Otolaryngology, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, USA.
Otolaryngology, Stanford University School of Medicine, Stanford, California, USA.
J Immunother Cancer. 2020 Sep;8(2). doi: 10.1136/jitc-2020-001048.
To explore lymphocyte infiltration as a potential mechanism behind CXCL14-mediated tumor growth suppression in oral cavity squamous cell carcinoma (OSCC).
We analyzed single cell RNA-sequencing (scRNA-seq) data from OSCC to identify expression changes among malignant cells in lymph nodes (LN) versus primary tumors. CXCL14 expression in murine OSCC cell lines was quantified using qRT-PCR. Short hairpin RNA knockdown of CXCL14 was performed in mouse oral cavity (MOC)1 cells, and CXCL14 overexpression was performed in MOC2 cells. Cells in each condition were injected into C57BL/6 mice with and without T cell depletion, and tumor volume was measured. At 30 days, tumors were dissociated and analyzed by flow cytometry for CD45CD3 T cells. CXCL14 expression was correlated with gene expression signatures of tumor infiltrating lymphocytes (TIL) in scRNA-seq data, as well as TCGA tumors.
scRNA-seq revealed CXCL14 as the most significantly downregulated gene among malignant cells in LNs relative to primary tumor, supporting a role in preventing invasion and/or metastasis. In a murine immunocompetent model, CXCL14 expression was higher in indolent MOC1 cells than in more aggressive MOC2 cells. Tumor growth was significantly increased by CXCL14 knockdown in MOC1 cells relative to control, with a corresponding decrease in TIL. In MOC2 cells, tumor growth was significantly reduced by CXCL14 overexpression relative to control and TIL were increased. Both effects were lost with T cell depletion. In a human tumor scRNA-seq cohort, we found that only malignant cell CXCL14, but not non-malignant cell or fibroblast CXCL14, was associated with TIL. Bulk CXCL14 from the TCGA cohort had no association with TIL.
Higher CXCL14 expression by tumor cells is associated with reduced tumor growth and increased TIL, supporting immune-mediated suppression of tumor growth in OSCC. Given that CXCL14 is downregulated in LN metastases compared with primary tumors, our data raise the possibility that CXCL14-mediated immune infiltration may discourage invasion and metastasis. In human scRNA-seq data, only malignant cell-specific CXCL14 was associated with TIL, suggesting a critical context-dependent effect of CXCL14 expression.
探讨淋巴细胞浸润作为 CXCL14 介导的口腔鳞状细胞癌 (OSCC) 肿瘤生长抑制潜在机制。
我们分析了 OSCC 的单细胞 RNA 测序 (scRNA-seq) 数据，以鉴定淋巴结 (LN) 与原发性肿瘤中恶性细胞的表达变化。使用 qRT-PCR 定量检测鼠 OSCC 细胞系中的 CXCL14 表达。在 MOC1 细胞中进行 CXCL14 的短发夹 RNA 敲低，在 MOC2 细胞中进行 CXCL14 的过表达。在有和没有 T 细胞耗竭的情况下，将每种条件下的细胞注射到 C57BL/6 小鼠中，并测量肿瘤体积。在 30 天时，将肿瘤解离并通过流式细胞术分析 CD45CD3 T 细胞。在 scRNA-seq 数据以及 TCGA 肿瘤中，CXCL14 的表达与肿瘤浸润淋巴细胞 (TIL) 的基因表达特征相关。
scRNA-seq 显示，与原发性肿瘤相比，LN 中的恶性细胞中 CXCL14 是下调最显著的基因，提示其在预防侵袭和/或转移中发挥作用。在鼠免疫活性模型中，惰性 MOC1 细胞中的 CXCL14 表达高于侵袭性更强的 MOC2 细胞。与对照相比，MOC1 细胞中 CXCL14 敲低导致肿瘤生长显著增加，同时 TIL 减少。在 MOC2 细胞中，与对照相比，CXCL14 过表达导致肿瘤生长显著减少，同时 TIL 增加。这两种作用都随着 T 细胞耗竭而丧失。在人类肿瘤 scRNA-seq 队列中，我们发现只有恶性细胞 CXCL14，而不是非恶性细胞或成纤维细胞 CXCL14，与 TIL 相关。来自 TCGA 队列的批量 CXCL14 与 TIL 无关。
肿瘤细胞中 CXCL14 的高表达与肿瘤生长减少和 TIL 增加相关，支持 OSCC 中免疫介导的肿瘤生长抑制。鉴于 LN 转移与原发性肿瘤相比，CXCL14 下调，我们的数据提出了 CXCL14 介导的免疫浸润可能阻止侵袭和转移的可能性。在人类 scRNA-seq 数据中，只有恶性细胞特异性 CXCL14 与 TIL 相关，这表明 CXCL14 表达具有关键的上下文依赖性效应。
