单区域样本评估肝细胞癌肿瘤免疫微环境的可靠性。
Reliability of a single-region sample to evaluate tumor immune microenvironment in hepatocellular carcinoma.
机构信息
Department of Medical Oncology, National Taiwan University Cancer Center, Taiwan; Department of Oncology, National Taiwan University Hospital, Taiwan; Graduate Institute of Oncology, School of Medicine, National Taiwan University, Taiwan.
Graduate Institute of Oncology, School of Medicine, National Taiwan University, Taiwan; Department of Medical Research, National Taiwan University Hospital, Taiwan.
出版信息
J Hepatol. 2020 Mar;72(3):489-497. doi: 10.1016/j.jhep.2019.09.032. Epub 2019 Oct 18.
BACKGROUND & AIMS: Intratumor heterogeneity has frequently been reported in patients with hepatocellular carcinoma (HCC). Thus, the reliability of single-region tumor samples for evaluation of the tumor immune microenvironment is also debatable. We conducted a prospective study to analyze the similarity in tumor immune microenvironments among different regions of a single tumor.
METHODS
Multi-region sampling was performed on newly resected tumors. The tumor immune microenvironment was evaluated by immunohistochemical staining of PD-L1, CD4, CD8, CD20, FoxP3, DC-LAMP (or LAMP3), CD68, MPO, and tertiary lymphoid structures (TLSs). PD-L1 expression was manually quantified according to the percentage of PD-L1-stained tumor or stromal cells. The densities (number/mm) of immune cells and the number of TLSs per sample were determined by whole-section counting. RNA-sequencing was applied in selected samples. Similarities in tumor immune microenvironments within each tumor were evaluated by multivariate Mahalanobis distance analyses.
RESULTS
Thirteen tumors were collected from 12 patients. The median diameter of tumors was 9 cm (range 3-16 cm). A median of 6 samples (range 3-12) were obtained from each tumor. Nine (69.2%) tumors exhibited uniform expression of PD-L1 in all regions of the tumor. Out of 13 tumors analyzed by immunohistochemical staining, 8 (61.5%) tumors displayed a narrow Mahalanobis distance for all regions within the tumor; while 8 (66.7%) of the 12 tumors analyzed by RNA-sequencing displayed a narrow Mahalanobis distance. Immunohistochemistry and RNA-sequencing had a high concordance rate (83.3%; 10 of 12 tumors) for the evaluation of similarities between tumor immune microenvironments within a tumor.
CONCLUSIONS
A single-region tumor sample might be reliable for the evaluation of tumor immune microenvironments in approximately 60-70% of patients with HCC.
LAY SUMMARY
Heterogeneity in the regional immune microenvironments of tumors has been reported in patients with hepatocellular carcinoma. This heterogeneity could be an obstacle when trying to reliably evaluate the immune microenvironment of an entire tumor using only a single-region tumor sample, which may be the only option in patients with more advanced disease. Our study utilized both immunohistochemical and transcriptomic analyses to demonstrate that a single-region sample is reliable for evaluation of tumor immune microenvironments in 60-70% of patients with hepatocellular carcinoma.
背景与目的
肝细胞癌（HCC）患者的肿瘤内异质性经常被报道。因此，单一区域肿瘤样本用于评估肿瘤免疫微环境的可靠性也存在争议。我们进行了一项前瞻性研究，以分析单个肿瘤内不同区域的肿瘤免疫微环境的相似性。
方法
对新切除的肿瘤进行多区域取样。通过 PD-L1、CD4、CD8、CD20、FoxP3、DC-LAMP（或 LAMP3）、CD68、MPO 和三级淋巴结构（TLSs）的免疫组织化学染色来评估肿瘤免疫微环境。根据 PD-L1 染色的肿瘤或基质细胞的百分比，手动量化 PD-L1 表达。通过全切片计数确定每个样本中免疫细胞的密度（每平方毫米的数量）和 TLSs 的数量。在选定的样本中应用 RNA 测序。通过多元马哈拉诺比斯距离分析评估每个肿瘤内的肿瘤免疫微环境的相似性。
结果
从 12 名患者中收集了 13 个肿瘤。肿瘤的中位直径为 9cm（范围 3-16cm）。每个肿瘤获得中位数为 6 个样本（范围 3-12 个）。9 个（69.2%）肿瘤的肿瘤在所有区域均表现出均匀的 PD-L1 表达。在 13 个用免疫组织化学染色分析的肿瘤中，8 个（61.5%）肿瘤的肿瘤内所有区域的马哈拉诺比斯距离均较窄；而在 12 个用 RNA 测序分析的肿瘤中，8 个（66.7%）肿瘤的马哈拉诺比斯距离较窄。免疫组织化学和 RNA 测序在评估肿瘤内肿瘤免疫微环境的相似性方面具有高一致性率（83.3%；12 个肿瘤中的 10 个）。
结论
对于 HCC 患者，约 60-70%的患者，单一区域的肿瘤样本可能足以评估肿瘤免疫微环境。
研究背景
在肝细胞癌患者中，肿瘤局部免疫微环境的异质性已被报道。当试图仅使用单个区域的肿瘤样本可靠地评估整个肿瘤的免疫微环境时，这种异质性可能成为一个障碍，而在疾病进展更严重的患者中，这可能是唯一的选择。我们的研究同时利用免疫组织化学和转录组分析，证明在 60-70%的肝细胞癌患者中，单一区域的样本可用于评估肿瘤免疫微环境。
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