title: Growth suppression of acute promyelocytic leukemia cells having increased expression of the non-rearranged alleles: RAR alpha or PML.
abstract: The balanced t(15;17) rearrangement found in acute promyelocytic leukemia (APL) cells fuses PML on chromosome 15 to the retinoic acid receptor alpha (RAR alpha) on chromosome 17. PML/RAR alpha is expressed in APL cells with the non-rearranged alleles, PML and RAR alpha. Clinical remissions induced by all-trans-retinoic acid (RA) treatment of APL patients are linked to expression of PML/RAR apha, a transcription factor with reported dominant negative functions. The roles of PML and RAR alpha in the RA response of APL have not yet been fully explored. This study examines these roles by individually transfecting RAR alpha and PML into NB4 APL cells. NB4 is the sole APL cell line containing the t(15;17). RA treatment represses NB4 cell growth and induces a myeloid phentoype. Full length cDNAs for RAR alpha and PML were individually cloned into a CMV-driven expression vector containing the neomycin resistance gene. Surprisingly, none of the obtained stable transfectants expressed exogenous RAR alpha or PML mRNAs even when reverse transcription polymerase chain reaction (RT-PCR) detection assays were used. All clones expressed the neomycin resistance gene and were similar to parental NB4 cells in their growth and differentiation properties. An explanation explored for this lack of gene expression was that increased levels of RAR alpha or PML might suppress APL cell growth. To examine this possibility, transfection experiments were repeated using an episomal vector-based expression system containing an SV40 driven RAR alpha or PML cDNA and the hygromycin B resistance gene. A new selection strategy augmented expression of the desired cDNAs. A control episomal vector lacked a cDNA insert. Following electroporation and selection, exogenous RAR alpha expression was obtained. Compared to controls, the growth of these transfectants was markedly inhibited before and after RA-treatment and these cells more prominently induced myeloid maturation markers. In contrast, exogenous PML expression was transient since these transfectants did not appear to propagate in culture. These findings indicate: (1) a growth disadvantage for NB4 cells having increased expression of RAR alpha or PML and (2) increased RAR alpha expression augmented RA-mediated maturation of NB4 cells. This implicates a role for RAR alpha or PML in regulating the growth or differentiation of APL cells. It is hypothesized this occurs through antagonism of PML/RAR alpha actions in these leukemic cells.
authors.affiliation: Department of Medicine, Memorial Hospital, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
authors.forename: M J
authors.initials: MJ
authors.lastname: Ahn
authors.forename: K
authors.initials: K
authors.lastname: Nason-Burchenal
authors.forename: M M
authors.initials: MM
authors.lastname: Moasser
authors.forename: E
authors.initials: E
authors.lastname: Dmitrovsky
mesh_terms: D000483:Alleles; D001483:Base Sequence; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002884:Chromosomes, Human, Pair 15; D002886:Chromosomes, Human, Pair 17; D002934:Cinnamates; D004351:Drug Resistance / Q000235:genetics; D006801:Humans; D006921:Hygromycin B / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D008969:Molecular Sequence Data; D009355:Neomycin / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D000071156:Promyelocytic Leukemia Protein; D012333:RNA, Messenger / Q000378:metabolism; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism*; D000072482:Retinoic Acid Receptor alpha; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014178:Translocation, Genetic; D014212:Tretinoin / Q000494:pharmacology; D025521:Tumor Suppressor Proteins
chemical_list: D002934:Cinnamates; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion; D000071156:Promyelocytic Leukemia Protein; C000607071:RARA protein, human; D012333:RNA, Messenger; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C093476:promyelocytic leukemia-retinoic acid receptor alpha fusion oncoprotein; C070576:PML protein, human; D006921:Hygromycin B; C026273:hygromycin A; D014212:Tretinoin; D009355:Neomycin