title: Identification of Mithramycin Analogues with Improved Targeting of the EWS-FLI1 Transcription Factor.
abstract: OBJECTIVE
  The goal of this study was to identify second-generation mithramycin analogues that better target the EWS-FLI1 transcription factor for Ewing sarcoma. We previously established mithramycin as an EWS-FLI1 inhibitor, but the compound's toxicity prevented its use at effective concentrations in patients.
  
  
  METHODS
  We screened a panel of mithralogs to establish their ability to inhibit EWS-FLI1 in Ewing sarcoma. We compared the IC50 with the MTD established in mice to determine the relationship between efficacy and toxicity. We confirmed the suppression of EWS-FLI1 at the promoter, mRNA, gene signature, and protein levels. We established an improved therapeutic window by using time-lapse microscopy to model the effects on cellular proliferation in Ewing sarcoma cells relative to HepG2 control cells. Finally, we established an improved therapeutic window using a xenograft model of Ewing sarcoma.
  
  
  RESULTS
  EC-8105 was found to be the most potent analogue and was able to suppress EWS-FLI1 activity at concentrations nontoxic to other cell types. EC-8042 was substantially less toxic than mithramycin in multiple species but maintained suppression of EWS-FLI1 at similar concentrations. Both compounds markedly suppressed Ewing sarcoma xenograft growth and inhibited EWS-FLI1 in vivo
  
  
  CONCLUSIONS
  These results provide a basis for the continued development of EC-8042 and EC-8105 as EWS-FLI1 inhibitors for the clinic. Clin Cancer Res; 22(16); 4105-18. ©2016 AACR.
authors.affiliation: Division of Pediatric Hematology/Oncology, Vanderbilt University School of Medicine, Nashville, Tennessee.
authors.forename: Christy L
authors.initials: CL
authors.lastname: Osgood
authors.affiliation: Division of Pediatric Hematology/Oncology, Vanderbilt University School of Medicine, Nashville, Tennessee.
authors.forename: Nichole
authors.initials: N
authors.lastname: Maloney
authors.affiliation: Division of Pediatric Hematology/Oncology, Vanderbilt University School of Medicine, Nashville, Tennessee.
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Kidd
authors.affiliation: Van Andel Research Institute, Grand Rapids, Michigan.
authors.forename: Susan
authors.initials: S
authors.lastname: Kitchen-Goosen
authors.affiliation: Division of Pediatric Hematology/Oncology, Vanderbilt University School of Medicine, Nashville, Tennessee. Pediatric Oncology Branch, National Cancer Institute, Bethesda, Maryland.
authors.forename: Laura
authors.initials: L
authors.lastname: Segars
authors.affiliation: Pediatric Oncology Branch, National Cancer Institute, Bethesda, Maryland.
authors.forename: Meti
authors.initials: M
authors.lastname: Gebregiorgis
authors.affiliation: Basic Science Program, Leidos Biomedical Research Laboratory, Inc., Molecular Targets Laboratory, Frederick National Laboratory for Cancer Research, Frederick, Maryland.
authors.forename: Girma M
authors.initials: GM
authors.lastname: Woldemichael
authors.affiliation: Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Min
authors.initials: M
authors.lastname: He
authors.affiliation: Department of Developmental Biology, Washington University School of Medicine, St. Louis, Missouri.
authors.forename: Savita
authors.initials: S
authors.lastname: Sankar
authors.affiliation: Center for Childhood Cancer and Blood Disorders, Nationwide Children's Hospital, Division of Pediatric Hematology/Oncology/BMT, The Ohio State University, Columbus, Ohio.
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Lessnick
authors.affiliation: Texas Tech University Health Science Center, School of Medicine, Lubbock, Texas.
authors.forename: Min
authors.initials: M
authors.lastname: Kang
authors.affiliation: Pediatric Oncology Branch, National Cancer Institute, Bethesda, Maryland. Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Malcolm
authors.initials: M
authors.lastname: Smith
authors.affiliation: Van Andel Research Institute, Grand Rapids, Michigan.
authors.forename: Lisa
authors.initials: L
authors.lastname: Turner
authors.affiliation: Van Andel Research Institute, Grand Rapids, Michigan.
authors.forename: Zachary B
authors.initials: ZB
authors.lastname: Madaj
authors.affiliation: Van Andel Research Institute, Grand Rapids, Michigan.
authors.forename: Mary E
authors.initials: ME
authors.lastname: Winn
authors.affiliation: EntreChem S.L., Oviedo, Spain.
authors.forename: Luz-Elena
authors.initials: LE
authors.lastname: Núñez
authors.affiliation: EntreChem S.L., Oviedo, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: González-Sabín
authors.affiliation: Pediatric Oncology Branch, National Cancer Institute, Bethesda, Maryland.
authors.forename: Lee J
authors.initials: LJ
authors.lastname: Helman
authors.affiliation: EntreChem S.L., Oviedo, Spain.
authors.forename: Francisco
authors.initials: F
authors.lastname: Morís
authors.affiliation: Division of Pediatric Hematology/Oncology, Vanderbilt University School of Medicine, Nashville, Tennessee. Van Andel Research Institute, Grand Rapids, Michigan. Helen De Vos Children's Hospital, Grand Rapids, Michigan. Department of Pediatrics, Michigan State University School of Medicine, East Lansing, Michigan. patrick.grohar@vai.org.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Grohar
mesh_terms: D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051379:Mice; D058990:Molecular Targeted Therapy; D015514:Oncogene Proteins, Fusion / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D008926:Plicamycin / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D051789:Proto-Oncogene Protein c-fli-1 / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D034802:RNA-Binding Protein EWS / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D012512:Sarcoma, Ewing / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism / Q000401:mortality; D014157:Transcription Factors; D023041:Xenograft Model Antitumor Assays
chemical_list: D000903:Antibiotics, Antineoplastic; C106031:EWS-FLI fusion protein; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D014157:Transcription Factors; D008926:Plicamycin