Gene expression profiling of oral squamous cell carcinoma

Abstract BACKGROUND: Oral squamous cell carcinoma (OSCC) is a frequent neoplasm, which is usually aggressive and has unpredictable biological behavior and unfavorable prognosis. The comprehension of the molecular basis of this variability should lead to the development of targeted therapies as well as to improvements in specificity and sensitivity of diagnosis. RESULTS: Samples of primary OSCCs and their corresponding surgical margins were obtained from male patients during surgery and their gene expression profiles were screened using whole-genome microarray technology. Hierarchical clustering and Principal Components Analysis were used for data visualization and One-way Analysis of Variance was used to identify differentially expressed genes. Samples clustered mostly according to disease subsite, suggesting molecular heterogeneity within tumor stages. In order to corroborate our results, two publicly available datasets of microarray experiments were assessed. We found significant molecular differences between OSCC anatomic subsites concerning groups of genes presently or potentially important for drug development, including mRNA processing, cytoskeleton organization and biogenesis, metabolic process, cell cycle and apoptosis. CONCLUSION: Our results corroborate literature data on molecular heterogeneity of OSCCs. Differences between disease subsites and among samples belonging to the same TNM class highlight the importance of gene expression-based classification and challenge the development of targeted therapies. Nine tumor samples differing in their TNM classification and their respective surgical margins were used in this study. They were obtained from individual patients during tongue and floor of the mouth tumor resection surgery. Microarray experiments were carried out using the microarray platform CodeLink (GE Healthcare). This platform utilizes bioarrays consisting of 30-base, single pre-validated oligonucleotide probe per gene target. CodeLink Whole-Genome bioarrays, containing 55,000 human transcripts, were used for all experiments. Hybridization procedures strictly followed protocols provided by the manufacturer (GE Healthcare). A total of 11 arrays were hybridized in this study. Arrays were scanned following the recommended scanning procedure and settings for use with CodeLink bioarrays (GE Healthcare) on GenePix 4000B Array Scanner/GenePix Pro 4.0 software (Axon Instruments).

摘要：
背景：口腔鳞状细胞癌（Oral Squamous Cell Carcinoma, OSCC）是一种常见恶性肿瘤，通常具有侵袭性强、生物学行为难以预测且预后不佳的特点。阐明该肿瘤异质性的分子基础，有助于开发靶向治疗方案，同时提升诊断的特异性与灵敏度。
结果：本研究从接受手术治疗的男性患者体内获取原发性OSCC样本及其对应手术切缘组织，采用全基因组微阵列技术（whole-genome microarray）筛选其基因表达谱。通过层次聚类（Hierarchical Clustering）与主成分分析（Principal Components Analysis）进行数据可视化，运用单因素方差分析（One-way Analysis of Variance）筛选差异表达基因（differentially expressed genes）。样本聚类结果主要按照疾病发病亚部位进行分组，提示同一肿瘤分期内存在分子异质性。为验证本研究结果，我们评估了两个公开可用的微阵列实验数据集。本研究发现，OSCC不同解剖亚部位之间，在与药物开发当前或潜在相关的基因集上存在显著分子差异，这些基因集涵盖mRNA加工（mRNA processing）、细胞骨架组织与生物发生（cytoskeleton organization and biogenesis）、代谢过程、细胞周期以及细胞凋亡（apoptosis）等通路。
结论：本研究结果证实了现有文献中关于OSCC分子异质性的结论。不同疾病亚部位之间，以及同一TNM分期样本之间的表达差异，既凸显了基于基因表达分类的重要性，也对靶向治疗的开发提出了挑战。
本研究共纳入9份TNM分期不同的肿瘤样本及其对应手术切缘组织，均来自接受舌部及口底肿瘤切除术的患者。本研究采用CodeLink（通用电气医疗集团，GE Healthcare）微阵列平台开展实验，该平台使用的生物芯片每个基因靶点对应1条预先验证过的30碱基单寡核苷酸探针。本研究所有实验均使用包含55000个人类转录本的CodeLink全基因组生物芯片。杂交实验严格遵循制造商（通用电气医疗集团，GE Healthcare）提供的操作流程。本研究共完成11张芯片的杂交实验，按照CodeLink生物芯片推荐的扫描流程与参数，使用GenePix 4000B阵列扫描仪/GenePix Pro 4.0软件（Axon Instruments）对芯片进行扫描。