The plant defensin NaD1 induces tumor cell death via a non-apoptotic, membranolytic process

Abstract

Cationic anti-microbial peptides (CAPs) have an important role in host innate defense against pathogens such as bacteria and fungi. Many CAPs including defensins also exhibit selective cytotoxic activity towards mammalian cells via both apoptotic and non-apoptotic processes, and are being investigated as potential anticancer agents. The anti-fungal plant defensin from ornamental tobacco, Nicotiana alata Defensin 1 (NaD1), was recently shown to induce necrotic-like cell death in a number of tumor cell types within 30 min of treatment, at a concentration of 10 μM. NaD1-mediated cell killing within these experimental parameters has been shown to occur via binding to the plasma membrane phosphatidylinositol 4,5-bisphosphate (PIP2) in target cells to facilitate membrane destabilization and subsequent lysis. Whether NaD1 is also capable of inducing apoptosis in tumor cells has not been reported previously. In this study, treatment of MM170 (melanoma) and Jurkat T (leukemia) cells with subacute (<10 μM) concentrations of NaD1 over 6–24 h was investigated to determine whether NaD1 could induce cell death via apoptosis. At subacute concentrations, NaD1 did not efficiently induce membrane permeabilization within 30 min, but markedly reduced cell viability over 24 h. In contrast to other CAPs that have been shown to induce apoptosis through caspase activation, dying cells were not sensitive to a pancaspase inhibitor nor did they display caspase activity or DNA fragmentation over the 24 h treatment time. Furthermore, over the 24 h period, cells exhibited necrotic phenotypes and succumbed to membrane permeabilization. These results indicate that the cytotoxic mechanism of NaD1 at subacute concentrations is membranolytic rather than apoptotic and is also likely to be mediated through a PIP2-targeting cell lytic pathway.

Moving beyond size and phosphatidylserine exposure: evidence for a diversity of apoptotic cell‐derived extracellular vesicles in vitro

“…To support this argument, large oncosomes (>1 to >10 μm in diameter) [77] were found to be distinct from ApoBDs based on the demonstration that apoptosis was not evident in culture conditions where large oncosomes were generated from human DU145 prostate cancer cells [78]. Similarly, we have also demonstrated previously that the plant defensin NaD1 can induce primary necrosis but not apoptosis [39,76,79]. However, Jurkat T cells treated with NaD1 resulted in the formation of EVs that resemble ApoBDs [39].…”

The induction and consequences of Influenza A virus-induced cell death

“…Although membrane permeabilisation during secondary necrosis has previously been thought to be an unregulated process, recent studies suggest that an N-terminal fragment generated from caspase-cleaved gasdermin E/DFNA5 may actively mediate this process 8 , 9 . In contrast, primary necrosis is directly induced by exposure to an array of stimuli such as antimicrobial peptides 10 , bacterial endotoxin 11 and heat shock 12 . Finally, similar to necrosis, necroptosis is an inflammatory form of cell death characterised by the formation of large necrotic blebs and membrane permeabilisation 13 .…”

Enediyne-activated, EGFR-targeted human β-defensin 1 has therapeutic efficacy against non-small cell lung carcinoma

“…This preference is likely due to the cationic properties of these proteins, which enable them to interact with the high percentage of anionic molecules, including phosphatidylserine and sialic acid residues, present in the cancer cell membrane [22,25,26]. Recently, defensin NaD1 was shown to induce tumor cell lysis by directly binding to PI(4,5)P2 in the plasma membrane [27]. A similar mechanism was also observed for HBD3 [8].…”