Inhibition of estrogen signaling in myeloid cells increases tumor immunity in melanoma
Binita Chakraborty, … , Ching-Yi Chang, Donald P. McDonnell
Binita Chakraborty, … , Ching-Yi Chang, Donald P. McDonnell
Published October 12, 2021
Citation Information: J Clin Invest. 2021;131(23):e151347. https://doi.org/10.1172/JCI151347.
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Research Article Oncology

Inhibition of estrogen signaling in myeloid cells increases tumor immunity in melanoma

Abstract

Immune checkpoint blockade (ICB) therapies have significantly prolonged patient survival across multiple tumor types, particularly in melanoma. Interestingly, sex-specific differences in response to ICB have been observed, with males receiving a greater benefit from ICB than females, although the mechanism or mechanisms underlying this difference are unknown. Mining published transcriptomic data sets, we determined that the response to ICBs is influenced by the functionality of intratumoral macrophages. This puts into context our observation that estrogens (E2) working through the estrogen receptor α (ERα) stimulated melanoma growth in murine models by skewing macrophage polarization toward an immune-suppressive state that promoted CD8+ T cell dysfunction and exhaustion and ICB resistance. This activity was not evident in mice harboring macrophage-specific depletion of ERα, confirming a direct role for estrogen signaling within myeloid cells in establishing an immunosuppressed state. Inhibition of ERα using fulvestrant, a selective estrogen receptor downregulator (SERD), decreased tumor growth, stimulated adaptive immunity, and increased the antitumor efficacy of ICBs. Further, a gene signature that determines ER activity in macrophages predicted survival in patients with melanoma treated with ICB. These results highlight the importance of E2/ER signaling as a regulator of intratumoral macrophage polarization, an activity that can be therapeutically targeted to reverse immune suppression and increase ICB efficacy.

Authors

Binita Chakraborty, Jovita Byemerwa, Jonathan Shepherd, Corinne N. Haines, Robert Baldi, Weida Gong, Wen Liu, Debarati Mukherjee, Sandeep Artham, Felicia Lim, Yeeun Bae, Olivia Brueckner, Kendall Tavares, Suzanne E. Wardell, Brent A. Hanks, Charles M. Perou, Ching-Yi Chang, Donald P. McDonnell

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Figure 4

E2 regulates TAM function.

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E2 regulates TAM function. (A–C) Ratio of M1/M2 macrophages in iBP (n = ...
(A–C) Ratio of M1/M2 macrophages in iBP (n = 6) (A) and B16F10 (n = 6–10) (B) tumors from placebo- and E2-treated mice and representative flow cytometric plots of M2 and M1 macrophages in the B16F10 model (C). (D) Growth of B16F10 tumors (n = 12) upon depletion of macrophages by clodronate liposomes (Clod Lipo) in ovariectomized mice supplemented with placebo or E2. Cont Lipo, control liposome. (E and F) Quantification of IFN-γ+CD8+ T cells (E) and GZMB+CD8+ T cells (F) (n = 3) that were cocultured with BMDMs differentiated in NM or TCM and treated with either DMSO or E2 (1 nM). (G and H) CFSE dilution and quantification representing the proliferation of CFSElo/–CD8+ T cells (n = 3) after coculturing with BMDMs from Esr1fl/fl and Esr1fl/fl LysMCre mice, differentiated in either normal media or TCM (B16F10), followed by treatment with either DMSO or E2 (1 nM). (I–K) Quantification of CD8+IFN-γ+, CD8+CD44+CD69+, and CD8+GZMB+ T cells (n = 3) from the same experiment as in G. (L) Schematic of tumor comixing methodology. (M) Syngeneic tumor growth of YuMM5.2 (5 × 105) cells comixed with BMDMs from either Esr1fl/fl LysMCre mice or their Esr1fl/fl littermate controls (1:1) in ovariectomized mice supplemented with either placebo or E2. Esr1fl/fl BMDMs plus YuMM5.2 (Placebo + WT MΦ; black; n = 10); Esr1fl/fl LysMCre BMDMs plus YuMM5.2 (Placebo + ER-KO MΦ; blue; n = 10); Esr1fl/fl BMDMs plus YuMM5.2 (E2 + WT MΦ; red; n = 10); and Esr1fl/fl LysMCre BMDMs plus YuMM5.2 (E2 + ER-KO MΦ; brown; n = 10). (N) uMAP representation of macrophage/monocyte subclusters as determined by scRNA-Seq. (O) Trajectory analysis depicting the differentiation of monocytes into different lineages of macrophages. (P) Density of cells in macrophage/monocyte subclusters along a pseudotime gradient. (Q) Expression of M2-associated genes (Cd163, Lgr2, Retnla, and Folr2) in macrophage clusters along the pseudotime axis. Data in E–K are representative of 2 independent experiments. Data are expressed as individual data points and indicate the mean ± SEM. *P < 0.05, **P < 0.01, and ****P < 0.0001, by Student’s t test (A and B), 1-way ANOVA (E–G, I–K) and by 2-way ANOVA followed by Bonferroni’s multiple-correction test (D and M). MΦ, macrophages.