Journal
FRONTIERS IN IMMUNOLOGY
Volume 11, Issue -, Pages -Publisher
FRONTIERS MEDIA SA
DOI: 10.3389/fimmu.2020.01514
Keywords
intravital imaging; adoptive T-cell therapy; immune checkpoint inhibitors; immunocompetent; melanoma
Categories
Funding
- CRUK Cambridge Centre [C9685/A25177]
- Cambridge Commonwealth, European and International Trust PhD Scholarship
- Cancer Research UK (CRUK) [C19212/A16628, C19212/A911376]
- CRUK Cancer Imaging Centre in Cambridge and Manchester [C197/A16465]
- Engineering and Physical Sciences Research Council (EPSRC) Cancer Imaging Centre in Cambridge and Manchester [C197/A16465]
- National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre
- BBSRC [BBS/E/B/000C0428, BBS/E/B/0000H329, BBS/E/B/000C0427] Funding Source: UKRI
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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 Levy-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.
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