title: Identification of synthetic lethality of PRKDC in MYC-dependent human cancers by pooled shRNA screening.
abstract: BACKGROUND
  MYC family members are among the most frequently deregulated oncogenes in human cancers, yet direct therapeutic targeting of MYC in cancer has been challenging thus far. Synthetic lethality provides an opportunity for therapeutic intervention of MYC-driven cancers.
  
  
  METHODS
  A pooled kinase shRNA library screen was performed and next-generation deep sequencing efforts identified that PRKDC was synthetically lethal in cells overexpressing MYC. Genes and proteins of interest were knocked down or inhibited using RNAi technology and small molecule inhibitors, respectively. Quantitative RT-PCR using TaqMan probes examined mRNA expression levels and cell viability was assessed using CellTiter-Glo (Promega). Western blotting was performed to monitor different protein levels in the presence or absence of RNAi or compound treatment. Statistical significance of differences among data sets were determined using unpaired t test (Mann-Whitney test) or ANOVA.
  
  
  RESULTS
  Inhibition of PRKDC using RNAi (RNA interference) or small molecular inhibitors preferentially killed MYC-overexpressing human lung fibroblasts. Moreover, inducible PRKDC knockdown decreased cell viability selectively in high MYC-expressing human small cell lung cancer cell lines. At the molecular level, we found that inhibition of PRKDC downregulated MYC mRNA and protein expression in multiple cancer cell lines. In addition, we confirmed that overexpression of MYC family proteins induced DNA double-strand breaks; our results also revealed that PRKDC inhibition in these cells led to an increase in DNA damage levels.
  
  
  CONCLUSIONS
  Our data suggest that the synthetic lethality between PRKDC and MYC may in part be due to PRKDC dependent modulation of MYC expression, as well as MYC-induced DNA damage where PRKDC plays a key role in DNA damage repair.
authors.forename: Zongxiang
authors.initials: Z
authors.lastname: Zhou
authors.forename: Manishha
authors.initials: M
authors.lastname: Patel
authors.forename: Nicholas
authors.initials: N
authors.lastname: Ng
authors.forename: Mindy H
authors.initials: MH
authors.lastname: Hsieh
authors.forename: Anthony P
authors.initials: AP
authors.lastname: Orth
authors.forename: John R
authors.initials: JR
authors.lastname: Walker
authors.forename: Serge
authors.initials: S
authors.lastname: Batalov
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Harris
authors.affiliation: Genomics Institute of the Novartis Research Foundation, 10675 John Jay Hopkins Drive, San Diego, CA 92121, USA. jliu@gnf.org.
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
mesh_terms: D002460:Cell Line; D002461:Cell Line, Transformed; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics; D016000:Cluster Analysis; D053903:DNA Breaks, Double-Stranded; D051747:DNA-Activated Protein Kinase / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006801:Humans; D009369:Neoplasms / Q000235:genetics* / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D055752:Small Cell Lung Carcinoma / Q000235:genetics / Q000378:metabolism
chemical_list: D009687:Nuclear Proteins; D016271:Proto-Oncogene Proteins c-myc; D034741:RNA, Small Interfering; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human