Callari Maurizio, Lembo Antonio, Bianchini Giampaolo, Musella Valeria, Cappelletti Vera, Gianni Luca, Daidone Maria Grazia, Provero Paolo
Department of Experimental Oncology and Molecular Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy.
Department of Molecular Biotechnology and Life Sciences, University of Turin, Turin, Italy.
PLoS One. 2014 Jan 29;9(1):e86511. doi: 10.1371/journal.pone.0086511. eCollection 2014.
Formalin fixed paraffin-embedded (FFPE) tumor specimens are the conventionally archived material in clinical practice, representing an invaluable tissue source for biomarkers development, validation and routine implementation. For many prospective clinical trials, this material has been collected allowing for a prospective-retrospective study design which represents a successful strategy to define clinical utility for candidate markers. Gene expression data can be obtained even from FFPE specimens with the broadly used Affymetrix HG-U133 Plus 2.0 microarray platform. Nevertheless, important major discrepancies remain in expression data obtained from FFPE compared to fresh-frozen samples, prompting the need for appropriate data processing which could help to obtain more consistent results in downstream analyses. In a publicly available dataset of matched frozen and FFPE expression data, the performances of different normalization methods and specifically designed Chip Description Files (CDFs) were compared. The use of an alternative CDFs together with fRMA normalization significantly improved frozen-FFPE sample correlations, frozen-FFPE probeset correlations and agreement of differential analysis between different tumor subtypes. The relevance of our optimized data processing was assessed and validated using two independent datasets. In this study we demonstrated that an appropriate data processing can significantly improve the reliability of gene expression data derived from FFPE tissues using the standard Affymetrix platform. Tools for the implementation of our data processing algorithm are made publicly available at http://www.biocut.unito.it/cdf-ffpe/.
福尔马林固定石蜡包埋（FFPE）肿瘤标本是临床实践中传统的存档材料，是生物标志物开发、验证和常规应用的宝贵组织来源。对于许多前瞻性临床试验，已收集了这种材料，从而实现了前瞻性-回顾性研究设计，这是确定候选标志物临床效用的成功策略。即使使用广泛使用的Affymetrix HG-U133 Plus 2.0微阵列平台，也可以从FFPE标本中获得基因表达数据。然而，与新鲜冷冻样本相比，从FFPE获得的表达数据仍存在重大差异，这促使需要进行适当的数据处理，以帮助在下游分析中获得更一致的结果。在一个公开可用的匹配冷冻和FFPE表达数据的数据集中，比较了不同归一化方法和专门设计的芯片描述文件（CDF）的性能。使用替代CDF与fRMA归一化一起可显著提高冷冻-FFPE样本相关性、冷冻-FFPE探针集相关性以及不同肿瘤亚型之间差异分析的一致性。使用两个独立数据集评估并验证了我们优化数据处理的相关性。在本研究中，我们证明了适当的数据处理可以显著提高使用标准Affymetrix平台从FFPE组织获得的基因表达数据的可靠性。我们的数据处理算法实施工具可在http://www.biocut.unito.it/cdf-ffpe/上公开获取。
