Department of Data Science, Dana-Farber Cancer Institute, Boston, Massachusetts.
Department of Biostatistics, Harvard T.H. Chan School of Public Health, Boston, Massachusetts.
Clin Cancer Res. 2021 Sep 15;27(18):5049-5061. doi: 10.1158/1078-0432.CCR-20-3251. Epub 2020 Dec 15.
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.
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.
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.
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.
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。为了评估不同中心的肿瘤 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）进行了一项系统的协调研究。
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，并分发给三个中心进行 WES 和 RNA-seq 分析。此外，还使用了两个已知突变的 10 plex HapMap 细胞系池来评估 WES 平台的准确性。
当在使用 > 50×常见覆盖度的位点评估时，WES 平台在 HapMap 池上实现了 > 0.98 的高精度和 > 0.87 的召回率。非同义突变根据肿瘤样本聚类，在重复、中心和样本处理之间实现了 > 0.67 的特异性一致性指数。RNA 的 DV200 > 24%，作为一种潜在的预测序 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，超过该指标的同一肿瘤样本在重复、中心和样本处理运行之间可以稳健聚类，并且可以进行 HLA 分型、免疫浸润和免疫受体库推断。
CIMAC 合作实验室平台有效地生成了一致的 WES 和 RNA-seq 数据，并能够在 NCI CIMAC-CIDC 网络中对高度复杂的免疫肿瘤生物标志物数据进行稳健的跨试验比较和荟萃分析。
