Abstract 1120: CD8 ImmunoPET imaging identifies an immunogenically active tumor microenvironment following radiotherapy in primary TNBC
Authors: Song Patrick N. , Lynch Shannon E. , DeMellier Chloe T. , Mascioni Alessandro , Fang Jia , Sorace Anna G. 
Published: March 2024

Background: Tumor immune composition has been shown to drive therapy response in triple negative breast cancer (TNBC). Radiation is a component of standard-of-care treatments for TNBC, however immune populations have not been studied in the context of radiation resistance in breast cancer. Positron emission tomography (PET) allows for quantifying molecular signatures of the tumor microenvironment, which can precede anatomical changes in tumor volume. Recently, development of immune-targeted imaging approaches has allowed for noninvasive quantification of immune cells, allowing for quantification and long-term therapy response. Preclinical studies have demonstrated that combining therapies, such as radiotherapy and immunotherapy, could be beneficial in treating cancer. The goal of this study is to quantify changes in immune infiltration with [89Zr]-CD8 ImmunoPET following radiation therapy to determine variations in immune activation that can affect therapeutic response in radiation sensitive or radiation resistant tumors.

Methods: Syngeneic radiation sensitive 4T1 or a radiation-resistant 4T1 tumors were treated with fractionated radiation (2 Gy/day) from day 0-5, injected with [89Zr]-CD8 minibody on day 6, and imaged with [89Zr]-CD8 PET on day 7. Following imaging, tumors were excised for biological assays (N = 20) or monitored for longitudinal changes in tumor volume (N = 15). Biological assays included CD8 immunofluorescence or assessment of immune population differences with flow cytometry against T-cells (CD3, CD4, CD8, IFN-gamma), macrophages (CD45, F4/80, CD86 and CD206), and natural killer cells (CD56 and CD16). A non-parametric T-test was used to assess for statistical differences.

Results: In radiation sensitive tumors, [89Zr]-CD8 PET indicated that radiation significantly increases CD8 immune infiltration (p = 0.02) compared to controls, which was correlated with immunofluorescence (p &lt; 0.01). Immune infiltration was increased in radiation sensitive tumors when treated with radiation therapy (4.6-fold increase in CD86+ M1-like macrophages, 4.7-fold increase in CD206+ M2-like macrophages, 5.3-fold increase in CD8 T-cells and 4.3-fold increase in CD4 T-cells) compared to control tumors. Radiation was observed to increase cytotoxic CD8 immune infiltration in radiation sensitive tumors. In radiation resistant tumors, no significant difference was observed in either CD8 immune infiltration or tumor volume in treated tumors compared to controls (p = 0.63).

Conclusions: [89Zr]-CD8 ImmunoPET reveals significant increases in CD8 immune infiltration in a radiation sensitive model of TNBC and identifies an immune altered tumor immune microenvironment. Fractionated radiotherapy was also observed to significantly increase the infiltration of innate and adaptive immune cells.

Citation Format: Patrick N. Song, Shannon E. Lynch, Chloe T. DeMellier, Alessandro Mascioni, Jia Fang, Anna G. Sorace. CD8 ImmunoPET imaging identifies an immunogenically active tumor microenvironment following radiotherapy in primary TNBC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1120.