The getaway CAR-T cell: High resolution microscopy of self-protection in the immune synapse

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The getaway CAR-T cell: High resolution microscopy of self-protection in the immune synapse

Walsh, Emily; (2024) The getaway CAR-T cell: High resolution microscopy of self-protection in the immune synapse. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Cytotoxic T lymphocytes (CTLs) are adaptive immune cells critical for anti-tumour immune activity. Chimeric antigen receptor T cells (CAR-T cells) are genetically engineered T cells that express a novel Chimeric antigen receptor (CAR) to direct T cell killing towards specific cancer cell targets. CAR-T cells show clinical success in treating B cell haematological cancers, but research is ongoing to overcome problems with resistance, side effects, and to develop treatments for T cell haematological cancers. CTLs act as unilateral serial killers of B cell and T cell targets. This function critically depends on self-protection against perforin, a pore-forming protein secreted into the immune synapse (IS). For CTLs, this protection is partially dependent on membrane order and charge. We hypothesised that CAR-T cells maintain similar membrane-based protection as T cells, including lipid order enhancement and phosphatidylserine externalisation, and that this local enhancement at the CAR-T cell membrane provides protection in the IS even in the case of T cell target synapses, i.e. when effector and target cell have the same global membrane properties. By using both primary CAR-T cell vs B cell, and primary CAR-T cell vs T cell models, we show that CAR-T cells and T cells both have higher global resistance to recombinant human perforin than various other cell types. We demonstrate, with evidence from extensive imaging of primary CAR-T cell vs T cell target synapses, that target T cells are killed in CAR-T cell vs T cell synapses while effectors remain unscathed, indicating that CAR-T cell effector cells are locally better protected against perforin than their T cell targets. We also show that effector externalisation of phosphatidylserine in the synapse is preserved. Our data reveals key details about killing mechanisms in CAR-T cells that may guide future CAR and CAR-T cell adjuvant design against T cell lymphoma.

Type:	Thesis (Doctoral)
Qualification:	Ph.D
Title:	The getaway CAR-T cell: High resolution microscopy of self-protection in the immune synapse
Open access status:	An open access version is available from UCL Discovery
Language:	English
Additional information:	Copyright © The Author 2024. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
UCL classification:	UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Medicine
URI:	https://discovery.ucl.ac.uk/id/eprint/10200495