Triple, Mutually Orthogonal Bioorthogonal Pairs through the Design of Electronically Activated Sulfamate-Containing Cycloalkynes

Interest
in mutually exclusive pairs of bioorthogonal labeling
reagents continues to drive the design of new compounds that are capable
of fast and predictable reactions. The ability to easily modify S-,
N-, and O-containing cyclooctynes (SNO-OCTs) enables electronic tuning
of various SNO-OCTs to influence their cycloaddition rates with Type
I–III dipoles. As opposed to optimizations based on just one
specific dipole class, the electrophilicity of the alkynes in SNO-OCTs
can be manipulated to achieve divergent reactivities and furnish mutually
orthogonal dual ligation systems. Significant reaction rate enhancements
of a difluorinated SNO-OCT derivative, as compared to the parent scaffold,
were noted, with the second-order rate constant in cycloadditions
with diazoacetamides exceeding 5.13 M–1 s–1. Computational and experimental studies were employed to inform
the design of triple ligation systems that encompass three orthogonal
reactivities. Finally, polar SNO-OCTs are rapidly internalized by
mammalian cells and remain functional in the cytosol for live-cell
labeling, highlighting their potential for diverse in vitro and in
vivo applications.