Noninvasive Prenatal Methylomic Analysis by Genomewide Bisulfite Sequencing of Maternal Plasma DNA

Lun, Fiona M.F.; Chiu, Rossa W.̉K.; Sun, Kun; Leung, Tak Yeung; Jiang, Peiyong; Chan, K. C. Allen; Sun, Hao; Lo, Y.M. Dennis

doi:10.1373/clinchem.2013.212274

Cited by 136 publications

(156 citation statements)

References 35 publications

(48 reference statements)

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“…Liquid biopsy has recently attracted the interest of clinicians as a possible approach that may overcome the influence of heterogeneity [5,6], with a number of studies being conducted to investigate intratumoral heterogeneity [7][8][9]. The potential of circulating tumor DNA (ctDNA) as the source of a template in minimally invasive diagnostics is becoming increasingly apparent for monitoring the status of tumors with temporal and spatial heterogeneities [10][11][12]. The detection and quantitation of specific nucleic acid sequences using a polymerase chain reaction (PCR) is fundamental to a large body of research and a growing number of molecular diagnostic tests, including HER2 amplification [13].…”

Section: Introductionmentioning

confidence: 99%

Monitoring the HER2 copy number status in circulating tumor DNA by droplet digital PCR in patients with gastric cancer

et al. 2016

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Background We previously demonstrated the potential of circulating tumor DNA (ctDNA) for the amplification of detecting HER2 in patients with gastric cancer (GC). In the present study, we focused on the clinical courses of patients who developed recurrence with GC, and investigated the potential clinical utility of the ddPCR-based HER2 copy number (CN) as a marker for the temporal and/or spatial heterogeneities of GC during treatment progress. Method We enrolled 30 healthy volunteers and 60 patients with GC who underwent surgery, including 17 patients who developed recurrence. Using ribonuclease P RNA component H1 (RPPH1) as a reference gene, plasma HER2 to RPPH1 ratios (the HER2 ratio) were determined using ddPCR. Results The preoperative plasma HER2 ratio correlated with the tumor HER2 status (p \ 0.001), and sensitivity and specificity were 0.733 and 0.933, respectively.Analyses of plasma samples during the postoperative follow-up periods revealed that high plasma HER2 ratios were detected at the time of recurrence in 7 of 13 cases, which were diagnosed as being HER2 negative at the time of surgery. These results were supported by continuously increasing HER2 ratios thereafter with the progression of recurrent cancer. Conclusion The plasma HER2 ratio determined by ddPCR is a repeatable and noninvasive approach for realtime evaluations of the HER2 status to monitor the effects of treatments for patients with HER2-positive GC and enable treatment options for patients with HER2-negative GC but positive conversion of the HER2 status after recurrence.

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Section: Introductionmentioning

confidence: 99%

Monitoring the HER2 copy number status in circulating tumor DNA by droplet digital PCR in patients with gastric cancer

et al. 2016

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“…Significantly elevated levels of total cell-free DNA and selected placenta-specific RNA transcripts have also been reported in the maternal plasma of women with PE (8)(9)(10)(11), restricted fetal growth (12), and preterm birth (13)(14)(15), supporting a role for cell-free nucleic acids as a noninvasive tool for placental monitoring. Previous studies have attempted to provide a comprehensive assessment of maternal plasma nucleic acids by microarray analysis, massively parallel transcriptomic, or methylomic sequencing (16)(17)(18)(19)(20)(21)(22). Several groups have explored the use of fetal-specific DNA polymorphisms, organ-specific DNA methylation (21), nucleosome footprinting (23), DNA fragmentation patterns (24), and tissuespecific RNA transcripts (19,20) to isolate the placental signal in the pool of circulating cell-free fetal nucleic acids and obtain changes of overall placental contribution.…”

mentioning

confidence: 99%

Integrative single-cell and cell-free plasma RNA transcriptomics elucidates placental cellular dynamics

Tsang

Vong

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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The human placenta is a dynamic and heterogeneous organ critical in the establishment of the fetomaternal interface and the maintenance of gestational well-being. It is also the major source of cell-free fetal nucleic acids in the maternal circulation. Placental dysfunction contributes to significant complications, such as preeclampsia, a potentially lethal hypertensive disorder during pregnancy. Previous studies have identified significant changes in the expression profiles of preeclamptic placentas using whole-tissue analysis. Moreover, studies have shown increased levels of targeted RNA transcripts, overall and placental contributions in maternal cell-free nucleic acids during pregnancy progression and gestational complications, but it remains infeasible to noninvasively delineate placental cellular dynamics and dysfunction at the cellular level using maternal cell-free nucleic acid analysis. In this study, we addressed this issue by first dissecting the cellular heterogeneity of the human placenta and defined individual cell-type-specific gene signatures by analyzing more than 24,000 nonmarker selected cells from full-term and early preeclamptic placentas using large-scale microfluidic single-cell transcriptomic technology. Our dataset identified diverse cellular subtypes in the human placenta and enabled reconstruction of the trophoblast differentiation trajectory. Through integrative analysis with maternal plasma cell-free RNA, we resolved the longitudinal cellular dynamics of hematopoietic and placental cells in pregnancy progression. Furthermore, we were able to noninvasively uncover the cellular dysfunction of extravillous trophoblasts in early preeclamptic placentas. Our work showed the potential of integrating transcriptomic information derived from single cells into the interpretation of cell-free plasma RNA, enabling the noninvasive elucidation of cellular dynamics in complex pathological conditions. single-cell transcriptomics | cell-free RNA | noninvasive prenatal testing | placenta | preeclampsia

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“…In most of the available methods, reactions can be prepared by massive parallel anchoring of numerous amplified molecules in a next generation sequencing (NGS) platform [24,27]. Thus, the number of DNA molecules that can be sequenced and consequently the amount of information produced is dramatically increased when compared to "first-generation" Sanger sequencing methods [34,35]. There are two different approaches in this concept: quantitative (either shotgun or targeted) and qualitative targeted single nucleotide polymorphism (SNP)-based methods [36].…”

Section: Available Nipt Techniquesmentioning

confidence: 99%

“…A very recent interesting development in using DNA methylation for NIPT has been the implementation of sodium bisulfite DNA treatment in combination with NGS [35]. This chemical treatment of the DNA is associated with a high degree of DNA degradation in > 90% of the template DNA [34].…”

Section: Epigenetic-based Nipt Approachesmentioning

confidence: 99%

“…This chemical treatment of the DNA is associated with a high degree of DNA degradation in > 90% of the template DNA [34]. So the accuracy and sensitivity of the test will be reduced because during pregnancy, the amount of fetal DNA in maternal plasma is already very low and further degradation will result in even fewer fetal DNA available for quantification [34,35]. On the other hand, manipulations for removal of sodium bisulfite will bias the correct interpretation of the results.…”

Section: Epigenetic-based Nipt Approachesmentioning

confidence: 99%

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Non-invasive prenatal testing (NIPT): limitations on the way to become diagnosis

Kotsopoulou¹,

Tsoplou²,

Mavrommatis

et al. 2015

Diagnosis

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With the discovery of existing circulating cellfree fetal DNA (ccffDNA) in maternal plasma and the advent of next-generation sequencing (NGS) technology, there is substantial hope that prenatal diagnosis will become a predominately non-invasive process in the future. At the moment, non-invasive prenatal testing (NIPT) is available for high-risk pregnancies with significant better sensitivity and specificity than the other existing non-invasive methods (biochemical and ultrasonographical). Mainly it is performed by NGS methods in a few commercial labs worldwide. However, it is expected that many other labs will offer analogous services in the future in this fastgrowing field with a multiplicity of in-house methods (e.g., epigenetic, etc.). Due to various limitations of the available methods and technologies that are explained in detail in this manuscript, NIPT has not become diagnostic yet and women may still need to undergo risky invasive procedures to verify a positive finding or to secure (or even expand) a negative one. Efforts have already started to make the NIPT technologies more accurate (even at the level of a complete fetal genome) and cheaper and thus more affordable, in order to become diagnostic screening tests for all pregnancies in the near future.

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Noninvasive Prenatal Methylomic Analysis by Genomewide Bisulfite Sequencing of Maternal Plasma DNA

Abstract: BACKGROUND:Epigenetic mechanisms play an important role in prenatal development, but fetal tissues are not readily accessible. Fetal DNA molecules are present in maternal plasma and can be analyzed noninvasively.

Cited by 136 publications

References 35 publications

Monitoring the HER2 copy number status in circulating tumor DNA by droplet digital PCR in patients with gastric cancer

Monitoring the HER2 copy number status in circulating tumor DNA by droplet digital PCR in patients with gastric cancer

Integrative single-cell and cell-free plasma RNA transcriptomics elucidates placental cellular dynamics

Non-invasive prenatal testing (NIPT): limitations on the way to become diagnosis

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