Background Human plasma contains RNA transcripts released by multiple cell types within the body. Single-cell transcriptomic analysis allows the cellular origin of circulating RNA molecules to be elucidated at high resolution and has been successfully utilized in the pregnancy context. We explored the application of a similar approach to develop plasma RNA markers for cancer detection. Methods Single-cell RNA sequencing was performed to decipher transcriptomic profiles of single cells from hepatocellular carcinoma (HCC) samples. Cell-type-specific transcripts were identified and used for deducing the cell-type-specific gene signature (CELSIG) scores of plasma RNA from patients with and without HCC. Results Six major cell clusters were identified, including hepatocyte-like, cholangiocyte-like, myofibroblast, endothelial, lymphoid, and myeloid cell clusters based on 4 HCC tumor tissues as well as their paired adjacent nontumoral tissues. The CELSIG score of hepatocyte-like cells was significantly increased in preoperative plasma RNA samples of patients with HCC (n = 14) compared with non-HCC participants (n = 49). The CELSIG score of hepatocyte-like cells declined in plasma RNA samples of patients with HCC within 3 days after tumor resection. Compared with the discriminating power between patients with and without HCC using the abundance of ALB transcript in plasma [area under curve (AUC) 0.72)], an improved performance (AUC: 0.84) was observed using the CELSIG score. The hepatocyte-specific transcript markers in plasma RNA were further validated by ddPCR assays. The CELSIG scores of hepatocyte-like cell and cholangiocyte trended with patients’ survival. Conclusions The combination of single-cell transcriptomic analysis and plasma RNA sequencing represents an approach for the development of new noninvasive cancer markers.

中文翻译：

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用于肝细胞癌 RNA 液体活检的单细胞和血浆 RNA 测序

背景 人血浆含有体内多种细胞类型释放的 RNA 转录物。单细胞转录组分析允许以高分辨率阐明循环 RNA 分子的细胞来源，并已成功用于妊娠环境。我们探索了应用类似方法开发用于癌症检测的血浆 RNA 标记物。方法 进行单细胞 RNA 测序以破译来自肝细胞癌 (HCC) 样本的单细胞的转录组谱。细胞类型特异性转录本被鉴定并用于推断患有和未患有 HCC 的患者血浆 RNA 的细胞类型特异性基因特征 (CELSIG) 评分。结果鉴定出6个主要细胞簇，包括肝细胞样、胆管细胞样、肌成纤维细胞、内皮细胞、淋巴样细胞、和基于 4 个 HCC 肿瘤组织及其配对的相邻非肿瘤组织的髓细胞簇。与非 HCC 参与者 (n = 49) 相比，HCC 患者 (n = 14) 的术前血浆 RNA 样本中肝细胞样细胞的 CELSIG 评分显着增加。肿瘤切除后 3 天内，HCC 患者血浆 RNA 样本中肝细胞样细胞的 CELSIG 评分下降。与使用血浆中丰富的 ALB 转录本 [曲线下面积 (AUC) 0.72)] 区分患有和不患有 HCC 的患者的能力相比，使用 CELSIG 评分观察到了改善的性能 (AUC: 0.84)。ddPCR 分析进一步验证了血浆 RNA 中的肝细胞特异性转录标记。肝细胞样细胞和胆管细胞的 CELSIG 评分随患者的生存而变化。