Time-Lapse Imaging of Neuroblastoma Cells to Determine Cell Fate upon Gene Knockdown

Neuroblastomais the most common extra-cranial solid tumor of early childhood. Standard therapies are not effective in case of poor prognosis and chemotherapy resistance. To improve drug therapy, it is imperative to discover new targets that play a substantial role in tumorigenesis of neuroblastoma. The mitotic machinery is an attractive target for therapeutic interventions and inhibitors can be developed to target mitotic entry, spindle apparatus, spindle activation checkpoint, and mitotic exit. We present an elaborate analysis pipeline to determine cancer specific therapeutic targets by first performing a focused gene expression analysis to select genes followed by a gene knockdownscreening assay of live cells. We interrogated gene expression studies of neuroblastomatumors and selected 240 genes relevant for tumorigenesis and cell cycle. With these genes we performed time-lapse screening of gene knockdownsin neuroblastomacells. We classified cellular phenotypes and used the temporal context of the perturbation effect to determine the sequence of events, particularly the mitotic entry preceding cell death. Based upon this phenotype kinetics from the gene knockdownscreening, we inferred dynamic gene functions in mitosisand cell proliferation. We identified six genes ( DLGAP5, DSCC1, SMO, SNRPD1, SSBP1, and UBE2C) with a vital role in mitosisand these are promising therapeutic targets for neuroblastoma. Images and movies of every time point of all screened genes are available at https://ichip.bioquant.uni-heidelberg.de.