Remodeling of the tumor microenvironment via disrupting Blimp1(+) effector Treg activity augments response to anti-PD-1 blockade.

Accumulation of Foxp3+ regulatory T (Treg) cells in the tumor often represents an important mechanism for cancer immune evasion and a critical barrier to anti-tumor immunity and immunotherapy. Many tumor-infiltrating Treg cells display an activated phenotype and express the transcription factor Blimp1. However, the specific impact of these Blimp1+ Treg cells and their follicular regulatory T (TFR) cell subset on tumor and the underlying mechanisms of action are not yet well-explored. Various transplantable tumor models were established in immunocompetent wild-type mice and mice with a Foxp3-specific ablation of Blimp1. Tumor specimens from patients with metastatic melanoma and TCGA datasets were analyzed to support the potential role of Treg and TFR cells in tumor immunity. In vitro culture assays and in vivo adoptive transfer assays were used to understand how Treg, TFR cells and antibody responses influence tumor control. RNA sequencing and NanoString analysis were performed to reveal the transcriptome of tumor-infiltrating Treg cells and tumor cells, respectively. Finally, the therapeutic effects of anti-PD-1 treatment combined with the disruption of Blimp1+ Treg activity were evaluated. Blimp1+ Treg and TFR cells were enriched in the tumors, and higher tumoral TFR signatures indicated increased risk of melanoma metastasis. Deletion of Blimp1 in Treg cells resulted in impaired suppressive activity and a reprogramming into effector T-cells, which were largely restricted to the tumor-infiltrating Treg population. This destabilization combined with increased anti-tumor effector cellular responses, follicular helper T-cell expansion, enhanced tumoral IgE deposition and activation of macrophages secondary to dysregulated TFR cells, remodeled the tumor microenvironment and delayed tumor growth. The increased tumor immunogenicity with MHC upregulation improved response to anti-PD-1 blockade. Mechanistically, Blimp1 enforced intratumoral Treg cells with a unique transcriptional program dependent on Eomesodermin (Eomes) expression; deletion of Eomes in Blimp1-deficient Treg cells restored tumor growth and attenuated anti-tumor immunity. These findings revealed Blimp1 as a new critical regulator of tumor-infiltrating Treg cells and a potential target for modulating Treg activity to treat cancer. Our study has also revealed two FCERIA-containing immune signatures as promising diagnostic or prognostic markers for melanoma patients.

Dixon ML ，Luo L ，Ghosh S ，Grimes JM ，Leavenworth JD ，Leavenworth JW ... -  《Molecular Cancer》

Tumor microenvironment dictates regulatory T cell phenotype: Upregulated immune checkpoints reinforce suppressive function.

Kim HR ，Park HJ ，Son J ，Lee JG ，Chung KY ，Cho NH ，Shim HS ，Park S ，Kim G ，In Yoon H ，Kim HG ，Jung YW ，Cho BC ，Park SY ，Rha SY ，Ha SJ ... -  《Journal for ImmunoTherapy of Cancer》

Therapeutic depletion of CCR8(+) tumor-infiltrating regulatory T cells elicits antitumor immunity and synergizes with anti-PD-1 therapy.

Modulation and depletion strategies of regulatory T cells (Tregs) constitute valid approaches in antitumor immunotherapy but suffer from severe adverse effects due to their lack of selectivity for the tumor-infiltrating (ti-)Treg population, indicating the need for a ti-Treg specific biomarker. We employed single-cell RNA-sequencing in a mouse model of non-small cell lung carcinoma (NSCLC) to obtain a comprehensive overview of the tumor-infiltrating T-cell compartment, with a focus on ti-Treg subpopulations. These findings were validated by flow cytometric analysis of both mouse (LLC-OVA, MC38 and B16-OVA) and human (NSCLC and melanoma) tumor samples. We generated two CCR8-specific nanobodies (Nbs) that recognize distinct epitopes on the CCR8 extracellular domain. These Nbs were formulated as tetravalent Nb-Fc fusion proteins for optimal CCR8 binding and blocking, containing either an antibody-dependent cell-mediated cytotoxicity (ADCC)-deficient or an ADCC-prone Fc region. The therapeutic use of these Nb-Fc fusion proteins was evaluated, either as monotherapy or as combination therapy with anti-programmed cell death protein-1 (anti-PD-1), in both the LLC-OVA and MC38 mouse models. We were able to discern two ti-Treg populations, one of which is characterized by the unique expression of Ccr8 in conjunction with Treg activation markers. Ccr8 is also expressed by dysfunctional CD4+ and CD8+ T cells, but the CCR8 protein was only prominent on the highly activated and strongly T-cell suppressive ti-Treg subpopulation of mouse and human tumors, with no major CCR8-positivity found on peripheral Tregs. CCR8 expression resulted from TCR-mediated Treg triggering in an NF-κB-dependent fashion, but was not essential for the recruitment, activation nor suppressive capacity of these cells. While treatment of tumor-bearing mice with a blocking ADCC-deficient Nb-Fc did not influence tumor growth, ADCC-prone Nb-Fc elicited antitumor immunity and reduced tumor growth in synergy with anti-PD-1 therapy. Importantly, ADCC-prone Nb-Fc specifically depleted ti-Tregs in a natural killer (NK) cell-dependent fashion without affecting peripheral Tregs. Collectively, our findings highlight the efficacy and safety of targeting CCR8 for the depletion of tumor-promoting ti-Tregs in combination with anti-PD-1 therapy.

Van Damme H ，Dombrecht B ，Kiss M ，Roose H ，Allen E ，Van Overmeire E ，Kancheva D ，Martens L ，Murgaski A ，Bardet PMR ，Blancke G ，Jans M ，Bolli E ，Martins MS ，Elkrim Y ，Dooley J ，Boon L ，Schwarze JK ，Tacke F ，Movahedi K ，Vandamme N ，Neyns B ，Ocak S ，Scheyltjens I ，Vereecke L ，Nana FA ，Merchiers P ，Laoui D ，Van Ginderachter JA ... -  《Journal for ImmunoTherapy of Cancer》

Lactic acid promotes PD-1 expression in regulatory T cells in highly glycolytic tumor microenvironments.

The balance of programmed death-1 (PD-1)-expressing CD8+ T cells and regulatory T (Treg) cells in the tumor microenvironment (TME) determines the clinical efficacy of PD-1 blockade therapy through the competition of their reactivation. However, factors that determine this balance remain unknown. Here, we show that Treg cells gain higher PD-1 expression than effector T cells in highly glycolytic tumors, including MYC-amplified tumors and liver tumors. Under low-glucose environments via glucose consumption by tumor cells, Treg cells actively absorbed lactic acid (LA) through monocarboxylate transporter 1 (MCT1), promoting NFAT1 translocation into the nucleus, thereby enhancing the expression of PD-1, whereas PD-1 expression by effector T cells was dampened. PD-1 blockade invigorated the PD-1-expressing Treg cells, resulting in treatment failure. We propose that LA in the highly glycolytic TME is an active checkpoint for the function of Treg cells in the TME via upregulation of PD-1 expression.

Kumagai S ，Koyama S ，Itahashi K ，Tanegashima T ，Lin YT ，Togashi Y ，Kamada T ，Irie T ，Okumura G ，Kono H ，Ito D ，Fujii R ，Watanabe S ，Sai A ，Fukuoka S ，Sugiyama E ，Watanabe G ，Owari T ，Nishinakamura H ，Sugiyama D ，Maeda Y ，Kawazoe A ，Yukami H ，Chida K ，Ohara Y ，Yoshida T ，Shinno Y ，Takeyasu Y ，Shirasawa M ，Nakama K ，Aokage K ，Suzuki J ，Ishii G ，Kuwata T ，Sakamoto N ，Kawazu M ，Ueno T ，Mori T ，Yamazaki N ，Tsuboi M ，Yatabe Y ，Kinoshita T ，Doi T ，Shitara K ，Mano H ，Nishikawa H ... -  《-》

Bcl6 and Blimp1 reciprocally regulate ST2(+) Treg-cell development in the context of allergic airway inflammation.

Bcl6 is required for the development of T follicular helper cells and T follicular regulatory (Tfr) cells that regulate germinal center responses. Bcl6 also affects the function of regulatory T (Treg) cells. The goal of this study was to define the functions of Bcl6 in Treg cells, including Tfr cells, in the context of allergic airway inflammation. We used a model of house dust mite sensitization to challenge wild-type, Bcl6fl/fl Foxp3-Cre, and Prdm1 (Blimp1)fl/fl Foxp3-Cre mice to study the reciprocal roles of Bcl6 and Blimp1 in allergic airway inflammation. In the house dust mite model, Tfr cells repress the production of IgE and Bcl6+ Treg cells suppress the generation of type 2 cytokine-producing cells in the lungs. In mice with Bcl6-deficient Treg cells, twice as many ST2+ (IL-33R+) Treg cells develop as are observed in wild-type mice. ST2+ Treg cells in the context of allergic airway inflammation are Blimp1 dependent, express type 2 cytokines, and share features of visceral adipose tissue Treg cells. Bcl6-deficient Treg cells are more susceptible, and Blimp1-deficient Treg cells are resistant, to acquiring the ST2+ Treg-cell phenotype in vitro and in vivo in response to IL-33. Bcl6-deficient ST2+ Treg cells, but not Bcl6-deficient ST2+ conventional T cells, strongly promote allergic airway inflammation when transferred into recipient mice. Lastly, ST2 is required for the exacerbated allergic airway inflammation in Bcl6fl/fl Foxp3-Cre mice. During allergic airway inflammation, Bcl6 and Blimp1 play dual roles in regulating Tfr-cell activity in the germinal center and in the development of ST2+ Treg cells that promote type 2 cytokine responses.

Koh B ，Ulrich BJ ，Nelson AS ，Panangipalli G ，Kharwadkar R ，Wu W ，Xie MM ，Fu Y ，Turner MJ ，Paczesny S ，Janga SC ，Dent AL ，Kaplan MH ... -  《-》