Purpose

Hepatoblastoma is the most common liver malignancy in children. In order to advance therapy against hepatoblastoma, novel immunologic targets and biomarkers are needed. Our purpose in this investigation is to examine hepatoblastoma transcriptomes for the expression of a class of genomic elements known as Human Endogenous Retrovirus (HERVs). HERVs are abundant in the human genome and are biologically active elements that have been associated with multiple malignancies and proposed as immunologic targets in a subset of tumors. A sub-family of HERVs, HERV-K(HML-2) (HERV-K), have been shown to be tightly regulated in fetal development, making investigation of these elements in pediatric tumors paramount.

Methods

We first created a HERVK-FASTA file utilizing 91 previously described HML-2 proviruses. We then concatenated the file onto the GRCh38.95 cDNA library from Ensembl. We used this reference database to evaluate existing RNA-seq data from 10 hepatoblastoma tumors and 3 normal liver controls (GEO accession ID: GSE8977575). Quantification and differential proviral expression analysis between hepatoblastoma and normal liver controls was performed using the pseudo-alignment program Salmon and DESeq2, respectively.

Results

HERV-K mRNA was expressed in hepatoblastoma from multiple proviral loci. All expressed HERV-K proviral loci were upregulated in hepatoblastoma compared to normal liver controls. Five HERV-K proviruses (1q21.3, 3q27.2, 7q22.2, 12q24.33 and 17p13.1) were significantly differentially expressed (p-adjusted value < 0.05, | log2 fold change | > 1.5) across conditions. The provirus at 17p13.1 had an approximately 300-fold increased expression in hepatoblastoma as compared to normal liver. This was in part due to the near absence of HERV-K mRNA at the 17p13.1 locus in fully differentiated liver samples.

Conclusions

Our investigation demonstrates that HERV-K is expressed from multiple loci in hepatoblastoma and that the expression is increased for several proviruses compared to normal liver controls. Our results suggest that HERV-K mRNA expression may be useful as a biomarker in hepatoblastoma, given the large differential expression profiles in hepatoblastoma, with very low mRNA levels in liver control samples.

中文翻译：

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肝母细胞瘤中人类内源性逆转录病毒-K（HML-2）mRNA的上调：潜在的新型免疫治疗靶标和生物标志物的鉴定

目的

肝母细胞瘤是儿童中最常见的肝恶性肿瘤。为了推进针对肝母细胞瘤的治疗，需要新的免疫学靶标和生物标记。我们在这项研究中的目的是检查肝母细胞瘤转录组中一类称为人类内源性逆转录病毒（HERV）的基因组元件的表达。HERV在人类基因组中含量很高，是与多种恶性肿瘤相关的生物活性元素，被提议作为肿瘤子集中的免疫学靶标。HERV的一个子家族HERV-K（HML-2）（HERV-K）已被证明在胎儿发育过程中受到严格的调控，这使得对小儿肿瘤中这些元素的研究成为重中之重。

方法

我们首先使用91个先前描述的HML-2前病毒创建了HERVK-FASTA文件。然后，我们将文件连接到Ensembl的GRCh38.95 cDNA文库中。我们使用该参考数据库评估了来自10个肝母细胞瘤肿瘤和3个正常肝脏对照（GEO登录号：GSE8977575）的现有RNA-seq数据。分别使用伪比对程序Salmon和DESeq2对肝母细胞瘤和正常肝脏对照进行定量和差异化前病毒表达分析。

结果

HERV-K mRNA在来自多个原病毒基因座的肝母细胞瘤中表达。与正常肝对照相比，肝母细胞瘤中所有表达的HERV-K前病毒基因座均上调。五个HERV-K前病毒（1q21.3、3q27.2、7q22.2、12q24.33和17p13.1）在不同条件下均显着差异表达（p调整值<  0.05，|  log2倍数变化 |> 1.5）。与正常肝脏相比，原发性肝癌中17p13.1的表达增加了约300倍。这部分是由于在完全分化的肝脏样品中17p13.1位点几乎没有HERV-K mRNA。

结论

我们的研究表明，HERV-K在肝母细胞瘤中由多个基因座表达，并且与正常肝脏对照相比，几种原病毒的表达增加。我们的结果表明，鉴于肝母细胞瘤中的差异表达谱较大，而肝对照样品中的mRNA水平非常低，HERV-K mRNA表达可能在肝母细胞瘤中用作生物标志物。