scholarly journals Frequent Aneuploidy in Primary Human T Cells Following CRISPR-Cas9 cleavage

2021 ◽  
Author(s):  
Alessio D Nahmad ◽  
Eli Reuveni ◽  
Ella Goldschmidt ◽  
Tamar Tenne ◽  
Meytal Liberman ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
Temporal Dynamics ◽  
Cell Receptor ◽  
T Cell Subsets ◽  
Chromosome 14 ◽  
T Cell Therapy ◽  
Guide Rna ◽  
Human T Cells

Multiple ongoing clinical trials use site-specific nucleases to disrupt T cell receptor (TCR) genes in order to allow for allogeneic T cell therapy. In particular, the first U.S. clinical trial using CRISPR-Cas9 entailed the targeted disruption of the TCR chains and programmed cell death protein 1 (PDCD1) in T cells of refractory cancer patients. Here, we used the same guide RNA sequences and applied single-cell RNA sequencing (scRNAseq) to more than 7000 primary human T cells, transfected with CRISPR-Cas9. Four days post-transfection, we found a loss of chromosome 14, harboring the TCRα locus, in up to 9% of the cells, and a chromosome 14 gain in up to 1.4% of the cells. We further identified truncations of chromosome 7, harboring the TCRβ locus, in 9.9% of the cells. Loss of heterozygosity (LOH) was further validated using fluorescence in situ hybridization (FISH) and the temporal dynamics of cleavage and incomplete repair were monitored using digital droplet PCR (ddPCR). Aneuploidy was found among all T cell subsets and was associated with transcriptional signatures of reduced proliferation and metabolism as well as with induced p53 activation and cell death. We conclude that aneuploidy and chromosomal truncations are frequent outcomes of CRISPR-Cas9 cleavage in clinical protocols. Monitoring and minimizing these aberrant products is crucial for future applications of genome editing in T cell engineering and beyond.

scholarly journals T Cell Receptor-induced Activation and Apoptosis In Cycling Human T Cells Occur throughout the Cell Cycle

1999 ◽  
Vol 10 (12) ◽  
pp. 4441-4450 ◽  
Author(s):  
Michael Karas ◽  
Tal Z. Zaks ◽  
Liu JL ◽  
Derek LeRoith
Keyword(s):  
Cell Cycle ◽  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
T Cell Receptor ◽  
Fas Ligand ◽  
Cell Receptor ◽  
Receptor Activation ◽  
Activation Induced Cell Death ◽  
Human T Cells

Previous studies have found conflicting associations between susceptibility to activation-induced cell death and the cell cycle in T cells. However, most of the studies used potentially toxic pharmacological agents for cell cycle synchronization. A panel of human melanoma tumor-reactive T cell lines, a CD8+ HER-2/neu-reactive T cell clone, and the leukemic T cell line Jurkat were separated by centrifugal elutriation. Fractions enriched for the G0–G1, S, and G2–M phases of the cell cycle were assayed for T cell receptor-mediated activation as measured by intracellular Ca2+flux, cytolytic recognition of tumor targets, and induction of Fas ligand mRNA. Susceptibility to apoptosis induced by recombinant Fas ligand and activation-induced cell death were also studied. None of the parameters studied was specific to a certain phase of the cell cycle, leading us to conclude that in nontransformed human T cells, both activation and apoptosis through T cell receptor activation can occur in all phases of the cell cycle.

scholarly journals Characterization of human T cells reactive with the Mycoplasma arthritidis-derived superantigen (MAM): generation of a monoclonal antibody against V beta 17, the T cell receptor gene product expressed by a large fraction of MAM-reactive human T cells.

1991 ◽  
Vol 174 (4) ◽  
pp. 891-900 ◽  
Author(s):  
S M Friedman ◽  
M K Crow ◽  
J R Tumang ◽  
M Tumang ◽  
Y Q Xu ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Cell Activity ◽  
Cell Receptor ◽  
Cytotoxic T Cell ◽  
Mycoplasma Arthritidis ◽  
Human T Cells

While all known microbial superantigens are mitogenic for human peripheral blood lymphocytes (PBL), the functional response induced by Mycoplasma arthritidis-derived superantigen (MAM) is unique in that MAM stimulation of PBL consistently results in T cell-dependent B cell activation characterized by polyclonal IgM and IgG production. These immunostimulatory effects of MAM on the humoral arm of the human immune system warranted a more precise characterization of MAM-reactive human T cells. Using an uncloned MAM reactive human T cell line as immunogen, we have generated a monoclonal antibody (mAb) (termed C1) specific for the T cell receptor V beta gene expressed by the major fraction of MAM-reactive human T cells, V beta 17. In addition, a V beta 17- MAM-reactive T cell population exists, assessed by MAM, induced T cell proliferation and cytotoxic T cell activity. mAb C1 will be useful in characterizing the functional properties of V beta 17+ T cells and their potential role in autoimmune disease.

scholarly journals Structure and specificity of T cell receptor gamma/delta on major histocompatibility complex antigen-specific CD3+, CD4-, CD8- T lymphocytes.

1988 ◽  
Vol 168 (5) ◽  
pp. 1899-1916 ◽  
Author(s):  
J A Bluestone ◽  
R Q Cron ◽  
M Cotterman ◽  
B A Houlden ◽  
L A Matis
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Gene Segment ◽  
Cell Receptor ◽  
Gamma Delta ◽  
Human T Cells ◽  
T Cell Lines ◽  
Gamma Delta T Cell ◽  
Delta Cells

Analyses of TCR-bearing murine and human T cells have defined a unique subpopulation of T cells that express the TCR-gamma/delta proteins. The specificity of TCR-gamma/delta T cells and their role in the immune response have not yet been elucidated. Here we examine alloreactive TCR-gamma/delta T cell lines and clones that recognize MHC-encoded antigens. A BALB/c nu/nu (H-2d)-derived H-2k specific T cell line and derived clones were both cytolytic and released lymphokines after recognition of a non-classical H-2 antigen encoded in the TL region of the MHC. These cells expressed the V gamma 2/C gamma 1 protein in association with a TCR-delta gene product encoded by a Va gene segment rearranged to two D delta and one J delta variable elements. A second MHC-specific B10 nu/nu (H-2b) TCR-gamma/delta T cell line appeared to recognize a classical H-2D-encoded MHC molecule and expressed a distinct V gamma/C gamma 4-encoded protein. These data suggest that many TCR-gamma/delta-expressing T cells may recognize MHC-linked antigens encoded within distinct subregions of the MHC. The role of MHC-specific TCR-gamma/delta cells in immune responses and their immunological significance are discussed.

scholarly journals Fas ligand mediates activation-induced cell death in human T lymphocytes.

1995 ◽  
Vol 181 (1) ◽  
pp. 71-77 ◽  
Author(s):  
M R Alderson ◽  
T W Tough ◽  
T Davis-Smith ◽  
S Braddy ◽  
B Falk ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
T Cell ◽  
Fas Ligand ◽  
Significant Proportion ◽  
Cell Receptor ◽  
Target Cells ◽  
Activated T Cells ◽  
Activation Induced Cell Death ◽  
Fas L

A significant proportion of previously activated human T cells undergo apoptosis when triggered through the CD3/T cell receptor complex, a process termed activation-induced cell death (AICD). Ligation of Fas on activated T cells by either Fas antibodies or recombinant human Fas-ligand (Fas-L) also results in cytolysis. We demonstrate that these two pathways of apoptosis are causally related. Stimulation of previously activated T cells resulted in the expression of Fas-L mRNA and lysis of Fas-positive target cells. Fas-L antagonists inhibited AICD of T cell clones and staphylococcus enterotoxin B (SEB)-specific T cell lines. The data indicate AICD in previously stimulated T cells is mediated by Fas/Fas-L interactions.

Susceptibility of human T cells, T-cell subsets, and B cells to cryopreservation

Cryobiology ◽  
1986 ◽  
Vol 23 (3) ◽  
pp. 199-208 ◽  
Author(s):  
M. Venkataraman ◽  
M.P. Westerman
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
T Cell Subsets ◽  
Human T Cells

Expression of T-cell receptor α and β variable genes in normal and malignant human T cells

1993 ◽  
Vol 84 (1) ◽  
pp. 39-48 ◽  
Author(s):  
Huaizhong Hu ◽  
Màrio Rui Queirò ◽  
Marcel G. J. Tilanus ◽  
Roel A. Weger ◽  
Henk-Jan Schuurman
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Cell Receptor ◽  
Human T Cells ◽  
Variable Genes

scholarly journals Oligoclonal T Cells Transiently Expand and Express Tim-3 and PD-1 Following Anti-CD19 CAR T Cell Therapy: A Case Report

2018 ◽  
Vol 19 (12) ◽  
pp. 4118 ◽  
Author(s):  
Christopher Funk ◽  
Christopher Petersen ◽  
Neera Jagirdar ◽  
Sruthi Ravindranathan ◽  
David Jaye ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Hematologic Malignancy ◽  
Drug Application ◽  
B Cell Lymphoma ◽  
T Cell Subsets ◽  
Phenotypic Analysis ◽  
T Cell Therapy ◽  
Car T Cell ◽  
Car T

Clinical trials of chimeric antigen receptor (CAR) T cells in hematologic malignancy associate remissions with two profiles of CAR T cell proliferation kinetics, which differ based upon costimulatory domain. Additional T cell intrinsic factors that influence or predict clinical response remain unclear. To address this gap, we report the case of a 68-year-old woman with refractory/relapsed diffuse large B cell lymphoma (DLBCL), treated with tisagenlecleucel (anti-CD19), with a CD137 costimulatory domain (4-1BB) on an investigational new drug application (#16944). For two months post-infusion, the patient experienced dramatic regression of subcutaneous nodules of DLBCL. Unfortunately, her CAR T exhibited kinetics unassociated with remission, and she died of DLBCL-related sequelae. Serial phenotypic analysis of peripheral blood alongside sequencing of the β-peptide variable region of the T cell receptor (TCRβ) revealed distinct waves of oligoclonal T cell expansion with dynamic expression of immune checkpoint molecules. One week prior to CAR T cell contraction, T cell immunoglobulin mucin domain 3 (Tim-3) and programmed cell death protein 1 (PD-1) exhibited peak expressions on both the CD8 T cell (Tim-3 ≈ 50%; PD-1 ≈ 17%) and CAR T cell subsets (Tim-3 ≈ 78%; PD-1 ≈ 40%). These correlative observations draw attention to Tim-3 and PD-1 signaling pathways in context of CAR T cell exhaustion.

scholarly journals Human CD4+ T cells specific for dominant epitopes of SARS-CoV-2 Spike and Nucleocapsid proteins with therapeutic potential

2021 ◽  
Author(s):  
Johan Verhagen ◽  
Edith Van der Meijden ◽  
Vanessa Lang ◽  
Andreas Kremer ◽  
Simon Völkl ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Cd4 T Cells ◽  
Cellular Therapy ◽  
Therapeutic Potential ◽  
Cell Receptor ◽  
Haematopoietic Stem Cell ◽  
T Cell Therapy ◽  
Immunological Protection

Since December 2019, Coronavirus disease-19 (COVID-19) has spread rapidly across the world, leading to a global effort to develop vaccines and treatments. Despite extensive progress, there remains a need for treatments to bolster the immune responses in infected immunocompromised individuals, such as cancer patients who recently underwent a haematopoietic stem cell transplantation. Immunological protection against COVID-19 is mediated by both short-lived neutralising antibodies and long-lasting virus-reactive T cells. Therefore, we propose that T cell therapy may augment efficacy of current treatments. For the greatest efficacy with minimal adverse effects, it is important that any cellular therapy is designed to be as specific and directed as possible. Here, we identify T cells from COVID-19 patients with a potentially protective response to two major antigens of the SARS-CoV-2 virus, Spike and Nucleocapsid protein. By generating clones of highly virus-reactive CD4+ T cells, we were able to confirm a set of 9 immunodominant epitopes and characterise T cell responses against these. Accordingly, the sensitivity of T cell clones for their specific epitope, as well as the extent and focus of their cytokine response was examined. Moreover, by using an advanced T cell receptor (TCR) sequencing approach, we determined the paired TCR sequences of clones of interest. While these data on a limited population require further expansion for universal application, the results presented here form a crucial first step towards TCR-transgenic CD4+ T cell therapy of COVID-19.

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