Trivalent ligands for CXCR4 bearing polyproline linkers show specific recognition for cells with increased CXCR4 expression.

The assembly status of G protein-coupled receptors (GPCR) on the cell surface is of interest because the multimerization of GPCR could play pivotal roles in cellular functions. A bivalent ligand with polyproline linkers for CXCR4 has been shown to serve as a "molecular ruler" as a result of the rigid structure of polyproline helices. To expand the utility of the ligands with rigid linkers and explore the possible multimeric forms of GPCR, trivalent ligands with polyproline helices were newly designed and synthesized. The binding affinities of the trivalent ligands for CXCR4 suggested that the ligands recognize the dimeric form of CXCR4 on the cell surface. The fluorescence imaging and analysis by flow cytometry revealed that the ligand with 9 proline linkers binds to CXCR4 with remarkable specificity. The results of the present study suggest that the ligand design with rigid linkers is useful in the multimeric form, but the design of trivalent ligands requires different strategic approaches.