ⁶⁷Ga-Deferoxamine−Folate, a Potential Radiopharmaceutical for Tumor Imaging

TLDR

The authors synthesized DF‑folate by carbodiimide coupling of folic acid to deferoxamine, separated the resulting α and γ isomers by anion‑exchange chromatography, radiolabeled the γ‑isomer with 67Ga, and evaluated its uptake in folate‑receptor‑positive KB cells. The γ‑isomer of DF‑folate binds the folate receptor with affinity similar to free folic acid, accumulates rapidly in KB cells (t½ ≈ 3 min) in a time‑, temperature‑, and concentration‑dependent manner, and its uptake is competitively inhibited by free folic acid, indicating specific receptor‑mediated internalization and potential use as a diagnostic agent for folate‑receptor‑positive tumors.

Abstract

The vitamin folic acid was covalently linked to the chelating agent deferoxamine (DF) via an amide bond using a simple carbodiimide coupling reaction. A mixture of two isomers, DF−folate(α) and DF−folate(γ), was produced involving the α- and γ-carboxyl group of folic acid, respectively. These two isomers were separated by anion-exchange chromatography using a NH4HCO3 gradient. Competitive binding studies revealed that only the DF−folate(γ) is recognized by the folate receptor on KB cells, interacting with an affinity comparable to unconjugated folic acid. The DF−folate conjugates were radiolabeled with the γ-emitting radionuclide 67Ga3+ and tested for uptake by cultured KB cells overexpressing the folate receptor. The cellular accumulation of 67Ga-DF−folate(γ) complex was found to be time-, temperature-, and concentration-dependent. The 67Ga-DF−folate(γ) tracer exhibited rapid uptake kinetics in cell culture with a t1/2 of ∼3 min. The KB cell association of 67Ga-DF−folate(γ) was competitively blocked by free folic acid, indicating that uptake of the 67Ga-DF−folate(γ) was specifically mediated by the folate receptor. Since the folate receptor is overexpressed on the surfaces of many neoplastic cells, these results suggest that 67Ga-DF−folate(γ) complex might be useful as a diagnostic agent for noninvasive imaging of folate receptor-positing tumors.