丝氨酸蛋白酶抑制剂 Kazal 型 5（SPINK5）通过免疫活性抑制食管鳞状细胞癌转移。
SPINK5 inhibits esophageal squamous cell carcinoma metastasis via immune activity.
机构信息
Department of Oncology, Jiangyin People's Hospital, Jiangyin Clinical College of Xuzhou Medical University, Jiangyin Hospital Affiliated to Nantong University, Jiangyin, Jiangsu, China.
The Health Supervision Institute of Suzhou, Suzhou, Jiangsu, China.
出版信息
J Gene Med. 2024 Mar;26(3):e3667. doi: 10.1002/jgm.3667.
BACKGROUND
Esophageal squamous cell carcinoma (ESCC) is a predominant subtype of esophageal cancer with relatively high mortality worldwide. Serine peptidase inhibitor Kazal-type 5 (SPINK5) is reported to be downregulated in ESCC. However, its explicit role in ESCC remains further investigation.
METHODS
The tumor tissues and adjacent non-cancerous tissues were obtained from 196 patients with ESCC for the determination of SPINK5 mRNA levels. Additionally, the relationship between SPINK5 mRNA levels and clinicopathological features of ESCC patients was explored. The effects of SPINK5 on the invasion and migration of ESCC cells were assessed using Transwell assays. Furthermore, SPINK5 mRNA and LEKTI protein were measured in ESCC cell lines after treatment with poly (I:C), lipopolysaccharide (LPS) or unmethylated CpG DNA. Moreover, the correlation between expression of SPINK5 and nuclear factor-kappa B (NF-κB) signaling pathway-related genes was analyzed in the TCGA-ESCC cohort, and the effects of SPINK5 on NF-κB transcription was analyzed using a luciferase reporter gene assay. Finally, the correlations between SPINK5 and infiltration of immune cells, immune scores, stromal scores and ESTIMATE (i.e., Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data) scores were explored.
RESULTS
SPINK5 mRNA levels were downregulated in tumor tissues, which was significantly correlated with higher lymph node metastases. Overexpressed SPINK5 inhibited cell invasion and migration in ESCC cell lines. Mechanistically, LPS-induced activation of Toll-like receptor 4 (TLR4) decreased SPINK5 mRNA and LEKTI in KYSE150 and KYSE70 cells. Spearman correlation analysis revealed that SPINK5 mRNA was significantly negatively correlated with a total of seven NF-κB signaling pathway-related genes in TCGA-ESCC patients. Moreover, downregulation of SPINK5 increased and upregulation of SPINK5 decreased the activity of the NF-κB promoter in HEK293T cells. Finally, immune cells infiltration analysis revealed that SPINK5 was significantly correlated with the infiltration of various immune cells, stromal scores, immune scores and ESTIMATE scores.
CONCLUSIONS
SPINK5 plays critical roles in the TLR4/NF-κB pathway and immune cells infiltration, which might contribute to the ESCC metastasis. The findings of the present study may provide a promising biomarker for the diagnosis and treatment of esophageal squamous cell carcinoma.
背景
食管鳞状细胞癌（ESCC）是一种主要的食管癌亚型，在全球范围内死亡率相对较高。丝氨酸肽酶抑制剂 Kazal 型 5（SPINK5）在 ESCC 中被报道下调。然而，其在 ESCC 中的明确作用仍需进一步研究。
方法
从 196 名 ESCC 患者中获得肿瘤组织和相邻非癌组织，以确定 SPINK5 mRNA 水平。此外，还探讨了 SPINK5 mRNA 水平与 ESCC 患者临床病理特征的关系。通过 Transwell 测定评估 SPINK5 对 ESCC 细胞侵袭和迁移的影响。此外，用多聚（I：C）、脂多糖（LPS）或未甲基化 CpG DNA 处理 ESCC 细胞系后，测量 SPINK5 mRNA 和 LEKTI 蛋白。此外，在 TCGA-ESCC 队列中分析了 SPINK5 表达与核因子-κB（NF-κB）信号通路相关基因之间的相关性，并通过荧光素酶报告基因测定分析了 SPINK5 对 NF-κB 转录的影响。最后，探讨了 SPINK5 与免疫细胞浸润、免疫评分、基质评分和 ESTIMATE（即使用表达数据估计恶性肿瘤组织中的基质和免疫细胞）评分之间的相关性。
结果
肿瘤组织中 SPINK5 mRNA 水平下调，与淋巴结转移较高显著相关。过表达 SPINK5 抑制 ESCC 细胞系的细胞侵袭和迁移。机制上，LPS 诱导的 Toll 样受体 4（TLR4）激活降低了 KYSE150 和 KYSE70 细胞中的 SPINK5 mRNA 和 LEKTI。Spearman 相关分析显示，TCGA-ESCC 患者的 SPINK5 mRNA 与 NF-κB 信号通路相关基因总数的七个基因呈显著负相关。此外，下调 SPINK5 增加，上调 SPINK5 减少 NF-κB 启动子在 HEK293T 细胞中的活性。最后，免疫细胞浸润分析表明，SPINK5 与各种免疫细胞、基质评分、免疫评分和 ESTIMATE 评分的浸润显著相关。
结论
SPINK5 在 TLR4/NF-κB 通路和免疫细胞浸润中发挥关键作用，可能有助于 ESCC 转移。本研究的结果可为食管鳞状细胞癌的诊断和治疗提供有前途的生物标志物。
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