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Research

Non-invasive prenatal assessment of trisomy 21 by multiplexed maternal plasma DNA sequencing: large scale validity study

BMJ 2011; 342 doi: https://doi.org/10.1136/bmj.c7401 (Published 11 January 2011) Cite this as: BMJ 2011;342:c7401
  1. Rossa W K Chiu, professor1,
  2. Ranjit Akolekar, clinical research fellow3,
  3. Yama W L Zheng, student 1,
  4. Tak Y Leung, professor2,
  5. Hao Sun, assistant professor1,
  6. K C Allen Chan, associate professor1,
  7. Fiona M F Lun, postdoctoral fellow1,
  8. Attie T J I Go, professor4,
  9. Elizabeth T Lau, department manager and honorary assistant professor5,
  10. William W K To, consultant6,
  11. Wing C Leung, consultant7,
  12. Rebecca Y K Tang, consultant8,
  13. Sidney K C Au-Yeung, consultant9,
  14. Helena Lam, consultant10,
  15. Yu Y Kung, obstetrician11,
  16. Xiuqing Zhang, manager1213,
  17. John M G van Vugt, professor4,
  18. Ryoko Minekawa, postdoctoral fellow3,
  19. Mary H Y Tang, consultant and honorary clinical associate professor5,
  20. Jun Wang, professor12, associate director13,
  21. Cees B M Oudejans, associate professor4,
  22. Tze K Lau, professor2,
  23. Kypros H Nicolaides, professor3,
  24. Y M Dennis Lo, professor112
  1. 1Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, Department of Chemical Pathology, The Chinese University of Hong Kong, Hong Kong SAR, China
  2. 2Department of Obstetrics and Gynaecology, The Chinese University of Hong Kong
  3. 3Harris Birthright Research Centre for Foetal Medicine, King’s College Hospital, London SE5 9RS, UK
  4. 4VU University Medical Center, 10081 HV Amsterdam, Netherlands
  5. 5Tsan Yuk Hospital, Department of Obstetrics and Gynaecology, University of Hong Kong, Hong Kong
  6. 6United Christian Hospital, Hospital Authority, Hong Kong
  7. 7Kwong Wah Hospital, Hospital Authority, Hong Kong
  8. 8Pamela Youde Nethersole Eastern Hospital, Hospital Authority, Hong Kong
  9. 9Tuen Mun Hospital, Hospital Authority, Hong Kong
  10. 10Princess Margaret Hospital, Hospital Authority, Hong Kong
  11. 11YY Kung Medical Centre, Hong Kong
  12. 12Joint Chinese University of Hong Kong-Beijing Genomics Institute Genome Research Centre, Hong Kong
  13. 13Beijing Genomics Institute at Shenzhen, Shenzhen, China
  1. Correspondence to: Y M D Lo loym{at}cuhk.edu.hk
  • Accepted 14 December 2010

Abstract

Objectives To validate the clinical efficacy and practical feasibility of massively parallel maternal plasma DNA sequencing to screen for fetal trisomy 21 among high risk pregnancies clinically indicated for amniocentesis or chorionic villus sampling.

Design Diagnostic accuracy validated against full karyotyping, using prospectively collected or archived maternal plasma samples.

Setting Prenatal diagnostic units in Hong Kong, United Kingdom, and the Netherlands.

Participants 753 pregnant women at high risk for fetal trisomy 21 who underwent definitive diagnosis by full karyotyping, of whom 86 had a fetus with trisomy 21.

Intervention Multiplexed massively parallel sequencing of DNA molecules in maternal plasma according to two protocols with different levels of sample throughput: 2-plex and 8-plex sequencing.

Main outcome measures Proportion of DNA molecules that originated from chromosome 21. A trisomy 21 fetus was diagnosed when the z score for the proportion of chromosome 21 DNA molecules was >3. Diagnostic sensitivity, specificity, positive predictive value, and negative predictive value were calculated for trisomy 21 detection.

Results Results were available from 753 pregnancies with the 8-plex sequencing protocol and from 314 pregnancies with the 2-plex protocol. The performance of the 2-plex protocol was superior to that of the 8-plex protocol. With the 2-plex protocol, trisomy 21 fetuses were detected at 100% sensitivity and 97.9% specificity, which resulted in a positive predictive value of 96.6% and negative predictive value of 100%. The 8-plex protocol detected 79.1% of the trisomy 21 fetuses and 98.9% specificity, giving a positive predictive value of 91.9% and negative predictive value of 96.9%.

Conclusion Multiplexed maternal plasma DNA sequencing analysis could be used to rule out fetal trisomy 21 among high risk pregnancies. If referrals for amniocentesis or chorionic villus sampling were based on the sequencing test results, about 98% of the invasive diagnostic procedures could be avoided.

Footnotes

  • We thank members of the Lo laboratory for technical assistance in performing the sequencing runs.

  • Contributors: RWKC and YMDL designed and initiated the study, coordinated and managed the research process, interpreted the results and wrote the first draft of the report. RA, ATJIG, ETL, JMGvV, and RM contributed to the implementation of the clinical recruitment protocols and management of clinical data. TYL, MHYT, CBMO, TKL, and KHN contributed to the design of the clinical recruitment protocols, coordination of the recruitment of patients and management of clinical data. YWLZ, KCAC, and FMFL contributed to the development of the laboratory protocols and monitoring of the laboratory analyses. HS and KCAC contributed to the development of the bioinformatics analysis protocols. WWKT, WCL, RYKT, SKCA-Y, HL, and YYK contributed to the recruitment, collection and management of clinical data. XZ and JW contributed to the laboratory analysis. All authors provided critical comments on the drafts of the manuscript and approved the final version. RWKC and YMDL had full access to all the data in the study, drafted the manuscript, take responsibility for the integrity of the data and the accuracy of the data analysis, and are the guarantors.

  • Competing interests: All authors have completed the Unified Competing Interest form at www.icmje.org/coi_disclosure.pdf (available on request from the corresponding author) and declare that (1) RWKC, HS, ETL, MHYT, TKL, and YMDL had support from the University Grants Committee of the Government of the Hong Kong Special Administrative Region, China, under the Areas of Excellence Scheme (AoE/M-04/06) for the submitted work; RWKC and YMDL had support from Sequenom for the submitted work; YMDL has support from an endowed chair from the Li Ka Shing Foundation. (2) YMDL is a consultant to, and holds equities in, Sequenom; RWKC and YMDL have received travel grants from Life Technologies and Illumina; RWKC, YWLZ, HS, KCAC, FMFL, and YMDL have received travel grants from University Grants Committee of the Government of the Hong Kong Special Administrative Region, China, under the Areas of Excellence Scheme (AoE/M-04/06). (3) The authors’ spouses, partners, and children have no financial relationships that may be relevant to the submitted work. (4) RWKC, YWLZ, KCAC, FMFL, and YMDL have filed patent applications on the detection of fetal nucleic acids in maternal plasma for non-invasive prenatal diagnosis. Part of this patent portfolio has been licensed to Sequenom and the Institut Jacques Boyd. The funders and sponsors have no role in the study design and the collection, analysis, and interpretation of data and the writing of the article and the decision to submit it for publication. The researchers are independent of the funders and sponsors.

  • Ethical approval: Approvals were obtained from the institutional review boards of each recruitment site: Joint Chinese University of Hong Kong-New Territories East Cluster Clinical Research Ethics Committee, Joint Institutional Review Board of the University of Hong Kong-Hospital Authority Hong Kong West Cluster, Clinical and Research Ethics Committees of the Hospital Authority in the Kowloon Central/Kowloon East, Kowloon West, Hong Kong East, New Territories West Clusters, King’s College Hospital Ethics Committee. and Ethics Committee of the VU University Medical Center. All participants gave informed written consent.

  • Data sharing: No additional data available.

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