Synthesis of BODIPY-Peptide Conjugates for Fluorescence Labeling of EGFR Overexpressing Cells.

Abstract

Regioselective functionalization of 2,3,5,6,8-pentachloro-BODIPY 1 produced unsymmetric BODIPY 5, bearing an isothiocyanate group suitable for conjugation, in only four steps. The X-ray structure of 5 reveals a nearly planar BODIPY core with aryl dihedral angles in the range 47.4-62.9°. Conjugation of 5 to two EGFR-targeting pegylated peptides, 3PEG-LARLLT (6) and 3PEG-GYHWYGYTPQNVI (7), under mild conditions (30 min at room temperature), afforded BODIPY conjugates 8 and 9 in 50-80% isolated yields. These conjugates showed red-shifted absorption and emission spectra compared with 5, in the near-IR region, and were evaluated as potential fluorescence imaging agents for EGFR overexpressing cells. SPR and docking investigations suggested that conjugate 8 bearing the LARLLT sequence binds to EGFR more effectively than 9 bearing the GYHWYGYTPQNVI peptide, in part due to the lower solubility of 9, and its tendency for aggregation at concentrations above 10 μM. Studies in human carcinoma HEp2 cells overexpressing EGFR demonstrated low dark and photo cytotoxicities for BODIPY 5 and the two peptide conjugates, and remarkably high cellular uptake for both conjugates 8 and 9, up to 90-fold compared with BODIPY 5 after 1 h. Fluorescence imaging studies in HEp2 cells revealed subcellular localization of the BODIPY-peptide conjugates mainly in the Golgi apparatus and the cell lysosomes. The low cytotoxicity of the new conjugates and their remarkably high uptake into EGFR overexpressing cells renders them promising imaging agents for cancers overexpressing EGFR.

• Authors: Ning Zhao, Tyrslai M Williams, Zehua Zhou, Frank R Fronczek, Martha Sibrian-Vazquez, Seetharama D Jois, M Graça H Vicente
• Journal: Bioconjugate chemistry (Bioconjug Chem) Vol. 28 Issue 5 Pg. 1566-1579 (05 17 2017) ISSN: 1520-4812 [Electronic] United States
• PMID: 28414435 (Publication Type: Journal Article)

Chemical References

• 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene
• Boron Compounds
• Peptide Fragments
• EGFR protein, human
• ErbB Receptors

Topics

• Boron Compounds (chemistry)
• Carcinoma, Squamous Cell (metabolism, pathology)
• Cell Survival
• ErbB Receptors (chemistry, metabolism)
• Fluorescence
• Humans
• Models, Molecular
• Optical Imaging (methods)
• Peptide Fragments (chemical synthesis)
• Tumor Cells, Cultured