Nimbolide, a neem limonoid inhibits cytoprotective autophagy to activate apoptosis via modulation of the PI3K/Akt/GSK-3β signalling pathway in oral cancer

Cell Death Dis. 2018 Oct 23;9(11):1087. doi: 10.1038/s41419-018-1126-4.

Abstract

Of late, nimbolide, a limonoid from the neem tree (Azadirachta indica) has gained increasing research attention owing to its potent antiproliferative and apoptosis-inducing effects. The present study was designed to investigate the effect of nimbolide on autophagy and the time point at which the phosphorylation status of GSK-3β and PI3K dictate the choice between autophagy and apoptosis in SCC131 and SCC4 oral cancer cells. Additionally, we analysed changes in the expression of proteins involved in autophagy and apoptosis after therapeutic intervention with nimbolide in a hamster model of oral oncogenesis. Furthermore, we also demonstrate changes in the expression of key genes involved in apoptosis and autophagy during the stepwise evolution of hamster and human OSCCs. Nimbolide-induced stereotypical changes in oral cancer cells characteristic of both apoptosis and autophagy. Time-course experiments revealed that nimbolide induces autophagy as an early event and then switches over to apoptosis. Nimbolide negatively regulates PI3K/Akt signalling with consequent increase in p-GSK-3βTyr216, the active form of GSK-3β that inhibits autophagy. Downregulation of HOTAIR, a competing endogenous RNA that sponges miR-126 may be a major contributor to the inactivation of PI3K/Akt/GSK3 signalling by nimbolide. Analysis of key markers of apoptosis and autophagy as well as p-AktSer473 during sequential progression of hamster and human OSCC revealed a gradual evolution to a pro-autophagic and antiapoptotic phenotype that could confer a survival advantage to tumors. In summary, the results of the present study provide insights into the molecular mechanisms by which nimbolide augments apoptosis by overcoming the shielding effects of cytoprotective autophagy through modulation of the phosphorylation status of Akt and GSK-3β as well as the ncRNAs miR-126 and HOTAIR. Development of phytochemicals such as nimbolide that target the complex interaction between proteins and ncRNAs that regulate the autophagy/apoptosis flux is of paramount importance in cancer prevention and therapeutics.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Anthracenes / pharmacology
  • Apoptosis / drug effects*
  • Autophagy / drug effects*
  • Azadirachta / chemistry
  • Carcinoma, Squamous Cell / chemically induced
  • Carcinoma, Squamous Cell / drug therapy
  • Carcinoma, Squamous Cell / metabolism*
  • Carcinoma, Squamous Cell / pathology
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Cricetinae
  • Disease Models, Animal
  • Glycogen Synthase Kinase 3 beta / metabolism*
  • Humans
  • Limonins / pharmacology*
  • Limonins / therapeutic use
  • Male
  • Mesocricetus
  • MicroRNAs / metabolism
  • Mouth Neoplasms / chemically induced
  • Mouth Neoplasms / drug therapy
  • Mouth Neoplasms / metabolism*
  • Mouth Neoplasms / pathology
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Phosphorylation / drug effects
  • Piperidines / pharmacology
  • Plant Extracts / pharmacology*
  • Plant Extracts / therapeutic use
  • Proto-Oncogene Proteins c-akt / metabolism*
  • RNA, Long Noncoding / antagonists & inhibitors
  • RNA, Long Noncoding / metabolism
  • Signal Transduction / drug effects

Substances

  • 3-(2,6-bis(4-fluorophenyl)-3-methylpiperidin-4-ylideneamino)-2-thioxoimidazolidin-4-one on 7,12-dimethylbenz(a)anthracene
  • Anthracenes
  • HOTAIR long untranslated RNA, human
  • Limonins
  • MIRN126 microRNA, human
  • MicroRNAs
  • Piperidines
  • Plant Extracts
  • RNA, Long Noncoding
  • nimbolide
  • Glycogen Synthase Kinase 3 beta
  • Proto-Oncogene Proteins c-akt