Development of a solid-phase protein labelling method for the synthesis of peptide-drug conjugates

Alanazi, Manal (2020). Development of a solid-phase protein labelling method for the synthesis of peptide-drug conjugates. University of Birmingham. Ph.D.

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Abstract

Protein-drug conjugates are emerging as a powerful tool for the targeting of specific cell populations, as demonstrated by the emergence of approved antibody-drug conjugates. In common with the broader field of protein modification, the selective synthesis of these agents represents a significant chemical challenge, as the conjugation reaction must modify the targeted residue in the presence of other competing unprotected amino acid side chains. While the alkylation of cysteine thiols remains the most commonly utilized method for the selective modification of proteins, there remain significant issues with this approach in terms of controlling cysteine oxidation, separation of any unreacted protein, and reagent compatibility.

In order to address these issues, we have developed a novel system containing a thiol reactive group (iodoacetamide) attached to the standard solid support (agarose) via a linker that can be readily cleaved under mild conditions. Following initial studies an allyl carbamate linker was selected due to its general stability and relatively straightforward deprotection with optimised catalytic Pd systems. Based on this we have developed a synthesis of the completed catch-release linker system in 7 steps and demonstrated that this linker can be immobilised on agarose and cleaved on the solid phase.

This system was then combined with a second solid phase reducing agent to allow the direct reduction and alkylation/trapping of a standard protein (BSA). The protein was successfully alkylated and separated from the unreacted protein by simple filtration. Unfortunately, the release of alkylated BSA from the solid support by using catalytic Pd did not proceed as expected from the model system and this step is currently under investigation.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):