TCR Repertoire Analysis by Next Generation Sequencing Allows Complex Differential Diagnosis of T Cell–Related Pathology

Dziubianau, Mikalai; Hecht, Jochen; Kuchenbecker, Léon; Sattler, Arne; Stervbo, Ulrik; Rödelsperger, Christian; Nickel, Peter; Neumann, Avidan U.; Robinson, Peter N.; Mundlos, Stefan; Volk, Hans‐Dieter; Thiel, Andreas; Reinke, Petra; Babel, Nina

doi:10.1111/ajt.12431

Cited by 127 publications

(113 citation statements)

References 57 publications

Supporting

Mentioning

108

Contrasting

Order By: Relevance

“…In addition to changes in the expression of cosignaling receptors on the surface of T cells, antigen-specific stimulation typically results in clonal expansion. TCR sequence immunoprofiling can be used to monitor T cell responses to a given immune challenge even without a priori knowledge of the specific epitope targeted, through determination of the abundance of specific clonotypes (29,30). However, there is limited knowledge regarding the TCR repertoire and the frequency of tumor-reactive clonotypes infiltrating human tumors.…”

Section: Introductionmentioning

confidence: 99%

PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors

Gros¹,

Robbins²,

Yao³

et al. 2014

J. Clin. Invest.

927

816

View full text Add to dashboard Cite

Adoptive transfer of tumor-infiltrating lymphocytes (TILs) can mediate regression of metastatic melanoma; however, TILs are a heterogeneous population, and there are no effective markers to specifically identify and select the repertoire of tumor-reactive and mutation-specific CD8 + lymphocytes. The lack of biomarkers limits the ability to study these cells and develop strategies to enhance clinical efficacy and extend this therapy to other malignancies. Here, we evaluated unique phenotypic traits of CD8 + TILs and TCR β chain (TCRβ) clonotypic frequency in melanoma tumors to identify patient-specific repertoires of tumor-reactive CD8 + lymphocytes. In all 6 tumors studied, expression of the inhibitory receptors programmed cell death 1 (PD-1; also known as CD279), lymphocyte-activation gene 3 (LAG-3; also known as CD223), and T cell immunoglobulin and mucin domain 3 (TIM-3) on CD8 + TILs identified the autologous tumor-reactive repertoire, including mutated neoantigen-specific CD8 + lymphocytes, whereas only a fraction of the tumor-reactive population expressed the costimulatory receptor 4-1BB (also known as CD137). TCRβ deep sequencing revealed oligoclonal expansion of specific TCRβ clonotypes in CD8 + PD-1 + compared with CD8 + PD-1 -TIL populations. Furthermore, the most highly expanded TCRβ clonotypes in the CD8 + and the CD8 + PD-1 + populations recognized the autologous tumor and included clonotypes targeting mutated antigens. Thus, in addition to the well-documented negative regulatory role of PD-1 in T cells, our findings demonstrate that PD-1 expression on CD8 + TILs also accurately identifies the repertoire of clonally expanded tumor-reactive cells and reveal a dual importance of PD-1 expression in the tumor microenvironment.

show abstract

Section: Introductionmentioning

confidence: 99%

PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors

Gros¹,

Robbins²,

Yao³

et al. 2014

J. Clin. Invest.

927

816

View full text Add to dashboard Cite

show abstract

“…Novel emerging technologies, such as next-generation sequencing (NGS), have recently begun to be used to assess TCR repertoires (12,13) and may be useful in understanding immunotherapy-induced immune system changes (14,15). The power of NGS is derived from its ability to examine unique sequences with inferred antigen specificity, or clonality, which mirrors the TCR repertoire, to enhance the speed of clonotyping analysis and to detect low-abundance clones (16).…”

Section: Introductionmentioning

confidence: 99%

Clonotypic Diversification of Intratumoral T Cells Following Sipuleucel-T Treatment in Prostate Cancer Subjects

et al. 2016

View full text Add to dashboard Cite

Sipuleucel-T is an autologous cellular therapy for asymptomatic, or minimally symptomatic, metastatic castrate-resistant prostate cancer, designed to stimulate an immune response against prostate cancer. In a recent clinical trial (NCT00715104), we found that neoadjuvant sipuleucel-T increased the number of activated T cells within the tumor microenvironment. The current analysis examined whether sipuleucel-T altered adaptive T-cell responses by expanding pre-existing T cells or by recruiting new T cells to prostate tissue.

show abstract

“…It is estimated that the total number of T cells in the human is approximately 10 12 , and the TRB repertoire consists of 10 6 different clones. 39 Thus, the higher variability among the sequencing results of different biological replicate libraries is likely a reflection of the enormous number of low-frequency clonotypes.…”

Section: Discussionmentioning

confidence: 99%

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

You

Zheng

et al. 2017

The Journal of Molecular Diagnostics

View full text Add to dashboard Cite

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 primerebased 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 b (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 <10À3 . A single library could identify >93% of the clones with frequency !10 À3 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 À3 and provides a methodologic framework for applying NGS immune repertoire methods to clinical testing applications beyond MRD. (J Mol Diagn 2017, 19: 72e83; http://dx.doi.org/10.1016/j.jmoldx.2016 Study and analysis of T-cell receptor (TCR) repertoires are essential for better understanding the development and reconstitution of immune systems, for providing insights into the pathogenesis of immune disorders, and for diagnosing and assessing therapeutic interventions for diseases driven by T cells. Next-generation sequencing (NGS) technology has become a standard method for assessing minimal residual disease (MRD) in patients with lymphoid malignancies. From a technical standpoint, MRD is a relatively simple application of NGS technology, essentially looking for known malignant or related somatically mutated clones; by combining replicate libraries, a sensitivity of 1 in 10 À5 has been demonstrated for MRD. Beyond detecting MRD, NGS is also an innovative and promising approach to assess clonal populations of T cells in healthy people during immune system development, patients with suspected malignancy, response to targeted therapies, immune reconstitution in patients undergoing hematopoietic cell transplant, and autoimmune diseases. 7,9e25 However, assessment of the T-cell repertoire in individuals is a far more complex task than assessment of MRD. In consideration of extending NGS methods beyond MRD applications in clinical laboratories, we examined the opportunity and technical limitations of the widely used BIOMED-2 primer sets and the MiSeq system (Illumina Inc., San Diego, CA) as an NGS application to assess the TCR repertoire. We evaluated replicate testing of individual libraries (th...

show abstract

TCR Repertoire Analysis by Next Generation Sequencing Allows Complex Differential Diagnosis of T Cell–Related Pathology

Cited by 127 publications

References 57 publications

PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors

PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors

Clonotypic Diversification of Intratumoral T Cells Following Sipuleucel-T Treatment in Prostate Cancer Subjects

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

Contact Info

Product

Resources

About