A Methodology for Utilization of Predictive Genomic Signatures in FFPE Samples

Purpose: Gene expression signatures developed to measure the activity of oncogenic signaling pathways have been used to dissect the heterogeneity of tumor samples and to predict sensitivity to various cancer drugs that target components of the relevant pathways, thus potentially identifying therapeutic options for subgroups of patients. To facilitate broad use, including in a clinical setting, the ability to generate data from formalin-fixed, paraffin-embedded (FFPE) tissues is essential. Experimental Design: Patterns of pathway activity in matched fresh-frozen and FFPE xenograft tumor samples were generated using the MessageAmp Premier methodology in combination with assays using Affymetrix arrays. Results generated were compared with those obtained from fresh-frozen samples using a standard Affymetrix assay. In addition, gene expression data from patient matched fresh-frozen and FFPE melanomas were also utilized to evaluate the consistency of predictions of oncogenic signaling pathway status. Results: Significant correlation of pathway activity predictions was observed between paired fresh-frozen and FFPE xenograft tumor samples. In addition, significant concordance of pathway activity predictions was also observed between patient matched fresh-frozen and FFPE melanomas. Conclusion: Reliable and consistent predictions of oncogenic pathway activities can be obtained from FFPE tumor tissue samples. The ability to reliably utilize FFPE patient tumor tissue samples for genomic analyses will lead to a better understanding of the biology of disease progression and, in the clinical setting, will provide tools to guide the choice of therapeutics to those most likely to be effective in treating a patient’s disease. 8 replicates of HMECs infected with adenovirus expressing GFP, 8 replicates of HMECs infected with adenovirus expressing RAS, 6 replicates of HMECs infected with adenovirus expressing MYC, 25 fresh-frozen melanoma xenografts, 25 FFPE melanoma xenografts, 6 FFPE human melanoma

【研究目的】 本研究中用于定量致癌信号通路活性的基因表达特征，已被用于解析肿瘤样本的异质性，并预测各类靶向相关通路组分的抗癌药物敏感性，从而有望为患者亚群筛选潜在治疗方案。为推动该技术的广泛应用（包括临床场景），从福尔马林固定石蜡包埋（formalin-fixed, paraffin-embedded, FFPE）组织中获取测序数据的能力至关重要。【实验设计】 本研究采用MessageAmp Premier试剂盒技术结合Affymetrix基因芯片（Affymetrix arrays）检测，对配对新鲜冷冻与FFPE异种移植肿瘤样本的通路活性模式进行表征；同时将所得结果与采用标准Affymetrix基因芯片检测的新鲜冷冻样本结果进行对比。此外，本研究还利用患者配对的新鲜冷冻与FFPE黑色素瘤样本的基因表达数据，评估致癌信号通路状态预测结果的一致性。【实验结果】 配对的新鲜冷冻与FFPE异种移植肿瘤样本的通路活性预测结果呈现显著相关性；此外，患者配对的新鲜冷冻与FFPE黑色素瘤样本的通路活性预测结果也表现出显著一致性。【研究结论】 从FFPE肿瘤组织样本中可获得可靠且一致的致癌通路活性预测结果。借助FFPE患者肿瘤组织样本开展可靠基因组分析的能力，将有助于我们更深入地解析疾病进展的生物学机制，并在临床场景中为筛选最有效的个体化治疗方案提供决策工具。本研究涉及的样本包括：8例表达绿色荧光蛋白（GFP）的腺病毒感染人乳腺上皮细胞（HMECs）重复样本、8例表达RAS的腺病毒感染HMECs重复样本、6例表达MYC的腺病毒感染HMECs重复样本、25例新鲜冷冻黑色素瘤异种移植样本、25例FFPE黑色素瘤异种移植样本以及6例FFPE人黑色素瘤样本。