Identification and characterization of a potential Biomarker and Modulator for HCC development derived from cancer type-specific long noncoding (Lnc) RNAs.

癌症研究已经进入了一个新的阶段，来自基因组学、生物信息学和现代遗传学的信息开始为潜在的癌症类型特异性生物标记物提供新的迹象。最近的ribo-seq数据显示，在人类细胞中表达的所有长非编码(Lnc)RNA中，有40%被翻译为“隐藏的多肽”。由于lncRNA表达人类细胞特异性、细胞类型特异性和癌症特异性，这些隐藏的多肽可能是癌症类型特异性抗原，也可能与生物学功能有关。揭开异质性肿瘤(如肝细胞癌)的基因组改变模式对于确定风险群体中的生物标记物以及改善患者护理的靶向治疗至关重要。事实上，我们最近发现Linc00176以肝癌特有的方式在外显子2内剪接，并被翻译成189个氨基酸长的多肽。此外，通过分析Ribo-Seq数据，我们还检测到了30个HepG2特异的潜在“隐藏多肽”。这项应用将进行以下工作：首先，将利用我们实验室最近建立的纳米孔技术来鉴定新的lncRNAs和lncRNAs的剪接变体。其次，为了检验这些多肽是否是内源性合成的，并且在细胞中是稳定的，我们将产生针对潜在的“隐藏多肽”的合成多肽特异性抗体。第三，我们将从HepG2细胞中分离并测序小肽。此外，为了检查它们的生物学功能，第一个蛋氨酸残基将被使用CRISPER的终止密码子取代，并将检查其表型。最后，使用针对这些多肽的抗体，我们将检测它们是否在原发性肝细胞癌、肝细胞腺瘤、肝硬变和正常肝脏中表达，以确定这些多肽是否可用作前肝癌和肝癌的生物标志物。在未来，这些多肽的抗原提呈将被检查。

Cancer research has entered a new phase in which information from genomics, bioinformatics and modern genetics are beginning to provide new indications of potential cancer type-specific biomarkers. Recent Ribo-seq data revealed that 40% of all long noncoding (lnc) RNAs expressed in human cells are translated as “hidden peptides.”. Since lncRNAs express human cell- specific, cell type-specific and cancer- specific properties, these hidden peptides may be cancer type-specific antigens and may also be associated with biological functions. Unraveling the patterns of genomic alterations in heterogeneous tumors such as Hepatocellular carcinoma (HCC) is pivotal for identifying biomarkers in risk groups, and for targeted therapies that could improve patient care. Indeed, we recently found that Linc00176 is spliced within Exon 2 in an HCC specific manner and is translated into a polypeptide 189 amino acids long. In addition, we detected 30 HepG2 specific potential “hidden peptides” by analyzing Ribo-seq data. This application is to undertake the following: Firstly, novel lncRNAs and splice variants of lncRNAs will be identified using nanopore technology that has recently been established in our laboratory. Secondly, to examine whether these peptides are endogenously synthesized and are stable in cells, we will generate synthetic peptide specific antibodies against potential “hidden peptides.”. Thirdly, we will isolate and sequence small peptides from HepG2 cells. Furthermore, to examine their biological functions, the first methionine residue will be replaced by a stop codon using CRISPER and the phenotype will be examined. Finally, using antibodies against these peptides we will examine whether they are expressed in primary HCC, hepatocellular adenoma, liver cirrhosis, and normal liver to determine whether these are useful as biomarkers for pre-HCCs and HCCs. In the future, antigen presentation of these peptides will be examined.