Ortho-phthalaldehyde (OPA) based bifunctional probes and chemoselective peptide cyclization and post-cyclization modification

Abstract

Chemical reactions in the native proteins under physiological conditions offer great opportunities to modify proteins and introduce exogenous moieties onto native proteins to generate bioconjugates. Thus, the development of a new chemoselective bioconjugation reaction is in great demand. In this thesis, the OPA/amine reaction and OPA/amine/thiol reaction have been developed to modify native proteins and construct cyclic peptides under the physiological conditions effectively. Firstly, OPA and its derivatives can rapidly and smoothly react with primary amine moieties in both peptides and proteins to achieve native protein bioconjugation. Furthermore, OPA-alkyne bifunctional linkers can further be functionalized with fluorophore via Click reaction, and the bifunctional linkers also can be used for proteome profiling and microarray technologies. Because the antibody-drug conjugates (ADCs) offer a very promising strategy for selective delivery of the cytotoxic drug to malignant tumor cells, various of OPA-based bifunctional linkers have been developed for drug conjugation via a maleimide-thiol reaction, followed by monoclonal antibody (mAb) linkage via OPA/amine bioconjugation reaction. Secondly, OPA/amine/thiol three-component reaction has been developed for chemoselective peptide cyclization, directly on unprotected peptides in the aqueous conditions. Moreover, this OPA-based cyclic peptide can be further modified with the N-maleimide moiety in a one-pot manner to introduce additional functionalities. Likewise, the thiophene-2,3-dicarboxaldehyde (TDA) can also be used to construct highly stable cyclic peptides in the same manner. Therefore, the development of OPA/TDA based reactions indeed extend the toolbox for protein chemical modification and construction of cyclic peptides.