title: Aldehyde dehydrogenase activity plays a Key role in the aggressive phenotype of neuroblastoma.
abstract: The successful targeting of neuroblastoma (NB) by associating tumor-initiating cells (TICs) is a major challenge in the development of new therapeutic strategies. The subfamily of aldehyde dehydrogenases 1 (ALDH1) isoenzymes, which comprises ALDH1A1, ALDH1A2, and ALDH1A3, is involved in the synthesis of retinoic acid, and has been identified as functional stem cell markers in diverse cancers. By combining serial neurosphere passages with gene expression profiling, we have previously identified ALDH1A2 and ALDH1A3 as potential NB TICs markers in patient-derived xenograft tumors. In this study, we explored the involvement of ALDH1 isoenzymes and the related ALDH activity in NB aggressive properties.
  
  
  
  ALDH activity and ALDH1A1/A2/A3 expression levels were measured using the ALDEFLUOR™ kit, and by real-time PCR, respectively. ALDH activity was inhibited using the specific ALDH inhibitor diethylaminobenzaldehyde (DEAB), and ALDH1A3 gene knock-out was generated through the CRISPR/Cas9 technology.
  
  
  
  We first confirmed the enrichment of ALDH1A2 and ALDH1A3 mRNA expression in NB cell lines and patient-derived xenograft tumors during neurosphere passages. We found that high ALDH1A1 expression was associated with less aggressive NB tumors and cell lines, and correlated with favorable prognostic factors. In contrast, we observed that ALDH1A3 was more widely expressed in NB cell lines and was associated with poor survival and high-risk prognostic factors. We also identified an important ALDH activity in various NB cell lines and patient-derived xenograft tumors. Specific inhibition of ALDH activity with diethylaminobenzaldehyde (DEAB) resulted in a strong reduction of NB cell clonogenicity, and TIC self-renewal potential, and partially enhanced NB cells sensitivity to 4-hydroxycyclophosphamide. Finally, the specific knock-out of ALDH1A3 via CRISPR/Cas9 gene editing reduced NB cell clonogenicity, and mediated a cell type-dependent inhibition of TIC self-renewal properties.
  
  
  
  Together our data uncover the participation of ALDH enzymatic activity in the aggressive properties and 4-hydroxycyclophosphamide resistance of NB, and show that the specific ALDH1A3 isoenzyme increases the aggressive capacities of a subset of NB cells.
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Marjorie
authors.initials: M
authors.lastname: Flahaut
authors.affiliation: Pediatric Surgery, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Jauquier
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.|Department of Physiology, University of Lausanne, Lausanne, Switzerland.
authors.forename: Nadja
authors.initials: N
authors.lastname: Chevalier
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Katya
authors.initials: K
authors.lastname: Nardou
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Katia
authors.initials: K
authors.lastname: Balmas Bourloud
authors.affiliation: Pediatric Surgery, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Joseph
authors.affiliation: SIB Swiss Institute of Bioinformatics, Bioinformatics Core Facility, Lausanne, Switzerland.
authors.forename: David
authors.initials: D
authors.lastname: Barras
authors.affiliation: Department of Physiology, University of Lausanne, Lausanne, Switzerland.
authors.forename: Christian
authors.initials: C
authors.lastname: Widmann
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Nicole
authors.initials: N
authors.lastname: Gross
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland.
authors.forename: Raffaele
authors.initials: R
authors.lastname: Renella
authors.affiliation: Pediatric Hematology-Oncology Research Laboratory, Pediatric Division, University Hospital CHUV, Lausanne, Switzerland. Annick.Muhlethaler@chuv.ch.
authors.forename: Annick
authors.identifier: 0000-0001-8274-0907
authors.initials: A
authors.lastname: Mühlethaler-Mottet
mesh_terms: D000444:Aldehyde Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D000080924:Aldehyde Dehydrogenase 1 Family; D000445:Aldehyde Oxidoreductases / Q000235:genetics / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D018450:Disease Progression; D004789:Enzyme Activation; D015870:Gene Expression; D055786:Gene Knockout Techniques; D064593:Heterografts; D006801:Humans; D007527:Isoenzymes; D051379:Mice; D009447:Neuroblastoma / Q000175:diagnosis* / Q000201:enzymology* / Q000235:genetics; D010641:Phenotype; D011379:Prognosis; D050697:Retinal Dehydrogenase / Q000235:genetics / Q000378:metabolism; D059467:Transcriptome
chemical_list: D007527:Isoenzymes; D000445:Aldehyde Oxidoreductases; D000080924:Aldehyde Dehydrogenase 1 Family; D000444:Aldehyde Dehydrogenase; C545372:ALDH1A2 protein, human; D050697:Retinal Dehydrogenase; C018699:aldehyde dehydrogenase (NAD(P)+)