title: Phenotypic drug screening and target validation for improved personalized therapy reveal the complexity of phenotype-genotype correlations in clear cell renal cell carcinoma.
abstract: OBJECTIVE
  Novel personalized therapeutic approaches are urgently needed for patients with metastatic clear cell renal cell carcinoma (ccRCC).
  
  
  METHODS
  We combined the development of a primary patient-derived ccRCC cell line with a phenotypic drug screen consisting of 101 approved anticancer compounds.
  
  
  RESULTS
  We identified the MNNG HOS transforming gene (MET)-anaplastic lymphoma receptor tyrosine kinase (ALK) inhibitor crizotinib as the top hit of our drug screen, whereas compounds targeting the vascular endothelial growth factor (VEGF) or mammalian target of rapamycin (mTOR) pathway showed no or only minor in vitro activity. Among the known major crizotinib targets MET, ALK, and ROS-1, only MET was expressed in our ccRCC cell line. Subsequent sequence analysis revealed a heterozygous R988C mutation of the MET gene and a VHL deletion in both the primary tumor and the tumor-derived ccRCC cell line. However, we were unable to show an activation of MET and, further, MET knockdown did not result in increased apoptosis or cytotoxicity. Therefore, our results suggest that MET R988C does not function as a major oncogenic driver mutation but rather represents a sequence variant. However, we provide evidence that the cytotoxic effect of crizotinib in our cell line model correlates with its ability to inhibit P-glycoprotein (ABCB1)-associated transport functions.
  
  
  CONCLUSIONS
  Our study shows that a phenotypic screen of a patient-derived tumor cell line can identify compounds with antitumor activity but with an unexpected mode of action. Our results underscore that target validation and phenotype-genotype correlations remain a major experimental challenge. The implications of our findings for a personalized management of patients with cancer are discussed.
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Molecular Urooncology, Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Meike
authors.initials: M
authors.lastname: Schneider
authors.affiliation: Oncotest GmbH, Freiburg, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Schüler
authors.affiliation: Molecular Urooncology, Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Rouven
authors.initials: R
authors.lastname: Höfflin
authors.affiliation: Molecular Urooncology, Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Korzeniewski
authors.affiliation: Department of Medical Oncology, National Center for Tumor Diseases, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Carsten
authors.initials: C
authors.lastname: Grüllich
authors.affiliation: Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany; Department of Pathology, University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Wilfried
authors.initials: W
authors.lastname: Roth
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Dogu
authors.initials: D
authors.lastname: Teber
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Boris
authors.initials: B
authors.lastname: Hadaschik
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Sascha
authors.initials: S
authors.lastname: Pahernik
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Hohenfellner
authors.affiliation: Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Molecular Urooncology, Department of Urology, University of Heidelberg School of Medicine, Heidelberg, Germany; Center for Kidney Tumors, National Center for Tumor Disease and University of Heidelberg School of Medicine, Heidelberg, Germany. Electronic address: stefan.duensing@med.uni-heidelberg.de.
authors.forename: Stefan
authors.initials: S
authors.lastname: Duensing
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D002292:Carcinoma, Renal Cell / Q000235:genetics / Q000473:pathology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D000077547:Crizotinib; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020022:Genetic Predisposition to Disease / Q000235:genetics*; D005838:Genotype; D006801:Humans; D015151:Immunoblotting; D007680:Kidney Neoplasms / Q000235:genetics / Q000473:pathology; D058990:Molecular Targeted Therapy / Q000379:methods*; D009154:Mutation; D010641:Phenotype; D057285:Precision Medicine / Q000379:methods*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019859:Proto-Oncogene Proteins c-met / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D034622:RNA Interference; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D011720:Pyrazoles; D011725:Pyridines; D000077547:Crizotinib; C491743:MET protein, human; D019859:Proto-Oncogene Proteins c-met
keywords: Crizotinib; Drug screen; MET; Personalized therapy; Renal cancer