Macrocyclization of a potent PACE4 inhibitor: Benefits and limitations.

PACE4, one of the seven members of the proprotein convertase family, plays an important role in the progression of prostate cancer. Therefore, its inhibition has become an attractive target to develop new therapies against this disease. Recently, we have developed a highly potent and selective PACE4 inhibitor, known as the multi-Leu peptide with the following sequence Ac-LLLLRVKR-NH. Herein, with the aim of improving the stability profile of this inhibitor for potential in vivo application, we investigated the impact of different cyclization strategies. The inhibitory activity of new peptides was tested and compared to their linear counterparts. The potent analogues were further selected for stability evaluation. Our results showed that the cyclization involving a C-terminal carboxylic acid (head-to-tail or side chain-to-tail) led to compounds with significantly diminished inhibitory potency towards PACE4, indicating that an appropriate balance between rigidity and flexibility of the structure is necessary to allow the optimal binding with the enzyme. On the other hand, the modification within a multi-Leu core in combination with the incorporation of a C-terminal 4-amidinobenzylamide (Amba) residue yielded potent cyclic analogues. The best compound derived from this group, (&)[Mpa]LLLC(&)RVK[Amba] (where & indicates cyclization, Mpa - 3-mercaptopropionic acid), exhibited promising overall profile comprising of potent inhibitory effect against PACE4 and prostate cancer cell lines as well as improved stability. We believe that this cyclic framework could be further used to design even more potent and stable PACE4 inhibitors.