title: Role of CYB5A in pancreatic cancer prognosis and autophagy modulation.
abstract: BACKGROUND
  Loss of 18q22.3 is a prognostic marker in pancreatic ductal adenocarcinoma (PDAC). This study investigated genes encoded by this cytoband.
  
  
  METHODS
  We studied mRNA/protein expression in radically resected (n = 130) and metastatic patients (n = 50). The role of CYB5A was tested in 11 PDAC cell lines and five primary cultures through retrovirus-mediated upregulation and small interfering RNA using wound-healing, invasion, annexin-V, electron microscopy, and autophagic assays, as well as autophagy genes and kinases arrays. CYB5A+ orthotopic models (n = 6 mice/group) were monitored by Firefly and Gaussia-luciferase bioluminescence, magnetic resonance imaging, and high-frequency ultrasound. Data were analyzed by t test, Fisher exact-test, log-rank test and Cox proportional hazards models. All statistical tests were two-sided.
  
  
  RESULTS
  Both resected and metastatic patients with low mRNA or protein expression of CYB5A had statistically significantly shorter survival (eg, median = 16.7 months, 95% confidence interval [CI] = 13.5 to 19.9; vs median = 24.8 months, 95% CI = 12.8 to 36.9; P = .02, two-sided log-rank test; n = 82 radically resected PDACs), and multivariable analyses confirmed prognostic relevance. Moreover, we characterized a novel function to CYB5A, autophagy induction, concomitant with reduced proliferation and migration/invasion of PDAC cells. Network analysis of proautophagic pathways suggested CYB5A interaction with TRAF6, which was confirmed by TRAF6 downregulation after CYB5A reconstitution (-69% in SU.86.86-CYB5A+; P = .005, two-sided t test). CYB5A silencing had opposite effects, restoring TRAF6 expression and wound healing. In vivo studies showed that CYB5A induced autophagy while inhibiting tumor growth/metastasis and increasing survival (median = 57 days, 95% CI = 52 to 61; vs median = 44 days, 95% CI = 21 to 57; P = .03, two-sided log-rank test).
  
  
  CONCLUSIONS
  These results define CYB5A as a novel prognostic factor for PDAC that exerts its tumor-suppressor function through autophagy induction and TRAF6 modulation.
authors.affiliation: Affiliations of authors: Department of Medical Oncology (EG, AA, GJP) and Department of Neurosurgery (TL, VC, TW), VU University Medical Center, and Centre for Integrative Bioinformatics (AvdV), VU University, Amsterdam, the Netherlands; Department of Ricerca Traslazionale e delle Nuove Tecnologie in Medicina e Chirurgia, University of Pisa, Pisa, Italy (NF, UB, EV); Department of Neurology, Stanford University, Stanford, CA (VC); Department of Neurology, Massachusetts General Hospital and Neuroscience Program, Harvard Medical School, Boston, MA (TW); Medical Oncology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD (QW, J-HL, YW, GG).
authors.forename: Elisa
authors.initials: E
authors.lastname: Giovannetti
authors.forename: Qiuyan
authors.initials: Q
authors.lastname: Wang
authors.forename: Amir
authors.initials: A
authors.lastname: Avan
authors.forename: Niccola
authors.initials: N
authors.lastname: Funel
authors.forename: Tonny
authors.initials: T
authors.lastname: Lagerweij
authors.forename: Jih-Hsiang
authors.initials: JH
authors.lastname: Lee
authors.forename: Viola
authors.initials: V
authors.lastname: Caretti
authors.forename: Arjan
authors.initials: A
authors.lastname: van der Velde
authors.forename: Ugo
authors.initials: U
authors.lastname: Boggi
authors.forename: Yisong
authors.initials: Y
authors.lastname: Wang
authors.forename: Enrico
authors.initials: E
authors.lastname: Vasile
authors.forename: Godefridus J
authors.initials: GJ
authors.lastname: Peters
authors.forename: Thomas
authors.initials: T
authors.lastname: Wurdinger
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Giaccone
mesh_terms: D001343:Autophagy / Q000235:genetics; D021441:Carcinoma, Pancreatic Ductal / Q000201:enzymology*; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D002887:Chromosomes, Human, Pair 18; D015786:Cytochromes b5 / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D009361:Neoplasm Invasiveness; D010190:Pancreatic Neoplasms / Q000201:enzymology*; D011237:Predictive Value of Tests; D061251:Primary Cell Culture; D011379:Prognosis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048029:TNF Receptor-Associated Factor 6 / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
chemical_list: D048029:TNF Receptor-Associated Factor 6; D015786:Cytochromes b5