title: Intragenic deletions and a deep intronic mutation affecting pre-mRNA splicing in the dihydropyrimidine dehydrogenase gene as novel mechanisms causing 5-fluorouracil toxicity.
abstract: Dihydropyrimidine dehydrogenase (DPD) is the initial enzyme acting in the catabolism of the widely used antineoplastic agent 5-fluorouracil (5FU). DPD deficiency is known to cause a potentially lethal toxicity following administration of 5FU. Here, we report novel genetic mechanisms underlying DPD deficiency in patients presenting with grade III/IV 5FU-associated toxicity. In one patient a genomic DPYD deletion of exons 21-23 was observed. In five patients a deep intronic mutation c.1129-5923C>G was identified creating a cryptic splice donor site. As a consequence, a 44 bp fragment corresponding to nucleotides c.1129-5967 to c.1129-5924 of intron 10 was inserted in the mature DPD mRNA. The deleterious c.1129-5923C>G mutation proved to be in cis with three intronic polymorphisms (c.483 + 18G>A, c.959-51T>G, c.680 + 139G>A) and the synonymous mutation c.1236G>A of a previously identified haplotype. Retrospective analysis of 203 cancer patients showed that the c.1129-5923C>G mutation was significantly enriched in patients with severe 5FU-associated toxicity (9.1%) compared to patients without toxicity (2.2%). In addition, a high prevalence was observed for the c.1129-5923C>G mutation in the normal Dutch (2.6%) and German (3.3%) population. Our study demonstrates that a genomic deletion affecting DPYD and a deep intronic mutation affecting pre-mRNA splicing can cause severe 5FU-associated toxicity. We conclude that screening for DPD deficiency should include a search for genomic rearrangements and aberrant splicing.
authors.affiliation: Department of Clinical Chemistry, Academic Medical Center, Emma Children's Hospital, University of Amsterdam, Amsterdam, The Netherlands. a.b.vanKuilenburg@amc.uva.nl
authors.forename: André B P
authors.initials: AB
authors.lastname: van Kuilenburg
authors.forename: Judith
authors.initials: J
authors.lastname: Meijer
authors.forename: Adri N P M
authors.initials: AN
authors.lastname: Mul
authors.forename: Rutger
authors.initials: R
authors.lastname: Meinsma
authors.forename: Veronika
authors.initials: V
authors.lastname: Schmid
authors.forename: Doreen
authors.initials: D
authors.lastname: Dobritzsch
authors.forename: Raoul C M
authors.initials: RC
authors.lastname: Hennekam
authors.forename: Marcel M A M
authors.initials: MM
authors.lastname: Mannens
authors.forename: Marion
authors.initials: M
authors.lastname: Kiechle
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Etienne-Grimaldi
authors.forename: Heinz-Josef
authors.initials: HJ
authors.lastname: Klümpen
authors.forename: Jan Gerard
authors.initials: JG
authors.lastname: Maring
authors.forename: Veerle A
authors.initials: VA
authors.lastname: Derleyn
authors.forename: Ed
authors.initials: E
authors.lastname: Maartense
authors.forename: Gérard
authors.initials: G
authors.lastname: Milano
authors.forename: Raymon
authors.initials: R
authors.lastname: Vijzelaar
authors.forename: Eva
authors.initials: E
authors.lastname: Gross
mesh_terms: D000328:Adult; D000368:Aged; D000964:Antimetabolites, Antineoplastic / Q000633:toxicity*; D055028:Comparative Genomic Hybridization; D042943:Dihydrouracil Dehydrogenase (NADP) / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005472:Fluorouracil / Q000633:toxicity*; D015321:Gene Rearrangement; D006801:Humans; D007438:Introns; D008297:Male; D008875:Middle Aged; D020125:Mutation, Missense; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D012326:RNA Splicing; D012333:RNA, Messenger; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion
chemical_list: D000964:Antimetabolites, Antineoplastic; D012333:RNA, Messenger; D042943:Dihydrouracil Dehydrogenase (NADP); D005472:Fluorouracil
references.citation: Pharmacogenomics. 2009 Jun;10(6):931-44
references.pmid: 19530960
references.citation: Acta Crystallogr D Biol Crystallogr. 2010 Apr;66(Pt 4):486-501
references.pmid: 20383002
references.citation: Mol Cancer Ther. 2006 Nov;5(11):2895-904
references.pmid: 17121937
references.citation: Nat Rev Genet. 2007 Oct;8(10):749-61
references.pmid: 17726481
references.citation: Adv Enzyme Regul. 2001;41:151-7
references.pmid: 11384742
references.citation: C R Biol. 2007 Oct;330(10):764-9
references.pmid: 17905396
references.citation: J Inherit Metab Dis. 1996;19(5):645-54
references.pmid: 8892022
references.citation: Biol Chem. 2005 Apr;386(4):319-24
references.pmid: 15899693
references.citation: Cancer Chemother Pharmacol. 2009 Aug;64(3):615-8
references.pmid: 19288105
references.citation: Hum Genet. 2009 Jun;125(5-6):581-90
references.pmid: 19296131
references.citation: J Clin Oncol. 2008 May 1;26(13):2131-8
references.pmid: 18299612
references.citation: Nucleic Acids Res. 1987 Sep 11;15(17):7155-74
references.pmid: 3658675
references.citation: Ther Drug Monit. 2007 Apr;29(2):190-6
references.pmid: 17417073
references.citation: Eur J Cancer. 2007 Jan;43(2):459-65
references.pmid: 17097873
references.citation: PLoS One. 2008;3(12):e4003
references.pmid: 19104657
references.citation: N Engl J Med. 2005 Jun 30;352(26):2696-704
references.pmid: 15987918
references.citation: Clin Cancer Res. 2000 Dec;6(12):4705-12
references.pmid: 11156223
references.citation: Nucleic Acids Res. 2002 Jun 15;30(12):e57
references.pmid: 12060695
references.citation: Eur J Cancer. 2004 May;40(7):939-50
references.pmid: 15093568
references.citation: Hum Mutat. 2003 Dec;22(6):498
references.pmid: 14635116
references.citation: Br J Cancer. 1999 Feb;79(3-4):627-30
references.pmid: 10027340
references.citation: Int J Cancer. 2002 Sep 20;101(3):253-8
references.pmid: 12209976
references.citation: Nat Rev Genet. 2004 May;5(5):389-96
references.pmid: 15168696
references.citation: Cancer Chemother Pharmacol. 2010 Jan;65(2):403-6
references.pmid: 19795123
references.citation: J Clin Invest. 1996 Aug 1;98(3):610-5
references.pmid: 8698850
references.citation: N Engl J Med. 2005 Feb 3;352(5):476-87
references.pmid: 15689586
references.citation: Clin Cancer Res. 2004 Apr 15;10(8):2652-8
references.pmid: 15102667
references.citation: Pharmacogenetics. 2000 Apr;10(3):217-23
references.pmid: 10803677
references.citation: Clin Cancer Res. 2005 Aug 15;11(16):5886-92
references.pmid: 16115930
references.citation: Clin Cancer Res. 2001 May;7(5):1149-53
references.pmid: 11350878
references.citation: Cancer Lett. 2007 Feb 8;246(1-2):82-91
references.pmid: 16556484
references.citation: Genomics. 1998 Aug 1;51(3):391-400
references.pmid: 9721209
references.citation: Pharmacogenetics. 2002 Oct;12(7):555-8
references.pmid: 12360106
references.citation: Neoplasma. 2009;56(4):303-16
references.pmid: 19473056
references.citation: J Hum Genet. 2007;52(10):804-819
references.pmid: 17828463
references.citation: EMBO J. 2001 Feb 15;20(4):650-60
references.pmid: 11179210