Transcriptomics-based analysis of key genes and potential mechanism and therapeutic agents in cadmium-induced esophageal squamous cell carcinoma progression

Chronic cadmium exposure is linked to esophageal squamous cell carcinoma (ESCC) progression, treatment resistance, and poor prognosis, but its molecular mechanisms remain unclear. Based on whole-transcriptome sequencing (RNA-seq) on chronic cadmium-treated (CCT) and untreated human EC109 ESCC cells, we performed lncRNA-mRNA co-expression and protein-protein interaction (PPI) network analyses to screen the hub genes related to cadmium exposure. Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses depicted that these genes were mainly enriched in nucleocytoplasmic transport, ferroptosis, Huntington disease, and cellular senescence. On the basis of lncRNA-miRNA-mRNA potential interaction, 3 lncRNAs (AC107068.1, TTN-AS1, MAPKAPK5-AS1) and 10 mRNAs (IPO5, NUDCD1, OSTM1, CCNB1, FANCD2, TFRC, POLR2B, HTT, NUP43, NBN) were identified for competitive endogenous RNA (ceRNA) network construction. Connectivity map (CMap) analysis for the target mRNAs showed that JAK3-inhibitor-I was the most promising therapeutic drug for cadmium-induced ESCC progression. The prognostic value and abnormal expression of the candidate mRNAs were then validated in TCGA cohorts, cadmium-treated cancer cells and 41 ESCC specimens. Moreover, migration and invasion assays were performed to assess the effects of the identified gene on cell malignant phenotypes. As a result, IPO5 was identified as the pivotal gene and MAPKAPK5-AS1-hsa-miR-379-5p-IPO5 was deemed as a potential ceRNA regulatory mechanism for cadmium carcinogenesis in ESCC. Using TIMER and EPIC algorithms, IPO5 was correlated with increased infiltration of CD4+ T cells and macrophages, while negatively associated with CD8+ T cells and NK cells in ESCC tissues. Collectively, our study provides valuable information in understanding the molecular mechanisms involved in cadmium-induced ESCC progression and treatment resistance. Furthermore, we predict potential agents for cadmium carcinogenicity prevention and treatment.

慢性镉暴露与食管鳞状细胞癌（esophageal squamous cell carcinoma, ESCC）的进展、治疗耐药及不良预后密切相关，但其分子机制迄今尚未明确。本研究以经慢性镉处理（chronic cadmium-treated, CCT）及未处理的人EC109食管鳞状细胞癌细胞为研究对象，开展全转录组测序（RNA-seq），并通过长链非编码RNA（long non-coding RNA, lncRNA）-mRNA共表达分析与蛋白质-蛋白质相互作用（protein-protein interaction, PPI）网络分析，筛选出与镉暴露相关的枢纽基因。随后，通过基因本体论（Gene Ontology, GO）及京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）通路富集分析，发现上述枢纽基因主要富集于核质运输、铁死亡、亨廷顿病及细胞衰老等生物学通路。基于lncRNA-微小RNA（microRNA, miRNA）-mRNA的潜在相互作用关系，本研究筛选出3种lncRNA（AC107068.1、TTN-AS1、MAPKAPK5-AS1）与10种mRNA（IPO5、NUDCD1、OSTM1、CCNB1、FANCD2、TFRC、POLR2B、HTT、NUP43、NBN）用于构建竞争性内源RNA（competing endogenous RNA, ceRNA）网络。针对靶mRNA的连通性图谱（Connectivity map, CMap）分析显示，JAK3-inhibitor-I是治疗镉诱导型食管鳞状细胞癌进展的最具潜力的候选药物。本研究随后在癌症基因组图谱（The Cancer Genome Atlas, TCGA）队列、镉处理的癌细胞及41例食管鳞状细胞癌标本中，验证了候选mRNA的异常表达及其预后价值。此外，本研究通过迁移与侵袭实验，评估了所鉴定基因对细胞恶性表型的影响。研究结果显示，IPO5被鉴定为核心枢纽基因，而MAPKAPK5-AS1-hsa-miR-379-5p-IPO5调控轴被视为镉诱导食管鳞状细胞癌发生的潜在ceRNA调控机制。借助TIMER与EPIC算法分析，本研究发现IPO5在食管鳞状细胞癌组织中与CD4+ T细胞及巨噬细胞的浸润水平呈正相关，而与CD8+ T细胞及自然杀伤细胞（natural killer cell, NK细胞）的浸润水平呈负相关。综上，本研究为阐明镉诱导食管鳞状细胞癌进展及治疗耐药的分子机制提供了宝贵的理论参考。此外，本研究还预测了可用于预防及治疗镉致癌作用的潜在候选药物。