Genome-wide RNA interference screen in cancer stem cell

Guillaume Pinna 1, Marie Vandamme 1, Celia Rouault 2, Emmanuelle Charafe-Jauffret 2, Christophe Ginestier 3

  • 1Plateforme ARN interférence (PARi), Université Paris Cité & Université Paris-Saclay, Inserm, iRCM/IBFJ CEA, Stabilité Génétique Cellules Souches et Radiations, Fontenay-aux-Roses, France.
  • 2CRCM, Inserm, CNRS, Institut Paoli-Calmettes, Aix-Marseille Université, Epithelial Stem Cells and Cancer Lab, Equipe labellisée LIGUE contre le cancer, Marseille, France.
  • 3CRCM, Inserm, CNRS, Institut Paoli-Calmettes, Aix-Marseille Université, Epithelial Stem Cells and Cancer Lab, Equipe labellisée LIGUE contre le cancer, Marseille, France. Electronic address: christophe.ginestier@inserm.fr.

Tumor heterogeneity represents a major hurdle for therapy. This cellular heterogeneity is mainly sustained by different subpopulations of tumorigenic cells, the so-called cancer stem cells (CSCs). CSCs burden is associated with disease progression and patient poor prognosis. In this context, deciphering molecular mechanisms regulating stemness is a key step in the development of new therapeutic strategy. Here, we provide a detailed protocol for high-throughput screening (HTS) strategy to detect modulators of CSC proportion. It is based on a miniaturized ALDEFLUOR-probed CSC assay quantitated by high-content imaging, that allows monitoring the changes in CSC proportions in response to gene silencing. Gene loss-of-function is achieved by transfecting a genome-wide RNA interference library. These genome-wide HTS strategies could lead to the identification of new therapeutic approaches in the treatment of various cancers.