Quantifying Spatiotemporal Immune Dynamics Following Chemoradiation
Abstract
Purpose
Head and neck squamous cell carcinoma (HNSCC) is a clinically aggressive malignancy with heterogeneous responses to chemoradiation, driven in part by dynamic immune remodeling. While immune infiltration has been studied at different time points, the spatiotemporal organization and communication of immune cells across time following chemoradiation remain poorly defined. This study longitudinally characterizes immune infiltration and immune network topology following chemoradiation, linking early innate responses to adaptive immune coordination and endpoint immune states.
Methods
Using an aggressive MOC2 murine oral squamous cell carcinoma model, 25 CCR2+/- or mice (6–11 weeks) were inoculated with 30,000 tumor cells in the right buccal mucosa. At tumor volumes of ~50 mm³, mice received 8 Gy radiation with 5 mg/kg cisplatin. Tumors were harvested at days 1 (n=7), 3 (n=6), and 7 (n=5) post-treatment, and at humane endpoint (≥10 mm in any dimension; n=7). H&E tumor slides were digitized and underwent pathomic analysis. A Deep learning model was applied to detect, segment, and classify nuclei and generate spatial graphs, where nodes represent cells and edges encode spatial proximity. High-throughput pathomic features quantified immune infiltration, and immune network connectivity.
Results
Immune infiltration increased from day 1 to day 7, with immune-to–non-immune cell density rising from 4.14×10⁻⁸ to 3.79×10⁻⁷ cells/µm², followed by reduced immune presence at endpoint (1.79×10⁻⁸ cells/µm²). Tumors with higher immune infiltration were smaller (inverse exponential correlation; R2=0.963), consistent with enhanced antitumor immunity, whereas larger tumors exhibited diminished immune presence suggesting immune escape. Graph-based analysis demonstrated increasing K-core values through day 7, indicating enhanced immune network connectivity consistent with a transition from innate to adaptive immunity. Subsequent decline at endpoint suggested immune dysregulation and chronic inflammation.
Conclusion
Immune response to chemoradiation is topologically dynamic. Ongoing studies involve increasing sample size, incorporating immunohistochemistry and RNA sequencing to improve robustness and further define immune cell–specific interactions.