NF-κB p65 repression by the sesquiterpene lactone, Helenalin, contributes to the induction of autophagy cell death.
BACKGROUND
Numerous studies have demonstrated that autophagy plays a vital role in maintaining cellular homeostasis. Interestingly, several anticancer agents were found to exert their anticancer effects by triggering autophagy. Emerging data suggest that autophagy represents a novel mechanism that can be exploited for therapeutic benefit. Pharmacologically active natural compounds such as those from marine, terrestrial plants and animals represent a promising resource for novel anticancer drugs. There are several prominent examples from the past proving the success of natural products and derivatives exhibiting anticancer activity. Helenalin, a sesquiterpene lactone has been demonstrated to have potent anti-inflammatory and antitumor activity. Albeit previous studies demonstrating helenalin's multi modal action on cellular proliferative and apoptosis, the mechanisms underlying its action are largely unexplained.


METHODS
To deduce the mechanistic action of helenalin, cancer cells were treated with the drug at various concentrations and time intervals. Using western blot, FACS analysis, overexpression and knockdown studies, cellular signaling pathways were interrogated focusing on apoptosis and autophagy markers.


RESULTS
We show here that helenalin induces sub-G1 arrest, apoptosis, caspase cleavage and increases the levels of the autophagic markers. Suppression of caspase cleavage by the pan caspase inhibitor, Z-VAD-fmk, suppressed induction of LC3-B and Atg12 and reduced autophagic cell death, indicating caspase activity was essential for autophagic cell death induced by helenalin. Additionally, helenalin suppressed NF-κB p65 expression in a dose and time dependent manner. Exogenous overexpression of p65 was accompanied by reduced levels of cell death whereas siRNA mediated suppression led to augmented levels of caspase cleavage, autophagic cell death markers and increased cell death.


CONCLUSIONS
Taken together, these results show that helenalin mediated autophagic cell death entails inhibition of NF-κB p65, thus providing a promising approach for the treatment of cancers with aberrant activation of the NF-κB pathway.
Division of Chemical Biology and Biotechnology, School of Biological Sciences, College of Science, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551, Singapore.
Chuan Bian
CB
Lim
Pan You
PY
Fu
Nung
N
Ky
Hong Shuang
HS
Zhu
XiaoLing
X
Feng
Jinming
J
Li
Kandhadayar Gopalan
KG
Srinivasan
Mohamed Sabry
MS
Hamza
Yan
Y
Zhao
D000590:Amino Acid Chloromethyl Ketones / Q000378:metabolism; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D018650:Arnica / Q000737:chemistry*; D001343:Autophagy / Q000187:drug effects*; D000071192:Autophagy-Related Protein 12; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D016193:G1 Phase / Q000187:drug effects; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009369:Neoplasms / Q000188:drug therapy / Q000378:metabolism*; D008517:Phytotherapy; D010936:Plant Extracts / Q000494:pharmacology / Q000627:therapeutic use; D034741:RNA, Small Interfering / Q000378:metabolism; D055630:Receptors, NK Cell Lectin-Like / Q000037:antagonists & inhibitors*; D012717:Sesquiterpenes / Q000494:pharmacology* / Q000627:therapeutic use; D045789:Sesquiterpenes, Guaiane; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism
C466624:ATG12 protein, human; D000590:Amino Acid Chloromethyl Ketones; D000972:Antineoplastic Agents, Phytogenic; D000071192:Autophagy-Related Protein 12; C548774:KLRF2 protein, human; C502070:MAP1LC3B protein, human; D008869:Microtubule-Associated Proteins; D010936:Plant Extracts; D034741:RNA, Small Interfering; D055630:Receptors, NK Cell Lectin-Like; D012717:Sesquiterpenes; D045789:Sesquiterpenes, Guaiane; D025841:Small Ubiquitin-Related Modifier Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C001329:helenalin; D020169:Caspases