title: An RNA editing fingerprint of cancer stem cell reprogramming.
abstract: BACKGROUND
  Deregulation of RNA editing by adenosine deaminases acting on dsRNA (ADARs) has been implicated in the progression of diverse human cancers including hematopoietic malignancies such as chronic myeloid leukemia (CML). Inflammation-associated activation of ADAR1 occurs in leukemia stem cells specifically in the advanced, often drug-resistant stage of CML known as blast crisis. However, detection of cancer stem cell-associated RNA editing by RNA sequencing in these rare cell populations can be technically challenging, costly and requires PCR validation. The objectives of this study were to validate RNA editing of a subset of cancer stem cell-associated transcripts, and to develop a quantitative RNA editing fingerprint assay for rapid detection of aberrant RNA editing in human malignancies.
  
  
  METHODS
  To facilitate quantification of cancer stem cell-associated RNA editing in exons and intronic or 3'UTR primate-specific Alu sequences using a sensitive, cost-effective method, we established an in vitro RNA editing model and developed a sensitive RNA editing fingerprint assay that employs a site-specific quantitative PCR (RESSq-PCR) strategy. This assay was validated in a stably-transduced human leukemia cell line, lentiviral-ADAR1 transduced primary hematopoietic stem and progenitor cells, and in primary human chronic myeloid leukemia stem cells.
  
  
  RESULTS
  In lentiviral ADAR1-expressing cells, increased RNA editing of MDM2, APOBEC3D, GLI1 and AZIN1 transcripts was detected by RESSq-PCR with improved sensitivity over sequencing chromatogram analysis. This method accurately detected cancer stem cell-associated RNA editing in primary chronic myeloid leukemia samples, establishing a cancer stem cell-specific RNA editing fingerprint of leukemic transformation that will support clinical development of novel diagnostic tools to predict and prevent cancer progression.
  
  
  CONCLUSIONS
  RNA editing quantification enables rapid detection of malignant progenitors signifying cancer progression and therapeutic resistance, and will aid future RNA editing inhibitor development efforts.
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. lcrews@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. lcrews@ucsd.edu.
authors.forename: Leslie A
authors.initials: LA
authors.lastname: Crews
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. q1jiang@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. q1jiang@ucsd.edu.
authors.forename: Qingfei
authors.initials: Q
authors.lastname: Jiang
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. mzipeto@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. mzipeto@ucsd.edu.
authors.forename: Maria A
authors.initials: MA
authors.lastname: Zipeto
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. elazzari@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. elazzari@ucsd.edu.|Doctoral School of Molecular and Translational Medicine, Department of Health Sciences, University of Milan, Milan, Italy. elazzari@ucsd.edu.
authors.forename: Elisa
authors.initials: E
authors.lastname: Lazzari
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. acourtrecart@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. acourtrecart@ucsd.edu.
authors.forename: Angela C
authors.initials: AC
authors.lastname: Court
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. sha003@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. sha003@ucsd.edu.
authors.forename: Shawn
authors.initials: S
authors.lastname: Ali
authors.affiliation: Division of Genome Information Sciences, Department of Pediatrics, University of California, La Jolla, CA, 92093, USA. cbarrett@ucsd.edu.
authors.forename: Christian L
authors.initials: CL
authors.lastname: Barrett
authors.affiliation: Division of Genome Information Sciences, Department of Pediatrics, University of California, La Jolla, CA, 92093, USA. kafrazer@ucsd.edu.
authors.forename: Kelly A
authors.initials: KA
authors.lastname: Frazer
authors.affiliation: Division of Regenerative Medicine, Department of Medicine, Moores Cancer Center at University of California, La Jolla, CA, 92093, USA. cjamieson@ucsd.edu.|Sanford Consortium for Regenerative Medicine, La Jolla, CA, 92037, USA. cjamieson@ucsd.edu.
authors.forename: Catriona H M
authors.initials: CH
authors.lastname: Jamieson
mesh_terms: D000243:Adenosine Deaminase / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000378:metabolism; D001752:Blast Crisis / Q000473:pathology; D065150:Cellular Reprogramming; D018920:Coculture Techniques; D018450:Disease Progression; D006801:Humans; D020014:K562 Cells; D016086:Lentivirus / Q000378:metabolism; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics / Q000473:pathology; D008954:Models, Biological; D014411:Neoplastic Stem Cells / Q000473:pathology*; D017393:RNA Editing / Q000235:genetics*; D015203:Reproducibility of Results
chemical_list: D014408:Biomarkers, Tumor; D000243:Adenosine Deaminase