BRCA Testing by Single-Molecule Molecular Inversion Probes

Neveling, Kornelia; Mensenkamp, Arjen R.; Derks, Ronny; Kwint, Michael; Ouchene, Hicham; Steehouwer, Marloes; Lier, Bart van; Bosgoed, Ermanno; Rikken, Alwin; Tychon, Marloes; Zafeiropoulou, Dimitra; Castelein, Steven; Hehir-Kwa, Jayne Y.; Thung, Djie Tjwan; Hofste, Tom; Lelieveld, Stefan H.; Bertens, Stijn M M; ADAN, IVO; Eijkelenboom, Astrid; Tops, Bastiaan B.J.; Yntema, Helger G.; Stokowy, Tomasz; Knappskog, Per M.; Høberg-Vetti, Hildegunn; Steen, Vidar M.; Boyle, Evan; Martin, Beth; Ligtenberg, Marjolijn J. L.; Shendure, Jay; Nelen, Marcel; Hoischen, Alexander

doi:10.1373/clinchem.2016.263897

Cited by 53 publications

(69 citation statements)

References 38 publications

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“…MIPs provide a high target sensitivity (99%) and specificity (98%) at low costs and minimal DNA requirements, and can be easily multiplexed to target regions of multiple genes in a single reaction [18]. Implementation of this methodology in routine diagnostics requires further adjustments to guarantee minimal costs and optimal turnaround times, which are highly dependent on sample supply, available infrastructure, and local personnel costs [19, 20]. For a MIP-based breast cancer susceptibility gene panel, a turnaround time of 4 days has been described [19].…”

Section: Discussionmentioning

confidence: 99%

“…Implementation of this methodology in routine diagnostics requires further adjustments to guarantee minimal costs and optimal turnaround times, which are highly dependent on sample supply, available infrastructure, and local personnel costs [19, 20]. For a MIP-based breast cancer susceptibility gene panel, a turnaround time of 4 days has been described [19]. Therefore, we consider MIP-based sequencing as a highly suitable replacement of Sanger sequencing for clinical genetic testing [17].…”

Section: Discussionmentioning

confidence: 99%

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A molecular inversion probe-based next-generation sequencing panel to detect germline mutations in Chinese early-onset colorectal cancer patients

et al. 2017

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The currently known Mendelian colorectal cancer (CRC) predisposition syndromes account for ∼5–10% of all CRC cases, and are caused by inherited germline mutations in single CRC predisposing genes. Using molecular inversion probes (MIPs), we designed a targeted next-generation sequencing panel to identify mutations in seven CRC predisposing genes: APC, MLH1, MSH2, MSH6, PMS2, MUTYH and NTHL1. From a consecutive series of 2,371 Chinese CRC patients, 140 familial and non-familial cases were selected that were diagnosed with CRC at or below the age of 35 years. Through MIP-based sequencing we identified pathogenic variants in six genes in 16 out of the 140 (11.4%) patients selected. In 10 patients, known pathogenic mutations in APC (five patients), MLH1 (three patients), or MSH2 (two patients) were identified. Three additional patients were found to carry novel, likely pathogenic truncating (n = 2) and missense (n = 1) mutations in the MSH2 gene and a concomitant loss of expression of both the MSH2 and MSH6 proteins in their respective tumor tissues. From our data, we conclude that targeted MIP-based sequencing is a reliable and cost-efficient approach to identify patients with a Mendelian CRC syndrome.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

A molecular inversion probe-based next-generation sequencing panel to detect germline mutations in Chinese early-onset colorectal cancer patients

et al. 2017

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“…Fifth, our analytic strategy enables the robust detection of MSI status without the need for sequencing patient-matched normal material (22). Lastly, the costs of library construction and required sequencing coverage by this approach are low, enabling economic testing of patient samples at scale (33); here, specimens were prepared and sequenced for <$80 USD in material costs each.…”

Section: Discussionmentioning

confidence: 99%

Accurate Pan-Cancer Molecular Diagnosis of Microsatellite Instability by Single-Molecule Molecular Inversion Probe Capture and High-Throughput Sequencing

et al. 2018

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BACKGROUND: Microsatellite instability (MSI) is an emerging, actionable phenotype in oncology that informs tumor response to immune checkpoint pathway immunotherapy. However, there remains a need for MSI diagnostics that are low cost, highly accurate, and generalizable across cancer types. We developed a method for targeted high throughput sequencing of numerous microsatellite loci with pan-cancer informativity for MSI using single-molecule molecular inversion probes (smMIPs). METHODS: We designed a smMIP panel targeting 111 loci highly informative for MSI across cancers. We developed an analytical framework taking advantage of smMIP-mediated error correction to specifically and sensitively detect instability events without the need for typing matched normal material. RESULTS: Using synthetic DNA mixtures, smMIPs were sensitive to at least 1% MSI positive cells and were highly consistent across replicates. The fraction of identified unstable microsatellites discriminated tumors exhibiting MSI from those lacking MSI with high accuracy across colorectal (100% diagnostic sensitivity and specificity), prostate (100% diagnostic sensitivity and specificity), and endometrial cancers (95.8% diagnostic sensitivity and 100% specificity). MSI-PCR, the current standard-of-care molecular diagnostic for MSI, proved equally robust for colorectal tumors, but evidenced multiple false negative results in prostate (81.8% diagnostic sensitivity and 100% specificity) and endometrial (75.0% diagnostic sensitivity and 100% specificity) tumors. CONCLUSIONS: smMIP capture provides an accurate, diagnostically sensitive, and economical means to diagnose MSI across cancer types without reliance on patient-matched normal material. The assay is readily scalable to large numbers of clinical samples, enables automated and quantitative analysis of microsatellite instability, and is readily standardized across clinical laboratories.

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“…Third, the quantitative nature of smMIP data enables computational inference of the genotypes of major and multiple minor cell populations without the need for independently genotyping donor and recipient material, as has proven necessary by both conventional STR (10,12) and PCR-based copy number deletion assays (12,13), and eliminates the need for calibration by standard curve (12). Fourth, smMIP capture is cost effective and readily scalable to multiple patient specimens (38), facilitating high throughput testing in clinical practice.…”

Section: Discussionmentioning

confidence: 99%

Ultrasensitive Detection of Chimerism by Single-Molecule Molecular Inversion Probe Capture and High-Throughput Sequencing of Copy Number Deletion Polymorphisms

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et al. 2018

Clinical Chemistry

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BACKGROUND: Genomic chimerism, the co-occurrence of cells from different genetic origins, provides important diagnostic information in diverse clinical contexts, including graft injury detection and longitudinal surveillance of hematopoietic stem cell transplantation patients, but existing assays are limiting. Here we applied single-molecule molecular inversion probes (smMIPs), a high throughput sequencing technology combining multiplexed target capture with read quantitation mediated by unique molecular identifiers, to detect chimerism based on the presence or absence of polymorphic genomic loci. METHODS: We designed a 159 smMIP panel targeting 40 autosomal regions of frequent homozygous deletion across human populations and two sex-linked loci. We developed methods for detecting and quantitating loci absent from one cell population but present in another, which could be used to sensitively identify chimeric cell populations. RESULTS: Unrelated individuals and first-degree relatives were highly polymorphic across the loci examined. Using synthetic DNA mixtures, limits of detection of at least 1 in 10,000 chimeric cells was demonstrated without prior knowledge of genotypes, and mixtures of up to four separate donors could be deconvoluted. Quantitative linearity over four orders of magnitude and false positive rates below 1 in 85,000 events were achieved. 11 of 11 post-transplant clinical specimens from patients with hematological malignancies testing positive for residual cancer by conventional methods had detectable chimeric populations by smMIP, while 11 of 11 specimens testing negative by conventional methods were low-positive for chimerism by smMIP. CONCLUSIONS: smMIPs are scalable to high sensitivity and large numbers of informative markers, enabling ultrasensitive chimerism detection for many clinical purposes.

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BRCA Testing by Single-Molecule Molecular Inversion Probes

Cited by 53 publications

References 38 publications

A molecular inversion probe-based next-generation sequencing panel to detect germline mutations in Chinese early-onset colorectal cancer patients

A molecular inversion probe-based next-generation sequencing panel to detect germline mutations in Chinese early-onset colorectal cancer patients

Accurate Pan-Cancer Molecular Diagnosis of Microsatellite Instability by Single-Molecule Molecular Inversion Probe Capture and High-Throughput Sequencing

Ultrasensitive Detection of Chimerism by Single-Molecule Molecular Inversion Probe Capture and High-Throughput Sequencing of Copy Number Deletion Polymorphisms

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