title: Noninvasive prenatal detection of fetal trisomy 18 by epigenetic allelic ratio analysis in maternal plasma: Theoretical and empirical considerations.
abstract: BACKGROUND
  The discovery of cell-free fetal DNA in maternal plasma has opened up new possibilities for noninvasive prenatal diagnosis. However, the use of maternal plasma fetal DNA for the direct detection of fetal chromosomal aneuploidies has not been reported. We postulate that the aneuploidy status of a fetus could be revealed by an epigenetic allelic ratio approach, i.e., by analyzing the allelic ratio of a single-base variation present within DNA molecules exhibiting a placental-specific epigenetic signature in maternal plasma.
  
  
  METHODS
  Placental-derived fetal-specific unmethylated maspin (SERPINB5) promoter sequences on human chromosome 18 were detectable in placental-maternal DNA mixtures and in maternal plasma by bisulfite modification followed by methylation-specific PCR (MSP) and primer extension. The ratios between the extension products of the 2 alleles were calculated for heterozygous placentas, placental-maternal blood cell DNA mixtures, and maternal plasma samples. The allelic ratios were compared between pregnancies carrying trisomy 18 and euploid fetuses.
  
  
  RESULTS
  The epigenetic allelic ratios of all tested trisomy 18 samples deviated from the reference range obtained from euploid samples (placental DNA, 1.135 to 2.052; placental-maternal DNA mixtures, 1.170 to 1.985; maternal plasma, 0.330 to 3.044; without skew correction on the raw mass spectrometric data). A theoretical model was established and validated that predicted that a minimum of 200 copies of genomic DNA after bisulfite conversion were required for distinguishing euploid and aneuploid fetuses with confidence.
  
  
  CONCLUSIONS
  Epigenetic allelic ratio analysis of maternal plasma DNA represents a promising approach for noninvasive prenatal diagnosis of fetal chromosomal aneuploidies.
authors.affiliation: Department of Chemical Pathology, Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China.
authors.forename: Yu K
authors.initials: YK
authors.lastname: Tong
authors.forename: Chunming
authors.initials: C
authors.lastname: Ding
authors.forename: Rossa W K
authors.initials: RW
authors.lastname: Chiu
authors.forename: Ageliki
authors.initials: A
authors.lastname: Gerovassili
authors.forename: Stephen S C
authors.initials: SS
authors.lastname: Chim
authors.forename: Tak Y
authors.initials: TY
authors.lastname: Leung
authors.forename: Tse N
authors.initials: TN
authors.lastname: Leung
authors.forename: Tze K
authors.initials: TK
authors.lastname: Lau
authors.forename: Kypros H
authors.initials: KH
authors.lastname: Nicolaides
authors.forename: Y M Dennis
authors.initials: YM
authors.lastname: Lo
mesh_terms: D002887:Chromosomes, Human, Pair 18; D004247:DNA / Q000032:analysis / Q000097:blood; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005240:Feasibility Studies; D005260:Female; D005787:Gene Frequency; D016147:Genes, Tumor Suppressor; D005838:Genotype; D006801:Humans; D013058:Mass Spectrometry; D010920:Placenta / Q000737:chemistry; D020641:Polymorphism, Single Nucleotide; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D011401:Promoter Regions, Genetic; D012016:Reference Values; D012680:Sensitivity and Specificity; D015843:Serpins / Q000032:analysis / Q000097:blood / Q000235:genetics*; D014314:Trisomy / Q000175:diagnosis*
chemical_list: C085344:SERPIN-B5; D015843:Serpins; D004247:DNA