Effect of APOBEC3 on Bladder Cancer Biology and Response to Immunotherapy

APOBEC3 对膀胱癌生物学和免疫治疗反应的影响

基本信息

批准号：
10092808
负责人：
Andrew Truong
金额：
$ 4.6万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2023-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10092808
关键词：
Adenoviruses Alleles Allografting Anchorage-Independent Growth Antigens Automobile Driving Bladder Bladder Neoplasm Cancer Biology Cancer Model Cancer Patient Cancer cell line Cisplatin Code Cytidine DNA biosynthesis Development Ear Enzymes Evolution Experimental Models Family Fibroblasts Genes Genetic Transcription Genetically Engineered Mouse Immune Immune checkpoint inhibitor Immunocompetent Immunotherapy Impairment In Vitro Individual Interferons Intuition Knock-in Knock-in Mouse Knowledge Malignant Neoplasms Malignant neoplasm of liver Malignant neoplasm of urinary bladder Mammalian Cell Mediating Missense Mutation Modeling Mus Mutagenesis Mutation Natural Immunity Nature Nonsense Mutation Nucleic Acids PD-1 blockade Patients Pattern Peptides Play Protein Family Protein Overexpression Proteins Refractory Resistance Role Shapes Signal Transduction Single Nucleotide Polymorphism Single-Stranded DNA Survival Rate Testing Transgenic Mice Transitional Cell Carcinoma UPK3 gene United States Uracil Urothelium Viral Virus Diseases Virus Replication anti-PD-1 anti-PD1 therapy anti-tumor immune response apolipoprotein B mRNA editing enzyme base cancer cell cancer type effective therapy exome sequencing immune checkpoint blockade improved inhibitor/antagonist member mouse model neoantigens novel overexpression polypeptide prevent response tool tumor tumor heterogeneity tumor progression

项目摘要

PROJECT SUMMARY The apolipoprotein B mRNA editing enzyme catalytic polypeptide 3 (APOBEC3) family of proteins play an important role in innate immunity by restricting retroviral replication. During a viral infection, these enzymes are upregulated by interferon signaling. They catalyze the conversion of cytidine to uracil within reverse-transcribed, single-stranded DNA, resulting in the degradation or integration of defective viral nucleic acid, and therefore, impaired viral replication. During DNA replication and transcription in mammalian cells, single-stranded DNA can also serve as the substrate for APOBEC3 enzymes. Given the highly proliferative nature of cancer cells, and a high level of interferon signaling in certain cancer types, APOBEC-induced mutagenesis can contribute significantly to their mutational spectrum. In cancer, high APOBEC expression correlates with an increased tumor mutational burden (TMB) and predicted neoantigen load. Studies in vitro and in liver cancer transgenic mouse models have shown that APOBEC overexpression can promote tumor progression. In urothelial carcinoma, the expression of APOBEC3B, a particular member of this protein family, is significantly upregulated. Strikingly, upwards of 70% of mutations can be attributed to APOBEC-induced mutagenesis. I have generated mouse bladder cancer cell lines with inducible murine Apobec3. I confirmed the induction of Apobec3 protein, induction of an APOBEC mutational pattern, and the ability of Apobec3 to promote anchorage-independent growth. In addition, we have generated a novel Rosa26-Apobec3 knock-in allele in which the expression of Apobec3 is Cre-inducible, allowing for spatial and temporal control of Apobec3 expression. Currently, no studies have examined APOBEC's role in mediating intratumoral heterogeneity, or in response to immune checkpoint blockade. High intratumoral heterogeneity has been shown to negatively correlate with response to immunotherapy. Additionally, because of the increased mutational load (and therefore, neoantigens) imparted by APOBEC-induced mutagenesis, one would expect a robust response to immune checkpoint blockade. However, many of the neoantigens may be subclonal and persistent APOBEC mutagenesis, in the context of a neoantigen driven anti-tumor immune response, may allow for antigen escape (i.e. due to nonsense mutations in the gene encoding the neoantigen). I will utilize the tools we have developed to answer these important questions. Specifically, in Aim 1, I will cross our novel Apobec3 knock-in allele to our established Pten and Trp53-deficient mouse model of bladder cancer and characterize the intratumoral heterogeneity by multiregional whole-exome sequencing. In Aim 2, I will utilize an allograft bladder cancer model with inducible Apobec3 to test whether Apobec3 mutagenic activity can dampen the efficacy of immunotherapy. Results of the proposed aims will contribute to a greater understanding of the role of APOBEC3 mutagenesis in fueling the evolution of bladder cancer. This expansion of knowledge will provide the rationale to develop more effective treatments for bladder cancer such as an APOBEC3 inhibitor.

项目概要载脂蛋白 B mRNA 编辑酶催化多肽 3 (APOBEC3) 家族蛋白发挥着通过限制逆转录病毒复制在先天免疫中发挥重要作用。在病毒感染期间，这些酶干扰素信号传导上调。它们在逆转录过程中催化胞苷转化为尿嘧啶，单链DNA，导致有缺陷的病毒核酸的降解或整合，因此，病毒复制受损。在哺乳动物细胞中 DNA 复制和转录过程中，单链 DNA 可以也可作为 APOBEC3 酶的底物。鉴于癌细胞的高度增殖性，以及 APOBEC 诱导的突变可能有助于某些癌症类型中高水平的干扰素信号传导对其突变谱有显着影响。在癌症中，APOBEC 高表达与肿瘤增加相关突变负荷（TMB）和预测的新抗原负荷。体外和肝癌转基因小鼠的研究模型显示 APOBEC 过度表达可以促进肿瘤进展。在尿路上皮癌中， APOBEC3B（该蛋白家族的一个特殊成员）的表达显着上调。引人注目的是，超过 70% 的突变可归因于 APOBEC 诱导的突变。我已经生成了鼠标具有诱导型小鼠 Apobec3 的膀胱癌细胞系。我确认了Apobec3蛋白的诱导，诱导 APOBEC 突变模式的研究，以及 Apobec3 促进锚定非依赖性生长的能力。在此外，我们还生成了一个新的 Rosa26-Apobec3 敲入等位基因，其中 Apobec3 的表达为 Cre 诱导型，允许对 Apobec3 表达进行空间和时间控制。目前，尚无研究研究了 APOBEC 在介导肿瘤内异质性或响应免疫检查点方面的作用封锁。肿瘤内的高异质性已被证明与对药物的反应呈负相关免疫疗法。此外，由于突变负荷（因此，新抗原）增加通过 APOBEC 诱导的突变，人们预计会对免疫检查点封锁产生强烈的反应。然而，许多新抗原可能是亚克隆和持久的 APOBEC 突变，在新抗原驱动的抗肿瘤免疫反应，可能允许抗原逃逸（即由于无义突变在编码新抗原的基因中）。我将利用我们开发的工具来回答这些重要的问题问题。具体来说，在目标 1 中，我将把我们新颖的 Apobec3 敲入等位基因与我们已建立的 Pten 和 Trp53 缺陷小鼠膀胱癌模型并通过多区域表征瘤内异质性全外显子组测序。在目标 2 中，我将利用具有诱导型 Apobec3 的同种异体移植膀胱癌模型来测试 Apobec3 诱变活性是否会削弱免疫疗法的功效。拟议目标的结果将有助于更好地了解 APOBEC3 突变在促进膀胱进化中的作用癌症。这种知识的扩展将为开发更有效的膀胱治疗方法提供依据癌症，例如 APOBEC3 抑制剂。