Electrophile-Integrating Smiles Rearrangement Provides Previously Inaccessible C4′‑<i>O</i>‑Alkyl Heptamethine Cyanine Fluorophores

New synthetic methods to rapidly access useful fluorophores are needed to advance modern molecular imaging techniques. A new variant of the classical Smiles rearrangement is reported that enables the efficient synthesis of previously inaccessible C4′-<i>O-</i>alkyl heptamethine cyanines. The key reaction involves <i>N</i>- to <i>O</i>- transposition with selective electrophile incorporation on nitrogen. A representative fluorophore exhibits excellent resistance to thiol nucleophiles, undergoes productive bioconjugation, and can be used in near-IR fluorescence imaging applications

Synthesis of Hindered Anilines: Three-Component Coupling of Arylboronic Acids, <i>tert</i>-Butyl Nitrite, and Alkyl Bromides

The synthesis of sterically hindered amines has been a significant challenge in organic chemistry. Herein, we report a modular, three-component coupling that constructs two carbon–nitrogen bonds including a sterically hindered C_sp³–N bond using commercially available materials. This process uses an earth-abundant copper catalyst and mild reaction conditions, allowing access to a variety of complex aromatic amines

Impact of C4′‑<i>O</i>‑Alkyl Linker on <i>in Vivo</i> Pharmacokinetics of Near-Infrared Cyanine/Monoclonal Antibody Conjugates

Near-infrared (NIR) fluorophores have several advantages over visible-light fluorophores, including superior tissue penetration and lower autofluorescence. We recently accessed a new class of readily synthesized NIR cyanines containing a novel C4′-<i>O</i>-alkyl linker, which provides both high chemical stability and excellent optical properties. In this study, we provide the first <i>in vivo</i> analysis of this new class of compounds, represented by the tetrasulfonate FNIR-774 (Frederick NIR 774). Monoclonal antibody (mAb) conjugates of FNIR-774 were compared to conjugates of the commercially available dye (IRDye800CW (IR800)), one of the most widely used NIR fluorophores for clinical translation. Both dyes were conjugated to panitumumab (pan) or cetuximab (cet) with ratios of 1:2 or 1:5. Conjugates of both dyes demonstrated similar quenching capacity, stability, and brightness in target cells <i>in vitro</i>. In contrast, <i>in vivo</i> imaging in mice showed different pharmacokinetics between pan-FNIR-774 (1:5) and pan-IR800 (1:5), or cet-FNIR-774 (1:5) and cet-IR800 (1:5). Particularly at the higher labeling density, mAb-FNIR-774 conjugates showed superior specific accumulation in tumors compared with mAb-IR800 conjugates. Thus, FNIR-774 conjugates showed superior <i>in vivo</i> pharmacokinetics compared with IR800 conjugates, independent of the mAb. These results suggest that FNIR-774 is a promising fluorescent probe for NIR optical imaging