de Morais Everton Freitas, de Oliveira Lilianny Querino Rocha, Marques Cintia Eliza, Téo Fábio Haach, Rocha Gisele Vieira, Rodini Camila Oliveira, Gurgel Clarissa A, Salo Tuula, Graner Edgard, Coletta Ricardo D
Graduate Program in Oral Biology, School of Dentistry, University of Campinas, Piracicaba 13414-018, SP, Brazil.
Department of Oral Diagnosis, School of Dentistry, University of Campinas, Piracicaba 13414-018, SP, Brazil.
Cells. 2025 Aug 24;14(17):1311. doi: 10.3390/cells14171311.
Cisplatin resistance remains a major therapeutic challenge in oral squamous cell carcinoma (OSCC), leading to treatment failure and poor outcomes. This study aimed to generate and characterize cisplatin-resistant OSCC models to elucidate resistance mechanisms. Two resistant OSCC cell lines (SCC-9R and HSC-3R) were developed through gradual dose escalation. Parental and resistant cells were analyzed via RNA-seq and gene set enrichment analysis, and validated through RT-qPCR, Western blot, immunofluorescence, and gelatin zymography. Functional assays, including 2D and 3D migration and invasion models, assessed phenotypic changes. A multi-omics analysis revealed molecular alterations in resistant cells, including 305 differentially expressed genes (DEGs) in HSC-3R (187 upregulated) and 782 in SCC-9R (298 upregulated) versus parental lines, with enrichment for extracellular matrix organization ( < 0.001) and consistent epithelial-mesenchymal transition (EMT) activation ( < 0.001), demonstrated by the upregulation of ZEB1, ZEB2, Vimentin, and TWIST1, and E-cadherin suppression. Functional validation confirmed an aggressive phenotype, including increased migration ( < 0.05), invasion ( < 0.01), and elevated MMP-2 ( < 0.01) and MMP-9 ( < 0.001) activity. Findings were verified in 3D spheroid models. Overall, cisplatin resistance in OSCC involves EMT, inflammatory signaling, and metabolic adaptation. The consistency of these features across both models supports the robustness of this in vitro system and reveals targets for therapeutic intervention.
顺铂耐药仍然是口腔鳞状细胞癌（OSCC）治疗中的一个主要挑战，导致治疗失败和预后不良。本研究旨在建立并表征顺铂耐药的OSCC模型，以阐明耐药机制。通过逐步增加剂量培养出了两种耐药的OSCC细胞系（SCC - 9R和HSC - 3R）。通过RNA测序和基因集富集分析对亲代细胞和耐药细胞进行了分析，并通过逆转录定量聚合酶链反应（RT - qPCR）、蛋白质免疫印迹法、免疫荧光法和明胶酶谱法进行了验证。包括二维和三维迁移及侵袭模型在内的功能分析评估了表型变化。多组学分析揭示了耐药细胞中的分子改变，与亲代细胞系相比，HSC - 3R中有305个差异表达基因（DEG）（187个上调），SCC - 9R中有782个（298个上调），细胞外基质组织富集（<0.001），并且一致的上皮 - 间质转化（EMT）激活（<0.001），这通过锌指蛋白E盒结合因子1（ZEB1）、锌指蛋白E盒结合因子2（ZEB2）、波形蛋白和 Twist 家族转录因子1（TWIST1）的上调以及E - 钙黏蛋白的抑制得以证明。功能验证证实了侵袭性表型，包括迁移增加（<0.05）、侵袭增加（<0.01）以及基质金属蛋白酶2（MMP - 2）（<0.01）和基质金属蛋白酶9（MMP - 9）活性升高（<0.001）。这些发现在三维球体模型中得到了验证。总体而言，OSCC中的顺铂耐药涉及EMT、炎症信号传导和代谢适应。这两种模型中这些特征的一致性支持了该体外系统的稳健性，并揭示了治疗干预的靶点。
