Coibamide A Induces mTOR-Independent Autophagy and Cell Death in Human Glioblastoma Cells

Andrew M Hau,Christiane V Löhr,Ian G Ganley,Jeffrey D Serrill,Philip J Proteau,Jane E Ishmael,Jeffrey A Greenwood,Kerry L Mcphail,Ben C.B Ko

doi:10.1371/journal.pone.0065250

Abstract

Coibamide A is an N-methyl-stabilized depsipeptide that was isolated from a marine cyanobacterium as part of an International Cooperative Biodiversity Groups (ICBG) program based in Panama. Previous testing of coibamide A in the NCI in vitro 60 cancer cell line panel revealed a potent anti-proliferative response and “COMPARE-negative” profile indicative of a unique mechanism of action. We report that coibamide A is a more potent and efficacious cytotoxin than was previously appreciated, inducing concentration- and time-dependent cytotoxicity (EC50<100 nM) in human U87-MG and SF-295 glioblastoma cells and mouse embryonic fibroblasts (MEFs). This activity was lost upon linearization of the molecule, highlighting the importance of the cyclized structure for both anti-proliferative and cytotoxic responses. We show that coibamide A induces autophagosome accumulation in human glioblastoma cell types and MEFs via an mTOR-independent mechanism; no change was observed in the phosphorylation state of ULK1 (Ser-757), p70 S6K1 (Thr-389), S6 ribosomal protein (Ser-235/236) and 4EBP-1 (Thr-37/46). Coibamide A also induces morphologically and biochemically distinct forms of cell death according to cell type. SF-295 glioblastoma cells showed caspase-3 activation and evidence of apoptotic cell death in a pattern that was also seen in wild-type and autophagy-deficient (ATG5-null) MEFs. In contrast, cell death in U87-MG glioblastoma cells was characterized by extensive cytoplasmic vacuolization and lacked clear apoptotic features. Cell death was attenuated, but still triggered, in Apaf-1-null MEFs lacking a functional mitochondria-mediated apoptotic pathway. From the study of ATG5-null MEFs we conclude that a conventional autophagy response is not required for coibamide A-induced cell death, but likely occurs in dying cells in response to treatment. Coibamide A represents a natural product scaffold with potential for the study of mTOR-independent signaling and cell death mechanisms in apoptotic-resistant cancer cells.

Highlights

There is high demand for new small molecules that can strategically target the dysregulated signaling pathways that underlie aggressive solid cancers such as glioblastoma
In the present study we investigated the cytotoxic potential of coibamide A against human glioblastoma cells and immortalized mouse embryonic fibroblasts (MEFs) from genetically manipulated animals
Using several independent measures of general cytotoxicity we found that coibamide A dramatically reduced the viability of U87-MG and SF-295 glioma cells in both a concentration- and time-dependent manner

Summary

Introduction

There is high demand for new small molecules that can strategically target the dysregulated signaling pathways that underlie aggressive solid cancers such as glioblastoma. Cell signaling driven by growth factors, such as the mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways, is dramatically enhanced. Together these aberrant signaling networks tend to promote cell survival and lend GBM a natural resistance to apoptosis, rendering conventional chemotherapeutic drugs that typically induce apoptosis ineffective for the treatment of this condition [3]. There is a great need for new pharmacologic tools that cause cell death in glioblastoma and other apoptosis-resistant cancer cells

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