Haplotype-Based Noninvasive Prenatal Diagnosis for Duchenne Muscular Dystrophy: A pilot study in South China

AbstractObjectiveTo explore the accuracy and feasibility of noninvasive prenatal diagnosis (NIPD) for Duchenne Muscular Dystrophy (DMD) based on the haplotype approach.MethodsWe recruited singleton pregnancies at-risk of DMD at 12-25 weeks of gestation from 17 families who all had a proband children affected by DMD. We have identified the pathogenic mutations in probands and their mothers by multiplex ligation-dependent probe amplification (MLPA). To construct parental haplotypes, we performed captured sequencing on genomic DNA from parents and probands. The integration analysis of parental haplotypes and targeted sequencing results of maternal plasma DNA were used to infer the fetal haplotype and genotypes in DMD gene. Fetal DMD genotypes were further confirmed by invasive prenatal diagnosis.ResultsWe have successfully performed the haplotype-based NIPD in all recruited families. Ten fetuses were identified as normal, including four female and six male fetuses. Four female fetuses were carriers and the other three male fetuses were affected by DMD with exons 49-52 deletion, exons 8-37 deletion and c.628G > T mutation, respectively. The results of NIPD were consistent with those of invasive diagnosis.ConclusionHaplotype-based NIPD for DMD by targeted sequencing is promising and has potential for clinical application.

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Noninvasive Prenatal Diagnosis of Monogenic Diseases by Targeted Massively Parallel Sequencing of Maternal Plasma: Application to β-Thalassemia

Clinical Chemistry ◽

10.1373/clinchem.2012.189589 ◽

2012 ◽

Vol 58 (10) ◽

pp. 1467-1475 ◽

Cited By ~ 112

Author(s):

Kwan-Wood G Lam ◽

Peiyong Jiang ◽

Gary J W Liao ◽

K C Allen Chan ◽

Tak Y Leung ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Massively Parallel Sequencing ◽

Globin Gene ◽

Maternal Plasma ◽

Massively Parallel ◽

Sequencing Data ◽

Plasma Dna ◽

Parallel Sequencing ◽

Noninvasive Prenatal Diagnosis ◽

Monogenic Diseases

Abstract BACKGROUND A genomewide genetic and mutational profile of a fetus was recently determined via deep sequencing of maternal plasma DNA. This technology could have important applications for noninvasive prenatal diagnosis (NIPD) of many monogenic diseases. Relative haplotype dosage (RHDO) analysis, a core step of this procedure, would allow one to elucidate the maternally inherited half of the fetal genome. For clinical applications, the cost and complexity of data analysis might be reduced via targeted application of this approach to selected genomic regions containing disease-causing genes. There is thus a need to explore the feasibility of performing RHDO analysis in a targeted manner. METHODS We performed target enrichment by using solution-phase hybridization followed by massively parallel sequencing of the β-globin gene region in 2 families undergoing prenatal diagnosis for β-thalassemia. We used digital PCR strategies to physically deduce parental haplotypes. Finally, we performed RHDO analysis with target-enriched sequencing data and parental haplotypes to reveal the β-thalassemic status for the fetuses. RESULTS A mean sequencing depth of 206-fold was achieved in the β-globin gene region by targeted sequencing of maternal plasma DNA. RHDO analysis was successful for the sequencing data obtained from the target-enriched samples, including a region in one of the families in which the parents had similar haplotype structures. Data analysis revealed that both fetuses were heterozygous carriers of β-thalassemia. CONCLUSIONS Targeted sequencing of maternal plasma DNA for NIPD of monogenic diseases is feasible.

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Haplotype-Based noninvasive prenatal diagnosis for duchenne muscular dystrophy: A pilot study in South China

European Journal of Obstetrics & Gynecology and Reproductive Biology ◽

10.1016/j.ejogrb.2019.05.005 ◽

2019 ◽

Vol 240 ◽

pp. 15-22 ◽

Cited By ~ 1

Author(s):

Min Chen ◽

Chao Chen ◽

Yingting Li ◽

Yuan Yuan ◽

Zhengfei Lai ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Pilot Study ◽

Muscular Dystrophy ◽

South China ◽

Noninvasive Prenatal Diagnosis

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Haplotype-based Noninvasive Prenatal Diagnosis of Hyperphenylalaninemia through Targeted Sequencing of Maternal Plasma

Scientific Reports ◽

10.1038/s41598-017-18358-y ◽

2018 ◽

Vol 8 (1) ◽

Cited By ~ 7

Author(s):

Jun Ye ◽

Chao Chen ◽

Yuan Yuan ◽

Lianshu Han ◽

Yaoshen Wang ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Maternal Plasma ◽

Targeted Sequencing ◽

Noninvasive Prenatal Diagnosis

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Noninvasive Prenatal Diagnosis for Duchenne Muscular Dystrophy Based on the Direct Haplotype Phasing

10.1101/720987 ◽

2019 ◽

Author(s):

Min Chen ◽

Chao Chen ◽

Xiaoyan Huang ◽

Jun Sun ◽

Lu Jiang ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Chorionic Villus ◽

Chorionic Villus Sampling ◽

Singleton Pregnancy ◽

High Molecular Weight Dna ◽

Haplotype Phasing ◽

Noninvasive Prenatal Diagnosis ◽

Mutational Status

AbstractObjectiveWe aimed to investigate the validity of noninvasive prenatal diagnosis (NIPD) based on direct haplotype phasing without the proband and its feasibility for clinical application in the case of Duchenne Muscular Dystrophy (DMD).MethodsThirteen singleton-pregnancy families affected by DMD were recruited. Firstly, we resolved maternal haplotypes for each family by performing targeted linked-read sequencing of their high molecular weight DNA, respectively. Then, we identified SNPs of the DMD gene in all carrier mothers and inferred the DMD gene mutation status of all fetuses. Finally, the fetal genotypes were further validated by using chorionic villus sampling.ResultsThe method of directly resolving maternal haplotype through targeted linked-read sequencing was smoothly performed in all participated families. The predicted mutational status of 13 fetuses was correct, which had been confirmed by invasive prenatal diagnosis.ConclusionDirect haplotyping of NIPD based on linked-read sequencing for DMD is accurate.

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AB095. Prenatal diagnosis of Duchenne muscular dystrophy by combining of multiplex Polymerase Chain Reaction and Multiplex Ligation dependent Probe Amplification

Annals of Translational Medicine ◽

10.21037/atm.2017.s095 ◽

2017 ◽

Vol 5 (S2) ◽

pp. AB095-AB095

Author(s):

Manh Tien Ngo ◽

Thi Phuong Mai Nguyen ◽

Thi Tuyet Nhung Ngo ◽

Thi Mai Huong Nguyen ◽

Thuy Lan An ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Multiplex Polymerase Chain Reaction ◽

Chain Reaction ◽

Polymerase Chain ◽

Dependent Probe

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Maternal Plasma DNA Analysis with Massively Parallel Sequencing by Ligation for Noninvasive Prenatal Diagnosis of Trisomy 21

Clinical Chemistry ◽

10.1373/clinchem.2009.136507 ◽

2010 ◽

Vol 56 (3) ◽

pp. 459-463 ◽

Cited By ~ 99

Author(s):

Rossa WK Chiu ◽

Hao Sun ◽

Ranjit Akolekar ◽

Christopher Clouser ◽

Clarence Lee ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Trisomy 21 ◽

Massively Parallel Sequencing ◽

Maternal Plasma ◽

Chromosome 21 ◽

Massively Parallel ◽

Plasma Dna ◽

Parallel Sequencing ◽

Noninvasive Prenatal Diagnosis ◽

Sequencing By Synthesis

Abstract Background: Noninvasive prenatal diagnosis of trisomy 21 (T21) has recently been shown to be achievable by massively parallel sequencing of maternal plasma on a sequencing-by-synthesis platform. The quantification of several other human chromosomes, including chromosomes 18 and 13, has been shown to be less precise, however, with quantitative biases related to the chromosomal GC content. Methods: Maternal plasma DNA from 10 euploid and 5 T21 pregnancies was sequenced with a sequencing-by-ligation approach. We calculated the genomic representations (GRs) of sequenced reads from each chromosome and their associated measurement CVs and compared the GRs of chromosome 21 (chr21) for the euploid and T21 pregnancies. Results: We obtained a median of 12 × 106 unique reads (21% of the total reads) per sample. The GRs deviated from those expected for some chromosomes but in a manner different from that previously reported for the sequencing-by-synthesis approach. Measurements of the GRs for chromosomes 18 and 13 were less precise than for chr21. z Scores of the GR of chr21 were increased in the T21 pregnancies, compared with the euploid pregnancies. Conclusions: Massively parallel sequencing-by-ligation of maternal plasma DNA was effective in identifying T21 fetuses noninvasively. The quantitative biases observed among the GRs of certain chromosomes were more likely based on analytical factors than biological factors. Further research is needed to enhance the precision for measuring for the representations of chromosomes 18 and 13.

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Haplotype-Based Noninvasive Prenatal Diagnosis of 21 Families With Duchenne Muscular Dystrophy: Real-World Clinical Data in China

Frontiers in Genetics ◽

10.3389/fgene.2021.791856 ◽

2021 ◽

Vol 12 ◽

Author(s):

Lingrong Kong ◽

Shaojun Li ◽

Zhenhua Zhao ◽

Jun Feng ◽

Guangquan Chen ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Family Members ◽

Chorionic Villus ◽

Genetic Diagnosis ◽

Blood Collection ◽

Chorionic Villus Sampling ◽

Clinical Environment ◽

Noninvasive Prenatal Diagnosis

Noninvasive prenatal diagnosis (NIPD) of single-gene disorders has recently become the focus of clinical laboratories. However, reports on the clinical application of NIPD of Duchenne muscular dystrophy (DMD) are limited. This study aimed to evaluate the detection performance of haplotype-based NIPD of DMD in a real clinical environment. Twenty-one DMD families at 7–12 weeks of gestation were prospectively recruited. DNA libraries of cell-free DNA from the pregnant and genomic DNA from family members were captured using a custom assay for the enrichment of DMD gene exons and spanning single-nucleotide polymorphisms, followed by next-generation sequencing. Parental haplotype phasing was based on family linkage analysis, and fetal genotyping was inferred using the Bayes factor through target maternal plasma sequencing. Finally, the entire experimental process was promoted in the local clinical laboratory. We recruited 13 complete families, 6 families without paternal samples, and 2 families without probands in which daughter samples were collected. Two different maternal haplotypes were constructed based on family members in all 21 pedigrees at as early as 7 gestational weeks. Among the included families, the fetal genotypes of 20 families were identified at the first blood collection, and a second blood collection was performed for another family due to low fetal concentration. The NIPD result of each family was reported within 1 week. The fetal fraction in maternal cfDNA ranged from 1.87 to 11.68%. In addition, recombination events were assessed in two fetuses. All NIPD results were concordant with the findings of invasive prenatal diagnosis (chorionic villus sampling or amniocentesis). Exon capture and haplotype-based NIPD of DMD are regularly used for DMD genetic diagnosis, carrier screening, and noninvasive prenatal diagnosis in the clinic. Our method, haplotype-based early screening for DMD fetal genotyping via cfDNA sequencing, has high feasibility and accuracy, a short turnaround time, and is inexpensive in a real clinical environment.

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Noninvasive prenatal diagnosis of duchenne muscular dystrophy in five Chinese families based on relative mutation dosage approach

BMC Medical Genomics ◽

10.1186/s12920-021-01128-1 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Ganye Zhao ◽

Xiaofeng Wang ◽

Lina Liu ◽

Peng Dai ◽

Xiangdong Kong

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Single Molecule ◽

De Novo ◽

Dna Barcode ◽

De Novo Mutations ◽

Molecular Alteration ◽

Noninvasive Prenatal Diagnosis ◽

Exon Deletion

Abstract Background Relative haplotype dosage (RHDO) approach has been applied in noninvasive prenatal diagnosis (NIPD) of Duchenne muscular dystrophy (DMD). However, the RHDO procedure is relatively complicated and the parental haplotypes need to be constructed. Furthermore, it is not suitable for the diagnosis of de novo mutations or mosaicism in germ cells. Here, we investigated NIPD of DMD using a relative mutation dosage (RMD)-based approach—cell-free DNA Barcode-Enabled Single-Molecule Test (cfBEST), which has not previously been applied in the diagnosis of exon deletion. Methods Five DMD families caused by DMD gene point mutations or exon deletion were recruited for this study. After the breakpoints of exon deletion were precisely mapped with multiple PCR, the genotypes of the fetuses from the five DMD families were inferred using cfBEST, and were further validated by invasive prenatal diagnosis. Results The cfBEST results of the five families indicated that one fetus was female and did not carry the familial molecular alteration, three fetuses were carriers and one was male without the familial mutation. The invasive prenatal diagnosis results were consistent with those of the cfBEST procedure. Conclusion This is the first report of NIPD of DMD using the RMD-based approach. We extended the application of cfBEST from point mutation to exon deletion mutation. The results showed that cfBEST would be suitable for NIPD of DMD caused by different kinds of mutation types.

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