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Intravital Imaging of Adoptive T-Cell Morphology, Mobility and Trafficking Following Immune Checkpoint Inhibition in a Mouse Melanoma Model.

Accepted version
Peer-reviewed

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Type

Article

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Authors

Lau, Doreen 
Garçon, Fabien 
Lechermann, Laura M 

Abstract

Efficient T-cell targeting, infiltration and activation within tumors is crucial for successful adoptive T-cell therapy. Intravital microscopy is a powerful tool for the visualization of T-cell behavior within tumors, as well as spatial and temporal heterogeneity in response to immunotherapy. Here we describe an experimental approach for intravital imaging of adoptive T-cell morphology, mobility and trafficking in a skin-flap tumor model, following immune modulation with immune checkpoint inhibitors (ICIs) targeting PD-L1 and CTLA-4. A syngeneic model of ovalbumin and mCherry-expressing amelanotic mouse melanoma was used in conjunction with adoptively transferred OT-1+ cytotoxic T-cells expressing GFP to image antigen-specific live T-cell behavior within the tumor microenvironment. Dynamic image analysis of T-cell motility showed distinct CD8+ T-cell migration patterns and morpho-dynamics within different tumor compartments in response to ICIs: this approach was used to cluster T-cell behavior into four groups based on velocity and meandering index. The results showed that most T-cells within the tumor periphery demonstrated Lévy-like trajectories, consistent with tumor cell searching strategies. T-cells adjacent to tumor cells had reduced velocity and appeared to probe the local environment, consistent with cell-cell interactions. An increased number of T-cells were detected following treatment, traveling at lower mean velocities than controls, and demonstrating reduced displacement consistent with target engagement. Histogram-based analysis of immunofluorescent images from harvested tumors showed that in the ICI-treated mice there was a higher density of CD31+ vessels compared to untreated controls and a greater infiltration of T-cells towards the tumor core, consistent with increased cellular trafficking post-treatment.

Description

Keywords

adoptive T-cell therapy, immune checkpoint inhibitors, immunocompetent, intravital imaging, melanoma, Adoptive Transfer, Animals, Antigens, Neoplasm, Cell Line, Tumor, Cell Movement, Combined Modality Therapy, Disease Models, Animal, Fluorescent Antibody Technique, Image Processing, Computer-Assisted, Immune Checkpoint Inhibitors, Immunotherapy, Adoptive, Lymphocyte Activation, Melanoma, Experimental, Mice, Molecular Imaging, T-Cell Antigen Receptor Specificity, T-Lymphocytes

Journal Title

Front Immunol

Conference Name

Journal ISSN

1664-3224
1664-3224

Volume Title

11

Publisher

Frontiers Media SA

Rights

All rights reserved
Sponsorship
Cancer Research UK (C12912/A27150)
Wellcome Trust (206618/Z/17/Z)
Cancer Research UK (C96/A25177)
This project was supported by the CRUK Cambridge Centre (C9685/A25177) and Cambridge Commonwealth, European and International Trust PhD Scholarship, Cancer Research UK (CRUK; C19212/A16628, C19212/A911376), the CRUK & Engineering and Physical Sciences Research Council (EPSRC) Cancer Imaging Centre in Cambridge and Manchester (C197/A16465) and the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre.