Generation of human parallel chimeric antigen receptor (pCAR) T cells to achieve synergistic T cell co-stimulation

Larcombe-Young, Daniel; Whilding, Lynsey M.; Davies, Daniel; Draper, Benjamin; Bechman, Natasha; Maher, John

doi:10.1016/j.xpro.2022.101414

Cited by 3 publications

(4 citation statements)

References 17 publications

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“…Vector production was performed as described [ 21 ]. In brief, 3 × 10 6 HEK293T cells were plated into 10 cm 2 tissue culture-treated plates.…”

Section: Methodsmentioning

confidence: 99%

“…Peripheral blood mononuclear cells (PBMC) were isolated using ficoll-based density gradient centrifugation as described [ 21 ]. Isolated cells were cultured in RPMI media supplemented with 2 mM L-glutamine and 5% human AB serum.…”

Section: Methodsmentioning

confidence: 99%

“…Co-cultivation assays between T-cells and target cells were established at effector-to-target (E:T) ratios as specified in individual experiments. Target cell destruction was quantified by in vitro MTT or luciferase assays, as described in [ 21 ]. Using either assay, the percentage of target cell viability was calculated using the following formula: (absorbance/luminescence of tumour cells cultured with T cells divided by absorbance/luminescence of tumour cells alone) × 100%.…”

Section: Methodsmentioning

confidence: 99%

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Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

Davies,

Pugliese,

Parente Pereira

et al. 2024

Biology

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γδ T-cells provide immune surveillance against cancer, straddling both innate and adaptive immunity. G115 is a clonal γδ T-cell receptor (TCR) of the Vγ9Vδ2 subtype which can confer responsiveness to phosphoantigens (PAgs) when genetically introduced into conventional αβ T-cells. Cancer immunotherapy using γδ TCR-engineered T-cells is currently under clinical evaluation. In this study, we sought to broaden the cancer specificity of the G115 γδ TCR by insertion of a tumour-binding peptide into the complementarity-determining region (CDR) three regions of the TCR δ2 chain. Peptides were selected from the foot and mouth disease virus A20 peptide which binds with high affinity and selectivity to αvβ6, an epithelial-selective integrin that is expressed by a range of solid tumours. Insertion of an A20-derived 12mer peptide achieved the best results, enabling the resulting G115 + A12 T-cells to kill both PAg and αvβ6-expressing tumour cells. Cytolytic activity of G115 + A12 T-cells against PAg-presenting K562 target cells was enhanced compared to G115 control cells, in keeping with the critical role of CDR3 δ2 length for optimal PAg recognition. Activation was accompanied by interferon (IFN)-γ release in the presence of either target antigen, providing a novel dual-specificity approach for cancer immunotherapy.

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“…Vector production was performed as described [ 21 ]. In brief, 3 × 10 6 HEK293T cells were plated into 10 cm 2 tissue culture-treated plates.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

Davies,

Pugliese,

Parente Pereira

et al. 2024

Biology

Self Cite

View full text Add to dashboard Cite

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“…To assess cytolytic activity of CAR T-cells, tumour and lymphoma cells were co-cultivated with T-cells at a one-to-one effector to target (E:T) ratio. In the case of T47D, tumour cell viability was determined using an MTT assay, as described in [ 25 ]. In the case of M-CSFR-expressing lymphoma cells, viability was determined by luciferase assays, as described in [ 26 ].…”

Section: Methodsmentioning

confidence: 99%

CAR T-Cell Targeting of Macrophage Colony-Stimulating Factor Receptor

Achkova

Beatson

Maher

2022

Cells

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Macrophage colony-stimulating factor receptor (M-CSFR) is found in cells of the mononuclear phagocyte lineage and is aberrantly expressed in a range of tumours, in addition to tumour-associated macrophages. Consequently, a variety of cancer therapies directed against M-CSFR are under development. We set out to engineer chimeric antigen receptors (CARs) that employ the natural ligands of this receptor, namely M-CSF or interleukin (IL)-34, to achieve specificity for M-CSFR-expressing target cells. Both M-CSF and IL-34 bind to overlapping regions of M-CSFR, although affinity of IL-34 is significantly greater than that of M-CSF. Matched second- and third-generation CARs targeted using M-CSF or IL-34 were expressed in human T-cells using the SFG retroviral vector. We found that both M-CSF- and IL-34-containing CARs enable T-cells to mediate selective destruction of tumour cells that express enforced or endogenous M-CSFR, accompanied by production of both IL-2 and interferon (IFN)-γ. Although they contain an additional co-stimulatory module, third-generation CARs did not outperform second-generation CARs. M-CSF-containing CARs mediated enhanced cytokine production and cytolytic activity compared to IL-34-containing CARs. These data demonstrate the feasibility of targeting M-CSFR using ligand-based CARs and raise the possibility that the low picomolar affinity of IL-34 for M-CSFR is detrimental to CAR function.

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CAR-Based Immunotherapy of Solid Tumours—A Survey of the Emerging Targets

Maher

Davies

2023

Cancers

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Immunotherapy with CAR T-cells has revolutionised the treatment of B-cell and plasma cell-derived cancers. However, solid tumours present a much greater challenge for treatment using CAR-engineered immune cells. In a partner review, we have surveyed data generated in clinical trials in which patients with solid tumours that expressed any of 30 discrete targets were treated with CAR-based immunotherapy. That exercise confirms that efficacy of this approach falls well behind that seen in haematological malignancies, while significant toxic events have also been reported. Here, we consider approximately 60 additional candidates for which such clinical data are not available yet, but where pre-clinical data have provided support for their advancement to clinical evaluation as CAR target antigens.

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Generation of human parallel chimeric antigen receptor (pCAR) T cells to achieve synergistic T cell co-stimulation

Cited by 3 publications

References 17 publications

Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

CAR T-Cell Targeting of Macrophage Colony-Stimulating Factor Receptor

CAR-Based Immunotherapy of Solid Tumours—A Survey of the Emerging Targets

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