Development of targeted therapies for melanoma – synthetic and analytical studies of duocarmycin-protein conjugates

Peptide-Drug Conjugates (PDCs) and Antibody-Drug Conjugates (ADCs) are compounds that aim to specifically deliver a high concentration of the cytotoxic agent to the tumour target . This project was designed to develop new PDCs based upon the potent cytotoxic agent CBI, to target melanoma. Chapter 1 provides an overall introduction to melanoma with particular emphasis on current treatments, the involvement of the melanocortin 1 receptor (MC1R) in cancer progression and how MC1R can be used as a potential target. Chapter 2 describes the synthesis of CBI analogues, ultrapotent cytotoxic agents that belong in the same family as CC-1065 and yatakeycin. CBI binds in the AT rich minor groove of the DNA causing their cyclopropyl group ring to open followed by adenine alkylation and then triggering a cascade of events that subsequently lead to cell apoptosis. These class of compounds are considered more effective as they can induce activity at any stage of the cell cycle. Herein, we describe the synthesis of a CBI analogue that can potentially be used in solid phase synthesis. The target compounds, which consist of an Fmoc-protected glutamic acid analogue carrying a CBI alkylating subunit on the side chain, were successfully synthesised and characterised. Off target toxicity is a crucial factor to consider when developing PDCs and ADCs with ultrapotent payloads. Therefore, Chapter 3 explores the solid phase synthesis of melanocyte stimulating hormone (MSH) as the targeting peptide to MC1R and its conjugation with the CBI analogues. The successful synthesis of several peptide conjugates was confirmed by mass spectrometry and represents the first description of this class of PDC. Preliminary studies of the biological activity of the CBI-MSH conjugates are described in chapter 4. The activity of the conjugates was compared with scrambled and protected versions of the active compounds and, while the activity was lower than expected, demonstrated an anti-proliferative effect in various assays. Flow cytometry also gave a preliminary indication that the peptide was binding to its target