CD4+ T cells induce rejection of urothelial tumors after immune checkpoint blockade
Yuji Sato, … , Jennifer K. Sehn, Vivek K. Arora
Yuji Sato, … , Jennifer K. Sehn, Vivek K. Arora
Published December 6, 2018
Citation Information: JCI Insight. 2018;3(23):e121062. https://doi.org/10.1172/jci.insight.121062.
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Research Article Immunology Oncology

CD4+ T cells induce rejection of urothelial tumors after immune checkpoint blockade

Abstract

Immune checkpoint blockade (ICB) provides clinical benefit to a minority of patients with urothelial carcinoma (UC). The role of CD4+ T cells in ICB-induced antitumor activity is not well defined; however, CD4+ T cells are speculated to play a supportive role in the development of CD8+ T cells that kill tumor cells after recognition of tumor antigens presented by MHC class I. To investigate the mechanisms of ICB-induced activity against UC, we developed mouse organoid-based transplantable models that have histologic and genetic similarity to human bladder cancer. We found that ICB can induce tumor rejection and protective immunity with these systems in a manner dependent on CD4+ T cells but not reliant on CD8+ T cells. Evaluation of tumor infiltrates and draining lymph nodes after ICB revealed expansion of IFN-γ–producing CD4+ T cells. Tumor cells in this system express MHC class I, MHC class II, and the IFN-γ receptor (Ifngr1), but none were necessary for ICB-induced tumor rejection. IFN-γ neutralization blocked ICB activity, and, in mice depleted of CD4+ T cells, IFN-γ ectopically expressed in the tumor microenvironment was sufficient to inhibit growth of tumors in which the epithelial compartment lacked Ifngr1. Our findings suggest unappreciated CD4+ T cell–dependent mechanisms of ICB activity, principally mediated through IFN-γ effects on the microenvironment.

Authors

Yuji Sato, Jennifer K. Bolzenius, Abdallah M. Eteleeb, Xinming Su, Christopher A. Maher, Jennifer K. Sehn, Vivek K. Arora

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Figure 5

IFN-γ mediates ICB activity and is sufficient to inhibit tumor growth.

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IFN-γ mediates ICB activity and is sufficient to inhibit tumor growth. (...
(A) αPD-1 and αCTLA-4 combination treatment from day 9 to 24 coadministered with IFN-γ–neutralizing antibodies administered i.p. every 3 days from day 8 to 23. Tumor sizes were compared for an additional 9 days after the last IFN-γ neutralization, a time frame within the reported half-life of the neutralizing antibody. Data represent mean tumor diameter ± SEM. n = 5 per group. (B) Quantification of Ck5 staining of MCB6C tumor sections obtained 5 days after initiation of combination ICB with and without IFN-γ neutralization. IFN-γ neutralization antibody was administered on days 8 and 11 after MCB6C injection. Quantification was performed using images at an original magnification of ×20. For each tumor, percentage Ck5 positivity was averaged from 4 independent fields and quantified using ImageJ software. The graph shows mean ± SD of 9 individual tumors from each treatment group. (C) Representative images used for B at low and high magnification. Scale bars: 1 mm (top); 200 μM (bottom). (D) MCB6C Infgr1-KO organoids constitutively expressing recombinant IFN-γ (rIFN-γ) were injected to mice. For all groups, mice were subjected to CD4+ T cell depletion that was started at day –1 and continued weekly throughout the duration of the experiment. IFN-γ neutralization or control treatments were also started at day –1 and continued weekly for the duration of the experiment. The low IFN-γ group was maintained on regular chow. The high IFN-γ group was initiated on doxycycline-containing chow at day 8. Constitutive low and high ectopic IFN-γ expression in tumor epithelial cells was confirmed by flow cytometry (see Supplemental Figure 7B). Data are plotted as mean ± SEM of n = 6–7 mice per group. (E) Mass of tumors described in D at day 34. (F) Ck5 staining and quantification as described in B. Representative images used for Ck5 quantification. Scale bars: 500 μM. Three tumors from each IFN-γ neutralization groups were scored for Ck5 positivity, and the remainder were utilized for flow cytometric analysis (See Supplemental Figure 7B). Comparisons for growth curves are by 2-way ANOVA for repeated measures and for column data are by Student’s t test. NS > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.