Background: Most immunologically “cold” tumors do not respond durably to checkpoint blockade because tumor antigen (TA) release and presentation are insufficient to prime effective T-cell immunity. While prior work demonstrated synergy between cisplatin and a TLR7/8/9 agonist (CR108) in 4T1 tumors, the underlying mechanism—particularly whether chemotherapy functions as a broad antigen-releasing agent enabling TLR-driven immune amplification—remained undefined. Methods: Using murine models of breast (4T1), melanoma (B16-F10), and colorectal cancer (CT26), we tested multiple chemotherapeutic classes combined with CR108. We quantified intratumoral and systemic soluble TAs, antigen presentation and cross-priming by antigen-presenting cells, tumor-infiltrating lymphocytes, and cytokine production by flow cytometry/ICS. T-cell receptor β (TCRβ) repertoire dynamics in tumor-draining lymph nodes were profiled to assess amplitude and breadth. Tumor microenvironment remodeling was analyzed, and public datasets (e.g., TCGA basal-like breast cancer) were interrogated for expression of genes linked to TA generation/processing and peptide loading. Results: Using cisplatin + CR108 in 4T1 as a benchmark, we demonstrate that diverse chemotherapies—especially platinum agents—broadly increase the repertoire of soluble tumor antigens available for immune recognition. Across regimens, chemotherapy combined with CR108 increased T-cell recognition of candidate TAs and enhanced IFN-γ+CD8+responses, with platinum agents producing the largest expansions in soluble TAs. TCRβ sequencing revealed increased clonal amplitude without loss of repertoire breadth, indicating focused yet diverse antitumor T-cell expansion. Notably, therapeutic efficacy was not predicted by canonical damage-associated molecular pattern (DAMP) signatures but instead correlated with antigen availability and processing capacity. In human basal-like breast cancer, higher expression of genes involved in TA generation and antigen processing/presentation correlated with improved survival. Conclusions: Our findings establish an antigen-centric mechanism underlying chemo–TLR agonist synergy: chemotherapy liberates a broadened repertoire of tumor antigens, which CR108 then leverages via innate immune activation to drive potent, T-cell-mediated antitumor immunity. This framework for rational selection of chemotherapy partners for TLR7/8/9 agonism and support clinical evaluation to convert “cold” tumors into immunologically responsive disease.
背景：大多数免疫学意义上的“冷”肿瘤对检查点阻断疗法无法产生持久应答，原因在于肿瘤抗原（TA）的释放与呈递不足以启动有效的T细胞免疫。尽管先前研究已证实顺铂与TLR7/8/9激动剂（CR108）在4T1肿瘤模型中具有协同作用，但其内在机制——尤其是化疗是否作为广谱抗原释放剂从而促进TLR驱动的免疫放大——尚未明确。 方法：本研究采用小鼠乳腺癌（4T1）、黑色素瘤（B16-F10）及结直肠癌（CT26）模型，测试了多类化疗药物与CR108的联合疗效。通过流式细胞术/细胞内因子染色技术，定量分析了瘤内及全身可溶性TA水平、抗原呈递细胞的抗原呈递与交叉呈递能力、肿瘤浸润淋巴细胞数量及细胞因子产生情况。对肿瘤引流淋巴结中T细胞受体β（TCRβ）谱系动态进行测序，以评估T细胞扩增的幅度与广度。同时分析肿瘤微环境重塑特征，并基于公共数据库（如TCGA基底样乳腺癌数据集）探究TA生成/加工及肽段装载相关基因的表达模式。 结果：以4T1模型中顺铂+CR108方案为基准，研究发现多种化疗药物（尤其是铂类药物）能广泛增加可供免疫识别的可溶性肿瘤抗原谱系。在所有治疗方案中，化疗联合CR108均能增强T细胞对候选TA的识别能力，并提升IFN-γ+CD8+ T细胞应答水平，其中铂类药物诱导的可溶性TA扩增最为显著。TCRβ测序显示克隆扩增幅度增加而谱系广度未受损失，表明抗肿瘤T细胞呈现聚焦性且多样化的扩增特征。值得注意的是，经典损伤相关分子模式（DAMP）特征无法预测疗效，而抗原可及性与加工能力则与疗效显著相关。在人类基底样乳腺癌中，TA生成及抗原加工/呈递相关基因的高表达与患者生存改善存在关联。 结论：本研究揭示了化疗-TLR激动剂协同作用的核心机制：化疗释放广谱肿瘤抗原库，CR108则通过天然免疫激活作用驱动强效的T细胞介导抗肿瘤免疫。该研究为TLR7/8/9激动剂的化疗联用方案选择提供了以抗原为中心的理论框架，并为将“冷”肿瘤转化为免疫应答性疾病的临床转化研究提供依据。
肿瘤(癌症)患者之家