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Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use.

ABSTRACT: Next-generation sequencing (NGS) of immune receptors has become a standard tool to assess minimal residual disease (MRD) in patients treated for lymphoid malignancy, and it is being used to study the T-cell repertoire in many clinical settings. To better understanding the potential clinical utility and limitations of this application outside of MRD, we developed a BIOMED-2 primer-based NGS method and characterized its performance in controls and patients with graft-versus-host disease (GVHD) after allogeneic hematopoietic transplant. For controls and patients with GVHD, replicate sequencing of the same T-cell receptor β (TRB) libraries was highly reproducible. Higher variability was observed in sequencing of different TRB libraries made from the same DNA stock. Variability was increased in patients with GVHD compared with controls; patients with GVHD also had lower diversity than controls. In the T-cell repertoire of a healthy person, approximately 99.6% of the CDR3 clones were in low abundance, with frequency <10^-3. A single library could identify >93% of the clones with frequency ≥10^-3 in the repertoire. Sequencing in duplicate increased the average detection rate to >97%. This work demonstrates that NGS reliably and robustly characterizes TRB populations in healthy individuals and patients with GVHD with frequency ≥10^-3 and provides a methodologic framework for applying NGS immune repertoire methods to clinical testing applications beyond MRD.

SUBMITTER: Xu L

PROVIDER: S-EPMC5225307 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use.

Xu Liwen L You Xiaoqing X Zheng PingPing P Zhang Bing M BM Gupta Puja K PK Lavori Philip P Meyer Everett E Zehnder James L JL

The Journal of molecular diagnostics : JMD 20161101 1

Next-generation sequencing (NGS) of immune receptors has become a standard tool to assess minimal residual disease (MRD) in patients treated for lymphoid malignancy, and it is being used to study the T-cell repertoire in many clinical settings. To better understanding the potential clinical utility and limitations of this application outside of MRD, we developed a BIOMED-2 primer-based NGS method and characterized its performance in controls and patients with graft-versus-host disease (GVHD) aft ...[more]

PMID: 27815002

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Project description:ObjectiveTimely and precise etiology diagnosis is crucial for optimized medication regimens and better prognosis in central nervous system infections (CNS infections). We aimed to analyze the impact of mNGS tests on the management of patients with CNS infections.MethodsWe conducted a single-center retrospective cohort study to analyze the value of mNGS in clinical applications. Three hundred sixty-nine patients with a CNS infection diagnosis were enrolled, and their clinical data were collected. CDI and DDI were defined in our study to describe the intensity of drug use in different groups. We used LOH and mRS to evaluate if the application of mNGS can benefit CNS infected patients.ResultsmNGS reported a 91.67% sensitivity in culture-positive patients and an 88.24% specificity compared with the final diagnoses. Patients who participated with the mNGS test had less drug use, both total (58.77 vs. 81.18) and daily (22.6 vs. 28.12, P < 0.1, McNemar) intensity of drug use, and length of hospitalization (23.14 vs. 24.29). Patients with a consciousness grading 1 and 3 had a decrease in CDI (Grade 1, 86.49 vs. 173.37; Grade 3, 48.18 vs. 68.21), DDI (Grade 1, 1.52 vs. 2.72; Grade 3, 2.3 vs. 2.45), and LOH (Grade 1, 32 vs. 40; Grade 3, 21 vs. 23) with the application of mNGS. Patients infected with bacteria in the CNS had a reduced CDI, DDI, and LOH in the mNGS group. This was compared with the TraE group that had 49% of patients altered medication plans, and 24.7% of patients reduced drug intensity four days after mNGS reports. This was because of the reduction of drug types.ConclusionmNGS showed its high sensitivity and specificity characteristics. mNGS may assist clinicians with more rational medication regimens and reduce the drug intensity for patients. The primary way of achieving this is to reduce the variety of drugs, especially for severe patients and bacterial infections. mNGS has the ability of improving the prognosis of CNS infected patients.

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Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use.

Publications

Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use.

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