采用钻取和激光显微切割技术对肿瘤组织进行肿瘤细胞富集，用于胰腺导管腺癌的蛋白质组学和基因组学分析。

Neoplastic cell enrichment of tumor tissues using coring and laser microdissection for proteomic and genomic analyses of pancreatic ductal adenocarcinoma.

作者信息

Li Qing Kay, Hu Yingwei, Chen Lijun, Schnaubelt Michael, Cui Zhou Daniel, Li Yize, Lu Rita Jui-Hsien, Thiagarajan Mathangi, Hostetter Galen, Newton Chelsea J, Jewell Scott D, Omenn Gil, Robles Ana I, Mesri Mehdi, Bathe Oliver F, Zhang Bing, Ding Li, Hruban Ralph H, Chan Daniel W, Zhang Hui

机构信息

Department of Pathology, the Johns Hopkins University, 400 N Broadway, Smith Bldg Rm 4011, Baltimore, MD, 21231, USA.

Department of Oncology, Sidney Kimmel Cancer Center, Johns Hopkins Medical Institutions, 600 N. Wolfe Street, Baltimore, MD, USA.

出版信息

Clin Proteomics. 2022 Oct 20;19(1):36. doi: 10.1186/s12014-022-09373-x.

DOI:10.1186/s12014-022-09373-x

PMID:36266629

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC9583468/

Abstract

BACKGROUND

The identification of differentially expressed tumor-associated proteins and genomic alterations driving neoplasia is critical in the development of clinical assays to detect cancers and forms the foundation for understanding cancer biology. One of the challenges in the analysis of pancreatic ductal adenocarcinoma (PDAC) is the low neoplastic cellularity and heterogeneous composition of bulk tumors. To enrich neoplastic cells from bulk tumor tissue, coring, and laser microdissection (LMD) sampling techniques have been employed. In this study, we assessed the protein and KRAS mutation changes associated with samples obtained by these enrichment techniques and evaluated the fraction of neoplastic cells in PDAC for proteomic and genomic analyses.

METHODS

Three fresh frozen PDAC tumors and their tumor-matched normal adjacent tissues (NATs) were obtained from three sampling techniques using bulk, coring, and LMD; and analyzed by TMT-based quantitative proteomics. The protein profiles and characterizations of differentially expressed proteins in three sampling groups were determined. These three PDACs and samples of five additional PDACs obtained by the same three sampling techniques were also subjected to genomic analysis to characterize KRAS mutations.

RESULTS

The neoplastic cellularity of eight PDACs ranged from less than 10% to over 80% based on morphological review. Distinctive proteomic patterns and abundances of certain tumor-associated proteins were revealed when comparing the tumors and NATs by different sampling techniques. Coring and bulk tissues had comparable proteome profiles, while LMD samples had the most distinct proteome composition compared to bulk tissues. Further genomic analysis of bulk, cored, or LMD samples demonstrated that KRAS mutations were significantly enriched in LMD samples while coring was less effective in enriching for KRAS mutations when bulk tissues contained a relatively low neoplastic cellularity.

CONCLUSIONS

In addition to bulk tissues, samples from LMD and coring techniques can be used for proteogenomic studies. The greatest enrichment of neoplastic cellularity is obtained with the LMD technique.

摘要

背景

鉴定差异表达的肿瘤相关蛋白以及驱动肿瘤形成的基因组改变，对于开发检测癌症的临床检测方法至关重要，也是理解癌症生物学的基础。胰腺导管腺癌（PDAC）分析中的一个挑战是大块肿瘤的肿瘤细胞含量低且组成异质性。为了从大块肿瘤组织中富集肿瘤细胞，已采用了取芯和激光显微切割（LMD）采样技术。在本研究中，我们评估了通过这些富集技术获得的样本中相关的蛋白质和KRAS突变变化，并评估了PDAC中用于蛋白质组学和基因组分析的肿瘤细胞比例。

方法

使用大块、取芯和LMD三种采样技术从三个新鲜冷冻的PDAC肿瘤及其肿瘤匹配的正常相邻组织（NATs）中获取样本，并通过基于TMT的定量蛋白质组学进行分析。确定了三个采样组中差异表达蛋白质的蛋白质谱和特征。这三个PDAC以及通过相同三种采样技术获得的另外五个PDAC的样本也进行了基因组分析，以表征KRAS突变。

结果

根据形态学评估，八个PDAC的肿瘤细胞含量范围从不到10%到超过80%。通过不同采样技术比较肿瘤和NATs时，揭示了某些肿瘤相关蛋白的独特蛋白质组模式和丰度。取芯组织和大块组织具有可比的蛋白质组谱，而与大块组织相比，LMD样本具有最独特的蛋白质组组成。对大块、取芯或LMD样本的进一步基因组分析表明，当大块组织的肿瘤细胞含量相对较低时，LMD样本中KRAS突变显著富集，而取芯在富集KRAS突变方面效果较差。