Purpose: Whole-exome (WES) and RNA sequencing (RNA-seq) are key components of cancer immunogenomic analyses. To evaluate the consistency of tumor WES and RNA-seq profiling platforms across different centers, the Cancer Immune Monitoring and Analysis Centers (CIMAC) and the Cancer Immunologic Data Commons (CIDC) conducted a systematic harmonization study. Experimental Design: DNA and RNA were centrally extracted from fresh frozen and formalin-fixed paraffin-embedded non–small cell lung carcinoma tumors and distributed to three centers for WES and RNA-seq profiling. In addition, two 10-plex HapMap cell line pools with known mutations were used to evaluate the accuracy of the WES platforms. Results: The WES platforms achieved high precision (> 0.98) and recall (> 0.87) on the HapMap pools when evaluated on loci using > 50× common coverage. Nonsynonymous mutations clustered by tumor sample, achieving an index of specific agreement above 0.67 among replicates, centers, and sample processing. A DV200 > 24% for RNA, as a putative presequencing RNA quality control (QC) metric, was found to be a reliable threshold for generating consistent expression readouts in RNA-seq and NanoString data. MedTIN > 30 was likewise assessed as a reliable RNA-seq QC metric, above which samples from the same tumor across replicates, centers, and sample processing runs could be robustly clustered and HLA typing, immune infiltration, and immune repertoire inference could be performed. Conclusions: The CIMAC collaborating laboratory platforms effectively generated consistent WES and RNA-seq data and enable robust cross-trial comparisons and meta-analyses of highly complex immuno-oncology biomarker data across the NCI CIMAC-CIDC Network.

中文翻译：

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CIMAC-CIDC 网络肿瘤 DNA 和 RNA 测序平台的跨站点一致性评估


目的：全外显子组 (WES) 和 RNA 测序 (RNA-seq) 是癌症免疫基因组分析的关键组成部分。为了评估不同中心之间肿瘤 WES 和 RNA-seq 分析平台的一致性，癌症免疫监测和分析中心 (CIMAC) 和癌症免疫数据共享中心 (CIDC) 进行了系统的协调研究。实验设计：从新鲜冷冻和福尔马林固定石蜡包埋的非小细胞肺癌肿瘤中集中提取 DNA 和 RNA，并分发到三个中心进行 WES 和 RNA-seq 分析。此外，还使用两个具有已知突变的 10 重 HapMap 细胞系库来评估 WES 平台的准确性。结果：当使用 > 50× 共同覆盖度对位点进行评估时，WES 平台在 HapMap 池上实现了高精度 (> 0.98) 和召回率 (> 0.87)。非同义突变按肿瘤样本聚集，在重复、中心和样本处理之间实现了 0.67 以上的特定一致性指数。 RNA 的 DV200 > 24% 作为假定的预测序 RNA 质量控制 (QC) 指标，被发现是在 RNA-seq 和 NanoString 数据中生成一致表达读数的可靠阈值。 MedTIN > 30 同样被评估为可靠的 RNA-seq QC 指标，高于该指标，来自同一肿瘤的重复、中心和样本处理运行的样本可以稳健地聚类，并且可以进行 HLA 分型、免疫浸润和免疫组库推断。结论：CIMAC 合作实验室平台有效地生成了一致的 WES 和 RNA-seq 数据，并能够对整个 NCI CIMAC-CIDC 网络中高度复杂的免疫肿瘤学生物标志物数据进行稳健的交叉试验比较和荟萃分析。