Tockyを使って、がん免疫療法研究を立ち上げるまで

2022

1. CD4 T cell dynamics shape the immune response to combination oncolytic herpes virus and BRAF inhibitor therapy for melanoma

   Galabina Bozhanova , Jehanne Hassan , Lizzie Appleton , Victoria Jennings , Shane Foo , Martin McLaughlin , Charleen ML Chan Wah Hak , Emmanuel C Patin , Eva Crespo-Rodriguez , Gabby Baker , Edward Armstrong , Matthew Chiu , Hardev Pandha , Adel Samson , Victoria Roulstone , Joan Kyula , Richard Vile , Fiona Errington-Mais , Malin Pedersen , Kevin Harrington , Masahiro Ono, and Alan Melcher

   Journal for ImmunoTherapy of Cancer, May 2022

   Abstract

   Background Combination herpes simplex virus (HSV) oncolytic virotherapy and BRAF inhibitors (BRAFi) represent promising immunogenic treatments for BRAF mutant melanoma, but an improved understanding of the immunobiology of combinations is needed to improve on the benefit of immune checkpoint inhibitors (ICI).Methods Using a BRAFV600E-driven murine melanoma model, we tested the immunogenicity of HSV/BRAFi in immunocompetent C57BL mice. In addition to standard FACS analysis, we used the ‘Timer of Cell Kinetics and Activity’ system, which can analyze the temporal dynamics of different T cell subsets. This immune data was used to inform the selection of ICI for triple combination therapy, the effects of which were then further characterized using transcriptomics.Results Adding BRAFi treatment to HSV improved anti-tumor effects in vivo but not in vitro. Immune characterization showed HSV or dual therapy led to fewer intratumoral Treg, although with a more activated phenotype, together with more effector CD8 +T cells. Tocky analysis further showed that HSV/BRAFi dual treatment reduced the Tocky signal (reflecting engagement with cognate antigen), in both Treg and conventional subsets of CD4+, but not in CD8 +cells. However, a higher percentage of Treg than of conventional CD4 +maintained frequent engagement with antigens on treatment, reflecting a predominance of suppressive over effector function within the CD4 +compartment. The only T cell subset which correlated with a reduction in tumor growth was within Tocky signal positive conventional CD4+, supporting their therapeutic role. Targeting CD25 high, antigen-engaged Treg with a depleting anti-CD25 ICI, achieved complete cures in 100% of mice with triple therapy. Transcriptomic analysis confirmed reduction in Foxp3 on addition of anti-CD25 to HSV/BRAFi, as well as increases in expression of genes reflecting interferon signaling and cytotoxic activity.Conclusions Combination HSV/BRAFi is an immunogenic therapy for BRAF mutant melanoma, but cannot fully control tumors. Dual therapy results in changes in T cell dynamics within tumors, with relatively maintained antigen signaling in Treg compared with conv CD4+. Antigen-engaged CD4 +effectors correlate with tumor growth control, and depletion of Treg by addition of an anti-CD25 ICI, releasing suppression of conventional CD4 +effectors by Treg, enhances survival and activates immune signaling within tumors.All data relevant to the study are included in the article or uploaded as online supplemental information. Not applicable.

2. Single-cell level temporal profiling of tumour-reactive T cells under immune checkpoint blockade

   Jehanne Hassan , Elizabeth Appleton , Bahire Kalfaoglu , Malin Pedersen , José Almeida-Santos , Hisashi Kanemaru , Nobuko Irie , Shane Foo , Omnia Reda , Benjy J.Y. Tan , Il-mi Okazaki , Taku Okazaki , Yorifumi Satou , Kevin Harrington , Alan Melcher , and Masahiro Ono

   bioRxiv, May 2022

   Abstract Publication

   The blockade of the immune checkpoints PD-1 and CTLA-4 enhances T cell response. However, it is largely unknown how antigen-reactive T cells regulate their checkpoint expression in vivo and whether and how the checkpoint blockade can change activation dynamics of tumour-reactive T cells. To address this, here we used Nr4a3-Timer-of-cell-kinetics-and-activity (Tocky), which allows analysis of temporal changes of activated T cells following TCR signalling in vivo. By analysing melanoma-bearing Nr4a3 Tocky mice, we elucidate hidden dynamics of tumour-reactive T cells in the steady-state. Checkpoint blockade depleted highly activated effector Treg, while promoting unique effector T cell populations, and thus differentially modulating activation of tumour-reactive T cell populations. Furthermore, multidimensional analysis and seamless analysis of Tocky and scRNA-seq revealed a full spectrum of T cell dynamics in response to tumour burden and treatment with checkpoint blockade. Lastly, we propose a rational design of combinatorial therapy to further enhance T cell activities.Competing Interest StatementThe authors have declared no competing interest.