LaFranzo Natalie A, Flanagan Kevin C, Quintanilha Danielle
Cofactor Genomics;
Cofactor Genomics.
J Vis Exp. 2020 Feb 25(156). doi: 10.3791/60645.
Immunotherapies show promise in the treatment of oncology patients, but complex heterogeneity of the tumor microenvironment makes predicting treatment response challenging. The ability to resolve the relative populations of immune cells present in and around the tumor tissue has been shown to be clinically-relevant to understanding response, but is limited by traditional techniques such as flow cytometry and immunohistochemistry (IHC), due the large amount of tissue required, lack of accurate cell type markers, and many technical and logistical hurdles. One assay (e.g., the ImmunoPrism Immune Profiling Assay) overcomes these challenges by accommodating both small amounts of RNA and highly degraded RNA, common features of RNA extracted from clinically archived solid tumor tissue. The assay is accessed via a reagent kit and cloud-based informatics that provides an end-to-end quantitative, high-throughput immuno-profiling solution for Illumina sequencing platforms. Researchers start with as few as two sections of formalin-fixed paraffin-embedded (FFPE) tissue or 20-40 ng of total RNA (depending on sample quality), and the protocol generates an immune profile report quantifying eight immune cell types and ten immune escape genes, capturing a complete view of the tumor microenvironment. No additional bioinformatic analysis is required to make use of the resulting data. With the appropriate sample cohorts, the protocol may also be used to identify statistically significant biomarkers within a patient population of interest.
免疫疗法在肿瘤患者治疗中显示出前景,但肿瘤微环境的复杂异质性使得预测治疗反应具有挑战性。已证明解析肿瘤组织内部及周围存在的免疫细胞相对数量的能力在临床上与理解治疗反应相关,但受传统技术(如流式细胞术和免疫组织化学(IHC))的限制,原因在于所需组织量巨大、缺乏准确的细胞类型标记以及诸多技术和后勤障碍。一种检测方法(例如免疫棱镜免疫分析检测法)通过适应少量RNA和高度降解的RNA克服了这些挑战,而这是从临床存档的实体瘤组织中提取的RNA的常见特征。该检测方法可通过试剂试剂盒和基于云的信息学访问,为Illumina测序平台提供端到端的定量、高通量免疫分析解决方案。研究人员从少至两片福尔马林固定石蜡包埋(FFPE)组织或20 - 40纳克总RNA(取决于样本质量)开始,该方案会生成一份免疫分析报告,对八种免疫细胞类型和十种免疫逃逸基因进行定量,全面呈现肿瘤微环境。利用所得数据无需额外的生物信息学分析。对于合适的样本队列,该方案还可用于在感兴趣的患者群体中识别具有统计学意义的生物标志物。