Functional Th1-oriented T follicular helper cells that infiltrate human breast cancer promote effective adaptive immunity
Grégory Noël, … , Denis Larsimont, Karen Willard-Gallo
Grégory Noël, … , Denis Larsimont, Karen Willard-Gallo
Published August 19, 2021
Citation Information: J Clin Invest. 2021;131(19):e139905. https://doi.org/10.1172/JCI139905.
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Functional Th1-oriented T follicular helper cells that infiltrate human breast cancer promote effective adaptive immunity

Abstract

We previously demonstrated that tumor-infiltrating lymphocytes (TIL) in human breast cancer sometimes form organized tertiary lymphoid structures (TLS) characterized by CXCL13-producing T follicular helper (Tfh) cells. The present study found that CD4+ Tfh TIL, CD8+ TIL, and TIL-B, colocalizing in TLS, all express the CXCL13 receptor CXCR5. An ex vivo functional assay determined that only activated, functional Th1-oriented Tfh TIL (PD-1hiICOSint phenotype) provide help for immunoglobulin and IFN-γ production. A functional Tfh TIL presence signals an active TLS, characterized by humoral (immunoglobulins, Ki-67+ TIL-B in active germinal centers) and cytotoxic (GZMB+CD8+ and GZMB+CD68+ TIL plus Th1 gene expression) immune responses. Analysis of active versus inactive TLS in untreated patients revealed that the former are associated with positive clinical outcomes. TLS also contain functional T follicular regulatory (Tfr) TIL, which are characterized by a CD25+CXCR5+GARP+FOXP3+ phenotype and a demethylated FOXP3 gene. Functional Tfr inhibited functional Tfh activities via a glycoprotein A repetitions predominant (GARP)-associated TGF-β–dependent mechanism. The activity of tumor-associated TLS was dictated by the relative balance between functional Tfh TIL and functional Tfr TIL. These data provide mechanistic insight into TLS processes orchestrated by functional Th1-oriented Tfh TIL, including TIL-B and CD8+ TIL activation and immunological memory generation. Tfh TIL, regulated by functional Tfr TIL, are an expected key target of PD-1/PD-L1 blockade.

Authors

Grégory Noël, Mireille Langouo Fontsa, Soizic Garaud, Pushpamali De Silva, Alexandre de Wind, Gert G. Van den Eynden, Roberto Salgado, Anaïs Boisson, Hanne Locy, Noémie Thomas, Cinzia Solinas, Edoardo Migliori, Céline Naveaux, Hugues Duvillier, Sophie Lucas, Ligia Craciun, Kris Thielemans, Denis Larsimont, Karen Willard-Gallo

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Figure 4

The balance of functional Tfh to functional Tfr TIL governs humoral immune responses in human BC.

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The balance of functional Tfh to functional Tfr TIL governs humoral immu...
(A) Correlation between the density of functional Tfh TIL (no./mg tumor; flow cytometry) and IgG (μg/mg tumor in the fresh tissue supernatant; n = 71; LumA [n = 20], LumB [n = 19], HER2+ [n = 12], and TN [n = 20]). (B) Representative FFPE sections (TNBC 1068 shown) were cIHC stained for scoring. Left: dual CD3/CD20 (brown/red) for T cell zones and B cell follicles; middle: dual PD-1/Ki-67 (brown/red); consecutive sections; right: zoomed image of the B cell follicle. The TLS is 1000 μm wide. Magnification: ×35 (left panels); ×140 (right panel). (C) Correlation between the frequency of PD-1+ cells (functional Tfh TIL surrogate) and Ki-67+ cells (GC TIL-B surrogate) in TLS-associated B cell follicles stained and scored as in B. Each point shows the mean score for all TLS in a given tumor. (D) The ratio of functional Tfh to functional Tfr TIL (flow cytometry) correlated with supernatant Igs (A). Upper panels: IgG (μg)/mg tumor; lower panels: IgA (μg)/mg tumor (n = 71; LumA [n = 20], LumB [n = 19], HER2+ [n = 12], TN [n = 20]). (E) mIHC for CD20 (green), Ki-67 (orange), CD4 (blue), PD-1 (red), ICOS (yellow), and Foxp3 (cyan) on a representative FFPE section (2 TLS; TNBC); zoomed image of the T cell zone. Contact scores between functional Tfr TIL (CD4+Foxp3+ICOS+) and Tfh TIL (CD4+PD-1–) or functional Tfh TIL (CD4+PD-1+) or Tfr TIL (CD4+Foxp3+ICOS–) or TIL-B (CD20+) in the T cell zone. Magnification: ×80 (upper right); ×100 (upper left); ×200 (lower panels). (F) Using the same mIHC-stained sections as in E, the ratios between (left) total Tfh TIL (PD-1–+PD-1+) and total Tfr TIL (ICOS–+ICOS+) and (right) functional Tfh TIL and functional Tfr TIL are shown for the T cell zones of 5 GC+ and 3 GC– TLS (TNBC; Student’s t test). Statistical testing for all correlations used linear regression analysis. *P < 0.05.