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2551 Specific PD-1 Deletion on Regulatory T Cells Leads to Enhanced Anti-Tumor Responses

Program: Oral and Poster Abstracts
Session: 203. Lymphocytes and Acquired or Congenital Immunodeficiency Disorders: Poster II
Hematology Disease Topics & Pathways:
Research, Fundamental Science, immune mechanism, immunology, Biological Processes
Sunday, December 10, 2023, 6:00 PM-8:00 PM

Konstantinos Aliazis, PhD1*, Anthos Christofides, MD1,2*, Halil-Ibrahim Aksoylar, PhD1*, Rushil Shah1*, Rinku Pal, BS1*, Nikolaos Patsoukis, PhD1,3* and Vassiliki A Boussiotis, MD, PhD1

1Hematology-Oncology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA
2Medicine, Yale University, New Heaven, CT
3Simcere of America, Cambridge, MA

Programmed cell death 1 (PD-1) is a T cell inhibitory receptor and the most extensively exploited therapeutic target of checkpoint immunotherapy in cancer. Ligation of PD-1 by its ligands PD-L1 or PD-L2 leads to inhibition of CD4+ and CD8+ T cell responses. However, very little is known about the specific role of PD-1 in regulatory T (Treg) cells. Previous studies have shown that PD-1 engagement during induction of Treg polarization by TCR/CD28 signaling in the presence of IL-2 and TGF-β promoted the generation and expansion of Treg cells with potent suppressive function. It has also been shown that patients with high numbers of PD-1+ Treg cells have unfavorable responses to PD-1 checkpoint immunotherapy. Moreover, patients who developed hyperprogressive disease after PD-1 blocking immunotherapy exhibited enhanced Treg cell proliferative ability compared to patients with non-hyperprogressive disease. In the present study, we sought to understand the role of PD-1 on Treg cells in the context of cancer. We generated mice with conditional targeting of the Pdcd1 gene (encoding for PD-1) and crossed them with FoxP3Cre-YFP (Pdcd1f/fFoxP3Cre) resulting in specific deletion of PD-1 in Treg cells. Under steady-state conditions, the percentage of Treg cells in the spleen and lymph nodes remained comparable between Pdcd1f/fFoxP3Cre and Pdcd1wt/wtFoxP3Cre control mice. However, PD-1-deficient Treg cells had a more activated phenotype compared to their counterparts in control mice, with higher expression of CD69, ICOS, and PD-L1. Furthermore, Treg cells in Pdcd1f/fFoxp3Cre mice expressed increased levels of GITR, CTLA-4, ICOS, and PD-L1 compared to endogenous CD4+ T conventional cells (CD4+Foxp3-, Tconv) and CD8+ T cells. To investigate the implications of these immunological changes in anti-tumor responses, we used the MC38 colon adenocarcinoma syngeneic mouse model. We found a smaller tumor size in Pdcd1f/fFoxP3Cre mice compared to their control counterparts. Treg cells from spleens and tumor-draining lymph nodes (tdLNs) showed minimal differences in their activation state, expression of checkpoint markers, and proliferation levels, as determined by direct ex vivo assessment. Similarly, CD4+ Tconv and CD8+ T cells from spleens and tdLNs showed a very low level of activation that was comparable between the two experimental groups. In the tumor site, however, Pdcd1f/fFoxp3Cre tumor-bearing mice had a significant reduction in the fractions of CD4+ Tconv and Treg cells and a significant increase in the fraction of CD8+ T cells compared to control tumor-bearing mice. Detailed analysis revealed that in tumor-bearing Pdcd1f/fFoxp3Cre mice intratumoral naive and central memory CD4+ T cells were reduced whereas CD4+ T effector cells were increased. Similarly, in Pdcd1f/fFoxp3Cre tumor-bearing mice we noticed a reduction in naïve and central memory CD8+ T cells but a significant expansion in the effector CD8+ T cells. No significant differences were detected in the expression of activation markers in tumor infiltrating Treg cells between the two groups. In contrast, in Pdcd1f/fFoxp3Cre tumor-bearing mice, tumor infiltrating CD4+ Tconv cells displayed an activated phenotype with upregulation of PD-L1, ICOS, TIM3, CD69, and CTLA-4, downregulated PD-1, and increased IFN-γ production. Tumor infiltrating CD8+ T cells also displayed an activated phenotype with upregulation of activation markers such as CD69, ICOS, PD-L1, TIM3, and LAG3 but reduced PD-1 expression. Notably, both CD4+ Tconv and CD8+ T cells proliferated more robustly in the Pdcd1f/fFoxp3Cre tumor-bearing mice as determined by the high expression of the proliferation marker Ki67. These results indicate that selective ablation of PD-1 in Treg cells confers a less suppressive immunological environment and allows for expansion and activation of CD4+ and CD8+ T cells, leading to anti-tumor responses from CD4+ and CD8+ T effector cells.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH