title: In vivo noninvasive optical imaging of receptor-mediated RGD internalization using self-quenched Cy5-labeled RAFT-c(-RGDfK-)(4).
abstract: We reported that regioselectively addressable functionalized template (RAFT)-c(-RGDfK-)(4 )presenting four cyclic (Arg-Gly-Asp) (cRGD) peptides targets integrin alpha(V)beta(3) with an improved specificity compared with monomeric cRGD. In this study, we improved this vector by creating a "stealth" molecule in which a fluorescence quencher (Q) is linked to Cy5 via a disulfide bond (-SS-). RAFT-c(-RGDfK-)(4)-Cy5-SS-Q fluorescence is quenched unless activated by reduction during internalization. RAFT-c(-RGDfK-)(4)-Cy5-SS-Q fluorescence was negligible when compared with the control but totally recovered after cleavage of the disulfide bridge. Confocal microscopy showed that only the intracellular Cy5 signal could be detected using RAFT-c(-RGDfK-)(4)-Cy5-SS-Q, confirming that uncleaved extracellular molecules are not visible. Whole-body imaging of mice bearing subcutaneous tumors injected intravenously with RAFT-c(-RGDfK-)(4)-Cy5-SS-Q showed a very significant enhancement of the fluorescent contrast in tumors compared with the unquenched molecule. Histology of the tumor confirmed the intracellular accumulation of Cy5. These results demonstrate that the presence of a labile disulfide bridge between the targeting vector and a drug mimetic is an efficient way to deliver a dye, or a drug, intracellularly. In addition, this quenched RAFT-c(-RGDfK-)(4)-Cy5-SS-Q probe is a very powerful vector for imaging tumor masses and investigating in vivo RGD-mediated internalization.
authors.affiliation: INSERM U578, Institut Albert Bonniot Université Joseph Fourier, Group de recherche sur le cancer du poumon, La Tronche, France.
authors.forename: Zhao-Hui
authors.initials: ZH
authors.lastname: Jin
authors.forename: Jesus
authors.initials: J
authors.lastname: Razkin
authors.forename: Veronique
authors.initials: V
authors.lastname: Josserand
authors.forename: Didier
authors.initials: D
authors.lastname: Boturyn
authors.forename: Alexeï
authors.initials: A
authors.lastname: Grichine
authors.forename: Isabelle
authors.initials: I
authors.lastname: Texier
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Favrot
authors.forename: Pascal
authors.initials: P
authors.lastname: Dumy
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Coll
mesh_terms: D000818:Animals; D002232:Carbocyanines / Q000032:analysis / Q000378:metabolism; D045744:Cell Line, Tumor; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000032:analysis*; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence; D009369:Neoplasms / Q000737:chemistry* / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism*; D055095:Optics and Photonics; D010456:Peptides, Cyclic / Q000032:analysis* / Q000378:metabolism; D051598:Whole Body Imaging / Q000379:methods*
chemical_list: D002232:Carbocyanines; D005456:Fluorescent Dyes; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D010456:Peptides, Cyclic; C524818:RAFT-c(RGDfK)4-Cy5-SS-Q; C085321:cyanine dye 5; C047981:arginyl-glycyl-aspartic acid