The potent immunostimulatory effects of toll-like receptor 8 (TLR8) agonism in combination with PD-1 blockade have resulted in various preclinical investigations, yet the mechanism of action in humans remains unknown. To decipher the combinatory mode of action of TLR8 agonism and PD-1 blockade, we employed a unique, open-label, phase 1b pre-operative window of opportunity clinical trial (NCT03906526) in head and neck squamous cell carcinoma (HNSCC) patients. Matched pre- and post-treatment tumor biopsies from the same lesion were obtained. We used single-cell RNA sequencing and custom multiplex staining to leverage the unique advantage of same-lesion longitudinal sampling. Patients receiving dual TLR8 agonism and anti-PD-1 blockade exhibited marked upregulation of innate immune effector genes and cytokines, highlighted by increased CLEC9A+ dendritic cell and CLEC7A/SYK expression. This was revealed via comparison with a previous cohort from an anti-PD-1 blockade monotherapy single-cell RNA sequencing study. Furthermore, in dual therapy patients, post-treatment mature dendritic cells increased in adjacency to CD8+ T-cells. Increased tumoral cytotoxic T-lymphocyte densities and expanded CXCL13+CD8+ T-cell populations were observed in responders, with increased tertiary lymphoid structures (TLSs) across all three patients. This study provides key insights into the mode of action of TLR8 agonism and anti-PD-1 blockade immune targeting in HNSCC patients.

中文翻译：

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树突状细胞效应机制和肿瘤免疫微环境浸润定义了 TLR8 调节和 PD-1 阻断


Toll 样受体 8 (TLR8) 激动剂与 PD-1 阻断相结合的有效免疫刺激作用已引起各种临床前研究，但其在人体中的作用机制仍不清楚。为了破译 TLR8 激动和 PD-1 阻断的组合作用模式，我们在头颈鳞状细胞癌 (HNSCC) 患者中采用了一项独特的、开放标签的 1b 期术前机会窗口临床试验 (NCT03906526)。获得来自同一病灶的匹配的治疗前和治疗后肿瘤活检。我们使用单细胞 RNA 测序和定制多重染色来利用同一病变纵向采样的独特优势。接受 TLR8 双重激动和抗 PD-1 阻断的患者表现出先天免疫效应基因和细胞因子的显着上调，其中 CLEC9A+ 树突状细胞和 CLEC7A/SYK 表达增加突出显示。这是通过与先前的抗 PD-1 阻断单一疗法单细胞 RNA 测序研究的队列进行比较得出的。此外，在双重治疗患者中，治疗后与 CD8+ T 细胞相邻的成熟树突状细胞有所增加。在应答者中观察到肿瘤细胞毒性 T 淋巴细胞密度增加和 CXCL13+CD8+ T 细胞群扩大，所有三名患者的三级淋巴结构 (TLS) 均增加。这项研究为 TLR8 激动剂和抗 PD-1 阻断免疫靶向在 HNSCC 患者中的作用模式提供了重要见解。