Improving Affinity while Reducing Kidney Uptake of CXCR4‐Targeting Radioligands Derived from the Endogenous Antagonist EPI‐X4

The C-X-C chemokine receptor 4 (CXCR4) is highly upregulated in most cancers, making it an ideal target for delivering radiation therapy to tumors. We previously demonstrated the feasibility of targeting CXCR4 in vivo using a radiolabeled derivative of EPI-X4, an endogenous CXCR4 antagonist, named DOTA-K-JM#173. However, this derivative showed undesirable accumulation in the kidneys, which would limit its clinical use. In this study, we identified that removing a positive charge from the peptide sequence significantly reduced renal uptake. We evaluated a series of optimized derivatives lacking this positive charge, in vitro and in vivo in a xenografted athymic nude mice model, after radiolabeling with ¹⁷⁷Lu. The most promising derivatives were further assessed in vivo after ⁶⁸Ga labeling. Among them, we identified DOTA-JM#173 and D-L¹-DOTA-JM#173, where the D-Ile¹ was replaced by D-Leu¹, two optimized derivatives with a lysine residue removed. These two molecules represent the most advanced DOTA-conjugated ligands derived from EPI-X4 for CXCR4-directed theranostic applications, offering enhanced potential for targeted cancer treatment.