Increasing melanoma cell death using an inhibitor of protein disulphide isomerases to abrogate stress-survival responses


Year:

Session type:

Penny Lovat1, Marco Corazzari2, Jane Armstrong1, Shaun Martin1, Vittoria Pagliarini2, David Hill1, Anna Brown1, Mauro Piacentini2, Mark Birch-Machin1, Chris Redfern1

1Newcastle University, Newcastle upon Tyne, UK, 2L’Spallanzani National Institute for Infectious Disease, Rome, Italy

Abstract

Increasing melanoma cell death using an inhibitor of protein disulphide isomerases to abrogate stress-survival responses

Background

Exploiting vulnerabilities in intracellular signalling pathways of tumor cells is a key strategy for drug development. The activation of cellular stress responses mediated by the endoplasmic reticulum (ER) allows cancer cells to survive outside their normal environment. Many proteins that protect cells against ER stress are active as protein disulfide isomerases (PDI) and the aim of this study was to test the hypothesis that apoptosis in response to ER stress can be increased by inhibiting PDI activity.

Method

The response of melanoma cell lines to fenretinide, velcade or PDI inhibitor was evaluated using biochemical markers of ER stress and assays for viability and apoptosis. The role of PDI inhibition was assessed by transfection with a wild-type and mutant PDI.

Results

The novel chemotherapeutic drugs fenretinide and velcade induced ER stress-mediated apoptosis in melanoma cells. Both the stress response and apoptosis were enhanced by bacitracin, a macrocyclic dodecapeptide antibiotic which inhibits PDI activity. Over-expression of the main cellular PDI, procollagen-proline, 2-oxoglutarate-4-dioxygenase-beta subunit (P4HB), resulted in increased PDI activity and abrogated the apoptosis-enhancing effect of bacitracin. In contrast, over-expression of a mutant P4HB lacking PDI activity did not increase cellular PDI activity or block the effects of bacitracin.

Conclusion

These results provide proof-of-principle for the concept that ER-stress-induced apoptosis can be enhanced using PDI inhibitors. Furthermore, the data imply that small-molecule PDI inhibitors designed to bind to the CXXC motif of the PDI active site may have significant potential for enhancing the efficacy of chemotherapy in a wide range of cancers.