Targeted T cell receptor gene editing provides predictable T cell product function for immunotherapy

Müller, Thomas; Jarosch, Sebastian; Hammel, Monika; Leube, Justin; Grassmann, Simon; Bernard, Bettina; Effenberger, Manuel; Andrä, Immanuel; Chaudhry, M. Zeeshan; Käuferle, Theresa; Malo, Antje; Čičin‐Šain, Luka; Schmitz, Peter; Feuchtinger, Tobias; Protzer, Ulrike; Schumann, Kathrin; Neuenhahn, Michael; Schober, Kilian; Busch, Dirk H.

doi:10.1016/j.xcrm.2021.100374

Cited by 32 publications

(30 citation statements)

References 72 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This technology places the transgenic TCR into the endogenous gene locus, resulting in physiological transgene expression, and simultaneously completely removes the endogenous TCR, leading to unbiased and complete reexpression of transgenic TCRs. Because the recipient T cells for engineering are the same for all TCRs, this approach allows for a systematic comparison of TCR functionality, irrespective of the phenotype of the original cells from which the TCR was isolated ( 62 , 63 ). For reexpression, we chose the eight most abundant TCRs across all samples [human TCRs (hTCRs) E14, E2, E4, E5, E23, E49, E13, and E74].…”

Section: Resultsmentioning

confidence: 99%

Signatures of recent activation identify a circulating T cell compartment containing tumor-specific antigen receptors with high avidity

et al. 2022

Self Cite

View full text Add to dashboard Cite

T cell receptor (TCR) avidity is assumed to be a major determinant of the spatiotemporal fate and protective capacity of tumor-specific T cells. However, monitoring polyclonal T cell responses with known TCR avidities in vivo over space and time remains challenging. Here, we investigated the fate and functionality of tumor neoantigen–specific T cells with TCRs of distinct avidities in a well-established, reductionist preclinical tumor model and human patients with melanoma. To this end, we used polyclonal T cell transfers with in-depth characterized TCRs together with flow cytometric phenotyping in mice inoculated with MC38 OVA tumors. Transfer of T cells from retrogenic mice harboring TCRs with high avidity resulted in best tumor protection. Unexpectedly, we found that both high- and low-avidity T cells are similarly abundant within the tumor and adopt concordant phenotypic signs of exhaustion. Outside the tumor, high-avidity TCR T cells were not generally overrepresented but, instead, selectively enriched in T cell populations with intermediate PD-1 protein expression. Single-cell sequencing of neoantigen-specific T cells from two patients with melanoma—combined with transgenic reexpression of identified TCRs by CRISPR-Cas9–mediated orthotopic TCR replacement—revealed high-functionality TCRs to be enriched in T cells with RNA signatures of recent activation. Furthermore, of 130 surface protein candidates, PD-1 surface expression was most consistently enriched in functional TCRs. Together, our findings show that tumor-reactive TCRs with high protective capacity circulating in peripheral blood are characterized by a signature of recent activation.

show abstract

Section: Resultsmentioning

confidence: 99%

Signatures of recent activation identify a circulating T cell compartment containing tumor-specific antigen receptors with high avidity

et al. 2022

Self Cite

View full text Add to dashboard Cite

show abstract

“…TI of the EF1α construct generated more potent T cell activation than the construct driven by the endogenous TCRα promoter while also resulting in less exhausted T cells. Recently, Müller and colleagues ( 65 ) showed that the orthotopic integration of viral specific TCR genes leads to homogeneous TCR expression and more predictable in vivo activity of engineered cells, when compared to retrovirally transduced, edited T cells. Our data suggest that optimal activation of the TCR is critical and dependent on genomic context as well as the right promoter strength.…”

Section: Discussionmentioning

confidence: 99%

CRISPR-based gene disruption and integration of high-avidity, WT1-specific T cell receptors improve antitumor T cell function

et al. 2022

View full text Add to dashboard Cite

T cell receptor (TCR)–based therapy has the potential to induce durable clinical responses in patients with cancer by targeting intracellular tumor antigens with high sensitivity and by promoting T cell survival. However, the need for TCRs specific for shared oncogenic antigens and the need for manufacturing protocols able to redirect T cell specificity while preserving T cell fitness remain limiting factors. By longitudinal monitoring of T cell functionality and dynamics in 15 healthy donors, we isolated 19 TCRs specific for Wilms’ tumor antigen 1 (WT1), which is overexpressed by several tumor types. TCRs recognized several peptides restricted by common human leukocyte antigen (HLA) alleles and displayed a wide range of functional avidities. We selected five high-avidity HLA-A*02:01–restricted TCRs, three that were specific to the less explored immunodominant WT1 37–45 and two that were specific to the noncanonical WT1 −78–64 epitopes, both naturally processed by primary acute myeloid leukemia (AML) blasts. With CRISPR-Cas9 genome editing tools, we combined TCR-targeted integration into the TCR α constant ( TRAC ) locus with TCR β constant ( TRBC ) knockout, thus avoiding TCRαβ mispairing and maximizing TCR expression and function. The engineered lymphocytes were enriched in memory stem T cells. A unique WT1 37–45 -specific TCR showed antigen-specific responses and efficiently killed AML blasts, acute lymphoblastic leukemia blasts, and glioblastoma cells in vitro and in vivo in the absence of off-tumor toxicity. T cells engineered to express this receptor are being advanced into clinical development for AML immunotherapy and represent a candidate therapy for other WT1-expressing tumors.

show abstract

“…Other sources of T cell heterogeneity can result when T cells are genetically modified, such as by CRISPR/Cas9 genome editing, which can produce mixed edited and unedited cells ( Rupp et al, 2017 ). Genome editing procedures may also be used to reduce heterogeneity in modified T cells ( Muller et al, 2021 ). An integrated readout of T cell activation combined with cancer cell death could be used to determine whether phenotypic differences that influence T cell activation result in altered T cell-mediated cytotoxicity.…”

Section: Discussionmentioning

confidence: 99%

Combined analysis of T cell activation and T cell-mediated cytotoxicity by imaging cytometry

Chanda

Shudde

Piper

et al. 2022

Journal of Immunological Methods

View full text Add to dashboard Cite

Targeted T cell receptor gene editing provides predictable T cell product function for immunotherapy

Cited by 32 publications

References 72 publications

Signatures of recent activation identify a circulating T cell compartment containing tumor-specific antigen receptors with high avidity

Signatures of recent activation identify a circulating T cell compartment containing tumor-specific antigen receptors with high avidity

CRISPR-based gene disruption and integration of high-avidity, WT1-specific T cell receptors improve antitumor T cell function

Combined analysis of T cell activation and T cell-mediated cytotoxicity by imaging cytometry

Contact Info

Product

Resources

About