Semisynthesis reveals apoptin as a tumour-selective protein prodrug that causes cytoskeletal collapse

Apoptin is a small viral protein capable of inducing cell death selectively in cancer cells. Despite its potential as an anticancer agent, relatively little is known about its mechanism of toxicity and cancer-selectivity. Previous experiments suggest that cancer-selective phosphorylation modulates apoptin toxicity, although a lack of chemical tools has hampered the dissection of underlying mechanisms. Here, we describe structure-function studies with site-specifically phosphorylated apoptin (apoptin-T108ph) in living cells which revealed that Thr108 phosphorylation is the selectivity switch for apoptin toxicity. Mechanistic investigations link T108ph to actin binding, cytoskeletal disruption and downstream inhibition of anoikis-resistance as well as cancer cell invasion. These results establish apoptin as a protein pro-drug, selectively activated in cancer cells by phosphorylation, which disrupts the cytoskeleton and promotes cell death. We anticipate that this mechanism provides a framework for the design of next generation anticancer proteins with enhanced selectivity and potency.

Automated summary

Apoptin is a small viral protein that selectively induces cell death in cancer cells. Phosphorylation of Thr108 is found to be the key factor that determines its toxicity. Mechanistic studies reveal that Thr108ph is associated with actin binding, cytoskeletal disruption, inhibition of anoikis-resistance, and cancer cell invasion. These findings establish apoptin as a pro-drug activated in cancer cells by phosphorylation, disrupting the cytoskeleton and promoting cell death, and provide a framework for designing next-generation anticancer proteins.