Targeting tumor-associated macrophages and granulocytic myeloid-derived suppressor cells augments PD-1 blockade in cholangiocarcinoma
Emilien Loeuillard, Jingchun Yang, EeeLN Buckarma, Juan Wang, Yuanhang Liu, Caitlin Conboy, Kevin D. Pavelko, Ying Li, Daniel O’Brien, Chen Wang, Rondell P. Graham, Rory L. Smoot, Haidong Dong, Sumera Ilyas
Emilien Loeuillard, Jingchun Yang, EeeLN Buckarma, Juan Wang, Yuanhang Liu, Caitlin Conboy, Kevin D. Pavelko, Ying Li, Daniel O’Brien, Chen Wang, Rondell P. Graham, Rory L. Smoot, Haidong Dong, Sumera Ilyas
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Research Article Gastroenterology Oncology

Targeting tumor-associated macrophages and granulocytic myeloid-derived suppressor cells augments PD-1 blockade in cholangiocarcinoma

Abstract

Immune checkpoint blockade (ICB) has revolutionized cancer therapeutics. Desmoplastic malignancies, such as cholangiocarcinoma (CCA), have an abundant tumor immune microenvironment (TIME). However, to date, ICB monotherapy in such malignancies has been ineffective. Herein, we identify tumor-associated macrophages (TAMs) as the primary source of programmed death–ligand 1 (PD-L1) in human and murine CCA. In a murine model of CCA, recruited PD-L1+ TAMs facilitated CCA progression. However, TAM blockade failed to decrease tumor progression due to a compensatory emergence of granulocytic myeloid-derived suppressor cells (G-MDSCs) that mediated immune escape by impairing T cell response. Single-cell RNA sequencing (scRNA-Seq) of murine tumor G-MDSCs highlighted a unique ApoE G-MDSC subset enriched with TAM blockade; further analysis of a human scRNA-Seq data set demonstrated the presence of a similar G-MDSC subset in human CCA. Finally, dual inhibition of TAMs and G-MDSCs potentiated ICB. In summary, our findings highlight the therapeutic potential of coupling ICB with immunotherapies targeting immunosuppressive myeloid cells in CCA.

Authors

Emilien Loeuillard, Jingchun Yang, EeeLN Buckarma, Juan Wang, Yuanhang Liu, Caitlin Conboy, Kevin D. Pavelko, Ying Li, Daniel O’Brien, Chen Wang, Rondell P. Graham, Rory L. Smoot, Haidong Dong, Sumera Ilyas

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Figure 1

TAMs are the predominant source of PD-L1 in CCA.

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TAMs are the predominant source of PD-L1 in CCA. (A) Representative imag...
(A) Representative images (left and middle panels) of PD-L1 (brown staining, black arrowhead) plus CD68 (red staining, red arrowhead) coimmunostaining (n = 33) and PD-L1 (brown staining) plus CK-19 (red staining) coimmunostaining (n = 18) in human resected CCA specimens. Percentage of patients with positive PD-L1/CD68 costaining and PD-L1/CK19 costaining, respectively (right panel). Scale bars: 40 μm. (B) Histograms show expression of PD-L1+ macrophages in human CCA tumors. (C–F) Flow cytometry analysis of normal WT mouse livers (from WT mice without tumors) as well as adjacent livers and tumors of mice 28 days after orthotopic implantation of 1 × 106 SB (murine CCA) cells. (C) Percentage of PD-L1+ macrophages (Mφ) of total macrophages (CD45+ CD11b+F4/80+) in WT mouse normal liver, tumor-adjacent liver, or tumor. Fluorescence Minus One (FMO) controls were used for each independent experiment to establish gates (See Supplemental Figure 1A for gating strategy) (n ≥ 8). Representative histograms show expression of PD-L1+ macrophages. (D) Percentage of CD206+ TAMs (left panel) and PD-L1+CD206+ TAMs (middle panel) of F4/80int macrophages (CD45+ CD11b+F4/80int) in WT mouse liver, tumor-adjacent liver, or tumor. Representative contour plots (right panel) show CD206 and PD-L1 expression of F4/80int macrophages (n ≥ 7). (E) Percentage of PD-L1+CD206– macrophages or PD-L1+CD206+ macrophages (CD11b+F4/80+) of CD45+ cells from SB tumors (n = 28). (F) Percentage of PD-L1 expression in myeloid cells from SB tumors. Macrophages, CD45+PD-L1+CD11b+F4/80+; MDSCs, CD45+PD-L1+CD11c–CD11b+F4/80–GR-1+; DCs, CD45+PD-L1+CD11chi; (n = 11). Data are represented as mean ± SD. Unpaired Student’s t test (E) and 1-way ANOVA with Bonferroni’s post hoc test (C, D, and F) were used. *P < 0.05; **P < 0.01; ***P < 0.001.