Massively parallel interrogation and mining of natively paired human TCRαβ repertoires

Spindler, Matthew J.; Nelson, Ayla L.; Wagner, Ellen K.; Oppermans, Natasha; Bridgeman, John S.; Heather, James; Adler, Adam S.; Asensio, Michael A.; Edgar, Robert C.; Lim, Yoong Wearn; Meyer, Everett; Hawkins, Robert E.; Cobbold, Mark; Johnson, D. S.

doi:10.1038/s41587-020-0438-y

Cited by 43 publications

(53 citation statements)

References 66 publications

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“…However, especially the identification and in‐depth characterisation of suitable TCRs for clinical use was so far a tedious process and only a handful of clinical studies with TCR re‐directed T‐cell products are reported 5–9 . Today, because of continuous improvements in the field of next‐generation sequencing, high‐throughput identification of full αβ TCR sequences is no longer a bottleneck 10–13 . Moreover, advanced bioinformatical analytical tools are developed to gain deep insight into such large TCR repertoire data and to predict antigen‐HLA specificity from raw TCR sequences 14,15 .…”

Section: Introductionmentioning

confidence: 99%

A T‐cell reporter platform for high‐throughput and reliable investigation of TCR function and biology

et al. 2020

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Objective Transgenic re‐expression enables unbiased investigation of T‐cell receptor (TCR)‐intrinsic characteristics detached from its original cellular context. Recent advancements in TCR repertoire sequencing and development of protocols for direct cloning of full TCRαβ constructs now facilitate large‐scale transgenic TCR re‐expression. Together, this offers unprecedented opportunities for the screening of TCRs for basic research as well as clinical use. However, the functional characterisation of re‐expressed TCRs is still a complicated and laborious matter. Here, we propose a Jurkat‐based triple parameter TCR signalling reporter endogenous TCR knockout cellular platform (TPRKO) that offers an unbiased, easy read‐out of TCR functionality and facilitates high‐throughput screening approaches. Methods As a proof‐of‐concept, we transgenically re‐expressed 59 human cytomegalovirus‐specific TCRs and systematically investigated and compared TCR function in TPRKO cells versus primary human T cells. Results We demonstrate that the TPRKO cell line facilitates antigen‐HLA specificity screening via sensitive peptide‐MHC‐multimer staining, which was highly comparable to primary T cells. Also, TCR functional avidity in TPRKO cells was strongly correlating to primary T cells, especially in the absence of CD8αβ co‐receptor. Conclusion Overall, our data show that the TPRKO cell lines can serve as a surrogate of primary human T cells for standardised and high‐throughput investigation of TCR biology.

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Section: Introductionmentioning

confidence: 99%

A T‐cell reporter platform for high‐throughput and reliable investigation of TCR function and biology

et al. 2020

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“…First, input linked scFv repertoires typically comprised approximately 2- to 4-fold more antibodies than the final CHO cell banks. Our prior work to clone T cell receptor (TCR) repertoires into Jurkat cells was much more efficient, 43 but that work used lentivirus rather than Flp-In site directed integration, and therefore many Jurkat clones expressed multiple TCRs. In the future, we will work to improve the efficiency of CHO engineering.…”

Section: Discussionmentioning

confidence: 99%

Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

Keating

Mizrahi

Ms³

et al. 2020

Preprint

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Plasma-derived polyclonal antibodies are polyvalent drugs used for many important clinical indications that require modulation of multiple drug targets simultaneously, including emerging infectious disease and transplantation. However, plasma-derived drugs suffer many problems, including low potency, impurities, constraints on supply, and batch-to-batch variation. In this study, we demonstrated proofs-of-concept for a technology that uses microfluidics and molecular genomics to capture diverse mammalian antibody repertoires as multivalent recombinant drugs. These “recombinant hyperimmune” drugs comprised thousands to tens of thousands of antibodies and were derived from convalescent human donors, or vaccinated human donors or immunized mice. Here we used our technology to build a highly potent recombinant hyperimmune for Severe Acute Respiratory Syndrome Coronavirus-2 (SARS CoV-2) in less than three months. We also validated a recombinant hyperimmune for Zika virus disease that abrogates antibody-dependent enhancement (ADE) through Fc engineering. For patients with primary immune deficiency (PID), we built high potency polyvalent recombinant hyperimmunes against pathogens that commonly cause serious lung infections. Finally, to address the limitations of rabbit-derived anti-thymocyte globulin (ATG), we generated a recombinant human version and demonstrated in vivo function against graft-versus-host disease (GVHD). Recombinant hyperimmunes are a novel class of drugs that could be used to target a wide variety of other clinical applications, including cancer and autoimmunity.

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“…Fast and inexpensive functional screening of transiently expressed TCRs in CD8+ and CD4+ Jurkats has been established, but the transient nature prevented further engineering approaches and selections. Current research in the field has been overwhelmingly focused on affinity-based selections of TCRs ( Kessels et al., 2001 ; Chervin et al., 2008 ; Malecek et al., 2013 ; Schmitt et al., 2017 ; Wagner et al., 2019 ; Spindler et al., 2020 ). As highlighted in previous sections, affinity maturation of scTCR-pMHC pairs has been done by phage and yeast display screening.…”

Section: Introductionmentioning

confidence: 99%

“…A number of reports demonstrate the use of mammalian cells for TCR screening and engineering, most of which involved viral transformations and affinity-based selections ( Kessels et al., 2001 ; Chervin et al., 2008 ; Malecek et al., 2013 ; Schmitt et al., 2017 ; Wagner et al., 2019 ; Spindler et al., 2020 ). In order to overcome the challenges related to TCR affinity and cross-reactivity, it is necessary to engineer TCR specificity not only on the basis of binding and function ( Rosskopf et al., 2018 ), but also while detecting cross-reactivity.…”

Section: Introductionmentioning

confidence: 99%

Immune Literacy: Reading, Writing, and Editing Adaptive Immunity

Csepregi¹,

Ehling²,

Wagner³

et al. 2020

iScience

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Advances in reading, writing, and editing DNA are providing unprecedented insights into the complexity of immunological systems. This combination of systems and synthetic biology methods is enabling the quantitative and precise understanding of molecular recognition in adaptive immunity, thus providing a framework for reprogramming immune responses for translational medicine. In this review, we will highlight state-of-the-art methods such as immune repertoire sequencing, immunoinformatics, and immunogenomic engineering and their application toward adaptive immunity. We showcase novel and interdisciplinary approaches that have the promise of transforming the design and breadth of molecular and cellular immunotherapies.

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Massively parallel interrogation and mining of natively paired human TCRαβ repertoires

Cited by 43 publications

References 66 publications

A T‐cell reporter platform for high‐throughput and reliable investigation of TCR function and biology

A T‐cell reporter platform for high‐throughput and reliable investigation of TCR function and biology

Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

Immune Literacy: Reading, Writing, and Editing Adaptive Immunity

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