Radiotherapy synergizes with CD24 blockade to initiate macrophage-driven systemic antitumor immunity in non–small cell lung cancer

BACKGROUND: Radiotherapy (RT) has been shown to elicit antitumor immune responses, yet it rarely induces durable systemic immunity. The identification of immune checkpoints that constrain RT-induced immune activation remains an unmet clinical need. METHODS: Using genetic deletion, pharmacologic blockade, syngeneic bilateral and orthotopic murine non-small cell lung cancer (NSCLC) models, and single-cell RNA sequencing, we dissected the role of CD24 in RT-induced immune responses. RESULTS: CD24 was overexpressed across NSCLC stages with stage-dependent bidirectional prognostic significance. CD24 exerted no tumor-intrinsic proliferative effects in vitro but drove macrophage-dependent immune evasion in vivo. Combined RT and CD24 blockade synergistically remodeled the tumor microenvironment-augmenting M1-polarised macrophages while suppressing M2 macrophages, regulatory T cells, and PMN-MDSCs-and induced reproducible abscopal responses across bilateral, orthotopic, and metastatic models regardless of radiation modality. Macrophage depletion abolished both local and distant tumor control, whereas CD8⁺ T cell depletion caused only partial attenuation, establishing macrophages as essential initiators of systemic immunity. Single-cell transcriptomics revealed that RT-induced CCL2 recruits peripheral CCR2⁺ monocytes that CD24 blockade reprograms into inflammatory macrophages; CCL2 neutralization completely abrogated the abscopal effect. Incorporating PD-1 blockade further amplified effector T cell responses and conferred durable tumor-specific immune memory. CONCLUSIONS: CD24 is a critical innate immune checkpoint restraining RT-induced systemic antitumor immunity. Combined RT, CD24 blockade, and PD-1 inhibition drives a macrophage-initiated, CCL2-CCR2-dependent innate-to-adaptive immune cascade, providing a mechanistic rationale for this combination strategy in NSCLC.