title: NOTCH1 and SOX10 are Essential for Proliferation and Radiation Resistance of Cancer Stem-Like Cells in Adenoid Cystic Carcinoma.
abstract: OBJECTIVE
  Although the existence of cancer stem cells (CSC) in adenoid cystic carcinoma (ACC) has been proposed, lack of assays for their propagation and uncertainty about molecular markers prevented their characterization. Our objective was to isolate CSC from ACC and provide insight into signaling pathways that support their propagation.
  
  
  METHODS
  To isolate CSC from ACC and characterize them, we used ROCK inhibitor-supplemented cell culture, immunomagnetic cell sorting, andin vitro/in vivoassays for CSC viability and tumorigenicity.
  
  
  RESULTS
  We identified in ACC CD133-positive CSC that expressed NOTCH1 and SOX10, formed spheroids, and initiated tumors in nude mice. CD133(+)ACC cells produced activated NOTCH1 (N1ICD) and generated CD133(-)cells that expressed JAG1 as well as neural differentiation factors NR2F1, NR2F2, and p27Kip1. Knockdowns ofNOTCH1, SOX10, and their common effectorFABP7had negative effects on each other, inhibited spheroidogenesis, and induced cell death pointing at their essential roles in CSC maintenance. Downstream effects ofFABP7knockdown included suppression of a broad spectrum of genes involved in proliferation, ribosome biogenesis, and metabolism. Among proliferation-linked NOTCH1/FABP7 targets, we identified SKP2 and its substrate p27Kip1. A γ-secretase inhibitor, DAPT, selectively depleted CD133(+)cells, suppressed N1ICD and SKP2, induced p27Kip1, inhibited ACC growthin vivo, and sensitized CD133(+)cells to radiation.
  
  
  CONCLUSIONS
  These results establish in the majority of ACC the presence of a previously uncharacterized population of CD133(+)cells with neural stem properties, which are driven by SOX10, NOTCH1, and FABP7. Sensitivity of these cells to Notch inhibition and their dependence on SKP2 offer new opportunities for targeted ACC therapies.
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut. Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, Tennessee.
authors.forename: Alex
authors.initials: A
authors.lastname: Panaccione
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Michael T
authors.initials: MT
authors.lastname: Chang
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Beatrice E
authors.initials: BE
authors.lastname: Carbone
authors.affiliation: Department of Cancer Biology, Vanderbilt University School of Medicine, Nashville, Tennessee.
authors.forename: Yan
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Department of Pathology, University of Virginia, Charlottesville, Virginia.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Moskaluk
authors.affiliation: Department of Pathology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Renu K
authors.initials: RK
authors.lastname: Virk
authors.affiliation: Department of Pathology, New York University (NYU), New York, New York.
authors.forename: Luis
authors.initials: L
authors.lastname: Chiriboga
authors.affiliation: Department of Pathology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Manju L
authors.initials: ML
authors.lastname: Prasad
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Judson
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut.
authors.forename: Saral
authors.initials: S
authors.lastname: Mehra
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut. H&N Disease Center, Smilow Cancer Hospital, New Haven, Connecticut. Molecular Virology Program, Yale Cancer Center, New Haven, Connecticut.
authors.forename: Wendell G
authors.initials: WG
authors.lastname: Yarbrough
authors.affiliation: Department of Surgery, Section of Otolaryngology, Yale School of Medicine, New Haven, Connecticut. sergey.ivanov@yale.edu.
authors.forename: Sergey V
authors.initials: SV
authors.lastname: Ivanov
mesh_terms: D000071916:AC133 Antigen / Q000235:genetics / Q000378:metabolism; D053829:Amyloid Precursor Protein Secretases / Q000378:metabolism; D000818:Animals; D014408:Biomarkers, Tumor; D003528:Carcinoma, Adenoid Cystic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism; D004195:Disease Models, Animal; D000071438:Fatty Acid-Binding Protein 7 / Q000378:metabolism; D020869:Gene Expression Profiling; D064593:Heterografts; D006801:Humans; D008024:Ligands; D051379:Mice; D008819:Mice, Nude; D060787:Neoplasm Grading; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D011485:Protein Binding; D011827:Radiation; D011836:Radiation Tolerance / Q000235:genetics; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism*; D044786:S-Phase Kinase-Associated Proteins / Q000378:metabolism; D055757:SOXE Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins / Q000378:metabolism
chemical_list: D000071916:AC133 Antigen; D014408:Biomarkers, Tumor; C107608:FABP7 protein, human; D000071438:Fatty Acid-Binding Protein 7; D008024:Ligands; D051881:Receptor, Notch1; D044786:S-Phase Kinase-Associated Proteins; C527923:SOX10 protein, human; D055757:SOXE Transcription Factors; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D053829:Amyloid Precursor Protein Secretases