Copper-Triggered Bioorthogonal Cleavage Reactions for Reversible Protein and Cell Surface Modifications

Abstract

Temporal and reversible control over protein and cell conjugations holds great potential for traceless release of antibody–drug conjugates (ADCs) on tumor sites as well as on-demand altering or removal of targeting elements on cell surface. We herein developed a bioorthogonal and traceless releasable reaction on proteins and intact cells to fulfill such purposes. A systematic survey of transition metals in catalyzing the bioorthogonal cleavage reactions revealed that copper complexes such as Cu­(I)-BTTAA and dual-substituted propargyl (dsPra) or propargyloxycarbonyl (dsProc) moieties offered a bioorthogonal releasable pair for reversible blockage and rescue of primary amines and phenol alcohols on small molecule drugs, protein side chains, as well as intact cell surface. For proof-of-concept, we employed such Cu­(I)-BTTAA/dsProc and Cu­(I)-BTTAA/dsPra pairs as a “traceless linker” strategy to construct cleavable ADCs to unleash cytotoxic compounds on cancer cells in situ and as a “reversible modification” strategy for cell surface engineering. Furthermore, by coupling with the genetic code expansion strategy, we site-specifically modulated ligand–receptor interactions on live cell membranes. Together, our work expanded the transition-metal-mediated bioorthogonal cleavage tool kit from terminal decaging to internal-linker breakage, which offered a temporal and reversible conjugation strategy on therapeutic proteins and cells.

Unleashing the Power of Bond Cleavage Chemistry in Living Systems

“…For example, BCRs represent an efficient approach in cell engineering, such as in remodeling cell surface glycans, 127 unmasking the epitope of major histocompatibility complex class I (MHC-I), 128 , 129 and removing chemically labeled molecular probes. 41 More innovative applications in this direction will likely emerge in the future.…”

“…Our group has been interested in Cu due to the wide availability of biocompatible ligands owing to the broad applications of the CuAAC reaction in living systems. A systematic survey of the ligands and metals identified the Cu-2-[4-[[bis[(1- tert butyltriazol-4-yl)methyl]amino]methyl]triazol-1-yl] acetic acid (BTTAA)/disubstituted propargyl group as a new bioorthogonal cleavage pair, which facilitated the further extension of this chemistry from terminal to internal bond cleavage of the substituted propargyl scaffold 41 ( Figure 3 a, entries 20 and 21). In addition, a recent report by Bernardes et al utilized the Pt-triggered deprotection of pentynoyl or propargyl groups on amines for the release of MMAE and 5-FU 44 ( Figure 3 a, entry 22).…”

“…By surveying a range of commonly used transition metal catalysts, we found that Cu(I)-BTTAA was able to catalyze the cleavage of disubstituted propargyl groups to release free amine or hydroxyl groups. Such a disubstituted propargyl moiety can be further extended by the conjugation of one arm with the ZHER affibody, which converts the propargyl moiety from a terminal decaging group to a traceless internal linker for releasing bioconjugates 41 ( Figure 6 , entries 11 and 12). Both Dox and etoposide (Etop) could be efficiently released from the cleavable ADCs by Cu-BTTAA 41 ( Figure 6 , entries 11 and 12).…”

Transition Metal‐Promoted Reactions in Aqueous Media and Biological Settings

“…The bioorthogonal removal of O ‐propargyl and N ‐propargyloxycarbonyl groups can also be carried out with copper complexes. In particular, Chen and co‐workers have recently shown that the complex Cu I ‐BTTA, originally developed for azide–alkyne biocompatible cycloadditions, is also able to promote the bioorthogonal cleavage of O ‐propargyl and N ‐Poc groups that hold a second alkyl substituent at the propargylic position [131a] . The method constitutes the first examples of a copper‐catalyzed biorthogonal cleavage reaction and was applied to the release of drugs bearing a phenol or a primary amine moiety, such as the anticancer agent doxorubicin (Figure 51 A).…”

Prodrug Activation by Gold Artificial Metalloenzyme‐Catalyzed Synthesis of Phenanthridinium Derivatives via Hydroamination

“…By incorporating metals into nanoparticles or using homogeneous metal complexes to overcome the inherent cytotoxicity of the abiotic catalyst, several groups have demonstrated methods for metal‐based uncaging of drugs applicable in living systems. Employing metals like iron, palladium, copper, ruthenium, platinum and gold, several studies focused on release of amine/hydroxyl groups through protecting groups such as phenyl azide, [10] allyl, [11] propargyl, [12] allyloxycarbonyl, [13] dual‐substituted propargyloxycarbonyl, [14] 2‐alkynlbenzamide, [15] as well as a strategy based on ring‐closing metathesis [16] …”