title: The enzymatic activity of human aldehyde dehydrogenases 1A2 and 2 (ALDH1A2 and ALDH2) is detected by Aldefluor, inhibited by diethylaminobenzaldehyde and has significant effects on cell proliferation and drug resistance.
abstract: There has been a new interest in using aldehyde dehydrogenase (ALDH) activity as one marker for stem cells since the Aldefluor flow cytometry-based assay has become available. Diethylaminobenzaldehyde (DEAB), used in the Aldeflour assay, has been considered a specific inhibitor for ALDH1A1 isoform. In this study, we explore the effects of human ALDH isoenzymes, ALDH1A2 and ALDH2, on drug resistance and proliferation, and the specificity of DEAB as an inhibitor. We also screened for the expression of 19 ALDH isoenzymes in K562 cells using TaqMan Low Density Array (TLDA). We used lentiviral vectors containing the full cDNA length of either ALDH2 or ALDH1A2 to over express the enzymes in K562 leukemia and H1299 lung cancer cell lines. Successful expression was measured by activity assay, Western blot, RT-PCR, and Aldefluor assay. Both cell lines, with either ALDH1A2 or ALDH2, exhibited higher cell proliferation rates, higher clonal efficiency, and increased drug resistance to 4-hydroperoxycyclophosphamide and doxorubicin. In order to study the specificity of known ALDH activity inhibitors, DEAB and disulfiram, we incubated each cell line with either inhibitor and measured the remaining ALDH enzymatic activity. Both inhibitors reduced ALDH activity of both isoenzymes by 65-90%. Furthermore, our TLDA results revealed that ALDH1, ALDH7, ALDH3 and ALDH8 are expressed in K562 cells. We conclude that DEAB is not a specific inhibitor for ALDH1A1 and that Aldefluor assay is not specific for ALDH1A1 activity. In addition, other ALDH isoenzymes seem to play a major role in the biology and drug resistance of various malignant cells.
authors.affiliation: Division of Hematology/Oncology, Department of Medicine, College of Medicine, University of Florida, Gainesville, FL, United States.
authors.forename: Jan S
authors.initials: JS
authors.lastname: Moreb
authors.forename: Deniz
authors.initials: D
authors.lastname: Ucar
authors.forename: Shuhong
authors.initials: S
authors.lastname: Han
authors.forename: John K
authors.initials: JK
authors.lastname: Amory
authors.forename: Alex S
authors.initials: AS
authors.lastname: Goldstein
authors.forename: Blanca
authors.initials: B
authors.lastname: Ostmark
authors.forename: Lung-Ji
authors.initials: LJ
authors.lastname: Chang
mesh_terms: D000444:Aldehyde Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000080924:Aldehyde Dehydrogenase 1 Family; D000071396:Aldehyde Dehydrogenase, Mitochondrial; D001547:Benzaldehydes / Q000737:chemistry / Q000494:pharmacology*; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003520:Cyclophosphamide / Q000031:analogs & derivatives / Q000494:pharmacology; D004221:Disulfiram / Q000737:chemistry / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D050697:Retinal Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001547:Benzaldehydes; D004791:Enzyme Inhibitors; C063555:4-hydroperoxydechlorocyclophosphamide; D004317:Doxorubicin; D003520:Cyclophosphamide; D000080924:Aldehyde Dehydrogenase 1 Family; C096127:ALDH2 protein, human; D000444:Aldehyde Dehydrogenase; D000071396:Aldehyde Dehydrogenase, Mitochondrial; C545372:ALDH1A2 protein, human; D050697:Retinal Dehydrogenase; D004221:Disulfiram
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