Hepatitis B virus integrations in KMT2B drive hepatocellular carcinoma

摘要 B型肝炎病毒（HBV）的慢性感染是肝细胞癌（HCC）的主要原因。HBV单药 占全球所有癌症部位死亡人数的3%以上。HBV如何促进 致癌作用尚未完全了解，导致晚期疾病的治疗选择有限。我们 先前的研究表明，10%的HBV相关的HCC肿瘤携带HBV在KMT 2B中的克隆整合， 癌细胞基因组中的基因位点。所有这些整合都位于KMT 2B的外显子3和6之间 基因KMT 2B是KMT 2A（也称为MLL 1）的唯一同源物。众所周知， 影响KMT 2A的染色体易位是婴儿混合谱系白血病的主要驱动因素。HBV HCC中KMT 2B的整合可能反映了白血病中KMT 2A的重排，但其功能尚不清楚。 研究了我们已经鉴定了携带HBV整合在KMT 2B中的患者来源的HCC细胞系。利用这些细胞 在KMT 2B中具有HBV整合的HCC肿瘤的细胞系和RNA-seq数据中，我们发现这些整合导致 涉及从N-末端至697至906个氨基酸残基的截短的KMT 2B的表达。敲除 KMT 2B截短携带整合的细胞系在体外抑制细胞增殖。的过表达 KMT 2B截短在体外转化正常人肝细胞，并在体内诱导肿瘤生长。 基于流体动力学注射的小鼠模型。从机制上讲，我们发现KMT 2B截短隔离了 肿瘤抑制因子MENIN来自KMT 2A/B组蛋白甲基转移酶复合物和染色体。A KMT2B 携带阻断MENIN结合的突变的截短不能促进体内肿瘤形成。基于 根据这些初步数据，我们的中心假设是HBV整合在KMT 2B中产生截短， 从KMT 2B复合物中隔离MENIN并促进肝细胞癌。为了验证这个假设， 我们将使用各种体外和体内模型确定KMT 2B截短的致癌功能（目的 1），并确定KMT 2B截短触发肿瘤发生（Aim 2）的机制。具体来说，一个新的 基因工程小鼠肝癌模型的建立，将成为一种潜在的肝癌HBV靶向治疗药物 将测试KMT 2B中的集成。拟议的研究旨在建立致癌驱动功能， 肝癌中KMT 2B中的HBV整合机制，每年导致35，000例肝癌病例。它是其中的 最常见的致癌基因改变，但它是显着研究不足。的完成 拟议的研究将发现一种新的癌病毒驱动的HCC发展的表观遗传机制， 为开发预防方法、早期检测测定和靶向治疗提供信息， 受影响的个人。

ABSTRACT Chronic infection by hepatitis B virus (HBV) is the leading cause of hepatocellular carcinoma (HCC). HBV alone is responsible for more than 3% of worldwide cancer deaths of all cancer sites combined. How HBV promotes carcinogenesis is not fully understood, leading to limited therapeutic options for advanced diseases. We previously revealed that 10% of HBV-associated HCC tumors carry clonal integrations of HBV in the KMT2B gene locus in cancer cell genomes. All these integrations are located between exons 3 and 6 of the KMT2B gene. KMT2B is the only close homolog of KMT2A (also known as MLL1). It has been well established that the chromosomal translocations affecting KMT2A are the primary drivers of infant mixed lineage leukemia. HBV integrations in KMT2B in HCC may mirror KMT2A rearrangements in leukemia, but its function has not been studied. We have identified patient-derived HCC cell lines carrying HBV integrations in KMT2B. Using these cell lines and RNA-seq data of HCC tumors with HBV integrations in KMT2B, we found that these integrations lead to expression of truncated KMT2B from the N-terminus to 697 to 906 amino acid residues. Knockout of the KMT2B truncation in an integration-carrying cell line suppressed cell proliferation in vitro. Overexpression of a KMT2B truncation transformed normal human hepatocytes in vitro and induced tumor growth in vivo in hydrodynamic injection-based mouse models. Mechanistically, we found the KMT2B truncation sequestered the tumor suppressor MENIN from the KMT2A/B histone methyltransferase complex and chromosome. A KMT2B truncation carrying a mutation that blocks MENIN binding failed to promote tumor formation in vivo. Based on these preliminary data, our central hypothesis is that HBV integrations in KMT2B produce truncations, which sequester MENIN from KMT2B complex and promote hepatocellular carcinoma. To test this hypothesis, we will determine the oncogenic function of KMT2B truncations, using various in vitro and in vivo models (Aim 1), and determine the mechanism of KMT2B truncation-triggered tumorigenesis (Aim2). Specifically, a new genetic engineered mouse model of HCC will be generated and a potential targeted agent for HCC with HBV integrations in KMT2B will be tested. The proposed studies aim to establish the oncogenic driving function and mechanism of HBV integrations in KMT2B in HCC, which cause 35,000 HCC cases every year. It is among the most common cancer-causing genetic alterations but it is significantly understudied. The completion of the proposed studies will discover a novel oncovirus-driving, epigenetic mechanism of HCC development and provide information for developing prevention approaches, early detection assays, and targeted therapeutics for affected individuals.