Introduction and Purpose:

CD4+CD25+Foxp3+ regulatory T cells (Tregs) are known to suppress immune responses against cancers. Treg-mediated immunosuppression is a key mechanism for tumor immune-evasion, which could lead cancer immunotherapies to failures. Systemic depletion of Tregs have been tried by using antibodies against them; however, intravenously delivering these antibodies might not sufficiently deplete Tregs in the tumor microenvironment or could substantially deplete effector cells, in case anti-CD25 antibody was used. In addition, development of auto-immune responses could cause potential side effects. To overcome these problems, we exploited near infrared photoimmunotherapy (NIR-PIT) at the tumor to quickly deplete only intratumoral Tregs, aiming to induce anti-tumor immune activation.

Experimental Design:

F(ab’)2 fragments of an anti-mouse CD25 antibody, PC61.3, were generated and conjugated with a phthalocyanine dye, IRDye-700DX (αnti-CD25-F(ab’)2-IR700). Using the anti-CD25-F(ab’)2-IR700, in vitro NIR-PIT effect was examined against CD25-expressing mouse T lymphocytes, HT-2 clone A5E cells. In vivo CD25-target-NIR-PIT of the tumor was performed after an intravenous injection of the conjugate to mice bearing subcutaneous, luciferase transfected, LL/2 (Lewis lung carcinoma, LL/2-luc) or MC38 (mouse colon cancer, MC38-luc) cancers. Tumor volume, bioluminescence signals (BLI), and Immune responses following a local CD25-target-NIR-PIT were examined.

Results:

In vitro NIR-PIT-induced cytotoxicity was light dose dependent. CD25-target-NIR-PIT at the tumor quickly and selectively depleted Tregs in the cancers, yet, Tregs in any other organs were not affected. The local CD25-target-NIR-PIT induced a rapid activation and cytotoxic action of CD8 T cells and NK cells within LL/2-luc or MC38-luc tumors. This led to significant reductions of tumor volume (p < 0.0001) and BLI (p < 0.05) in both LL/2-luc and MC38-luc models and prolonged the survival of the mice (p < 0.0001) compared to the non-treated controls. Intriguingly, this local CD25-target-NIR-PIT induced a transient systemic cytokine storm and antitumor-effects on distant non-irradiated specific tumors. Effects of local CD25-target-NIR-PIT were significantly (p < 0.0001) inhibited by a CD8-, NK-, or INFγ−depletion, suggesting the anti-tumor roles of CD8 T cells and NK cells.

Conclusions:

Quick depletion of intratumoral Tregs by a local CD25-target-NIR-PIT rapidly induced CD8 T- and NK-cell activation, thereby restoring local anti-tumor immunity. Consequently, activated immunity led to regression of not only NIR-PIT-treated tumors but also non-NIR light exposed tumors in separate parts of the body. These observations suggest that using local CD25-target-NIR-PIT may be a promising new strategy for cancer immunotherapy.

Citation Format: Kazuhide Sato, Noriko Sato, Biying Xu, Yuko Nakamura, Tadanobu Nagaya, Peter L. Choyke, Hisataka Kobayashi. Cancer treatment by near infrared photoimmunotherapy targeting intratumoral regulatory T cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1470.