Targeting lung tumors expressing mutant p53 with oncolytic viruses and suicide genes

用溶瘤病毒和自杀基因靶向表达突变型 p53 的肺部肿瘤

基本信息

批准号：
10655337
负责人：
Sumitra Deb
金额：
$ 46.46万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
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：
2020-07-01 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10655337
关键词：
Acceleration Adenovirus Vector Adenoviruses Affect Alleles Amino Acids Automobile Driving Base Sequence Biological Assay Biological Models Bystander Effect Cell Death Cells Cessation of life Cloning Cytolysis Cytomegalovirus Cytosine deaminase DNA Sequence Early Promoters Exposure to Ganciclovir Gene Activation Gene Expression Gene Proteins Genes Goals Growth Herpesvirus 1 Human Lung Lung Neoplasms Lytic Lytic Phase Malignant Neoplasms Malignant neoplasm of lung Mediating Modeling Mus Mutation Mutation Analysis Normal Cell Oncogenic Oncogenic Viruses Oncolytic Oncolytic viruses Phenotype Process Prodrugs Property Proteins Reporter Research Response Elements Specificity Suicide System TP53 gene Technology Testing Therapeutic Thymidine Kinase Time Transactivation Viral Virus Virus Replication Work Xenograft procedure cancer cell cell type gain of function gene cloning improved individual response innovation lung cancer cell mouse model mutant novel null mutation oncolysis oncolytic adenovirus patient derived xenograft model promoter response suicide gene suicide virus targeted treatment therapeutic target thymidine kinase 1 tumor tumor xenograft vector weapons

项目摘要

Mutations of the p53 gene are found in the majority of lung cancers and most of these mutations are single amino acid changes instilling gain-of-function (GOF) oncogenic phenotypes, making the GOF p53 oncoprotein an excellent cancer therapeutic target. We propose a radically different approach to current GOF p53 targeting therapeutic concepts in which instead of inhibiting the protein, we weaponize GOF p53 in promoting lung cancer cell death, either by suicide, viral lysis, or both, while leaving normal cells unscathed. This innovative strategy is possible based on our discovery of a unique transactivation mechanism for GOF p53, and from that, our creation of a GOF p53 inducible promoter. Our GOF p53 inducible promoter directs expression of any gene cloned downstream only if the cell has a GOF p53 mutation, with wild-type (WT) p53 having no effect on the promoter and cells with WT p53 or p53 null mutations showing no expression. For our first major goal, we propose using an exciting new oncolytic virus that only replicates, propagates, and kills cancer cells with GOF p53 while having no effect on normal cells. We have placed two adenoviral early genes, E1A and E1B, the genes needed for adenoviral replication, under the control of the GOF p53 inducible promoter within an adenoviral vector. Initial studies show that this virus has remarkable oncolytic ability and specificity for lung cancer cells with GOF p53, with no effect or viral growth whatsoever in cells with WT p53. The killing effects in xenograft tumors with GOF p53 appear as though there is sustained accelerated tumor killing after a short delay of when the virus is injected. We propose to enhance the oncolytic virus by adding additional lysis abilities and by combining the suicide strategy with the oncolytic strategy. Preliminary results look very promising for this combination. For our second goal, we propose devising a means of specifically killing lung cancer cells with GOF p53 mutations by cloning a suicide gene downstream of our GOF p53 inducible promoter. This construct will be introduced into an adenoviral vector so that when the virus infects cells, only cells with a GOF p53 mutation (cancer cells) will die from prodrug treatment. We have created such a virus using the Herpes Thymidine Kinase suicide gene and show striking killing effects and specificity for lung cancer cells with GOF p53 both in culture and in xenograft tumors. We aim to further discover how this strategy works and ways to improve it. We propose the use of the bacterial Cytosine Deaminase suicide gene (bCD) to enhance the bystander effect of our GOF p53 specific suicide virus. In addition, we propose to improve the inducibility of our GOF p53 inducible promoter to further enhance the suicide and oncolytic viruses. The potential impact of this work is far-reaching since these strategies should be applicable for any cancer with GOF p53 mutations, which constitutes over half of all cancers.

在大多数肺癌中发现了p53基因的突变，其中大多数是单个突变氨基酸改变灌输功能获得（GOF）致癌表型，使GOF p53癌蛋白一个极好的癌症治疗靶标。我们提出了针对当前GOF P53靶向的一种根本不同的方法在促进肺癌中，我们将GOF p53武器化，而不是抑制蛋白质，而不是抑制蛋白质的治疗概念通过自杀，病毒裂解或两者兼而有之的细胞死亡，同时毫发无损地离开正常细胞。这种创新的策略基于我们发现GOF p53的独特反式激活机制，因此我们可能是可能的创建GOF p53诱导启动子。我们的GOF p53诱导启动子指导任何基因的表达仅当细胞具有GOF p53突变时，才将其克隆到下游，野生型（WT）p53对启动子和WT p53或p53无效突变的细胞显示未表达。对于我们的第一个主要目标，我们建议使用一种令人兴奋的新溶瘤病毒，该病毒仅重复，传播和杀死GOF p53虽然对正常细胞没有影响。我们已经放置了两个腺病毒早期基因E1A和E1B，在GOF p53诱导型启动子的控制下，腺病毒复制所需的基因腺病毒载体。初步研究表明，该病毒具有明显的肺溶性能力和肺的特异性具有GOF p53的癌细胞，在WT p53的细胞中无效或病毒生长无效。杀人效应带有GOF p53的异种移植肿瘤似乎在短暂延迟后持续加速肿瘤杀死注射病毒的时间。我们建议通过增加额外的裂解能力和通过将自杀策略与溶瘤策略相结合。初步结果看起来很有希望组合。对于我们的第二个目标，我们提出了一种专门杀死肺癌细胞的方法 GOF p53突变通过克隆我们GOF p53诱导启动子的下游自杀基因。这个结构将被引入腺病毒载体中，以便当病毒感染细胞时，只有具有GOF p53突变的细胞（癌细胞）将死于前药治疗。我们使用疱疹胸苷激酶创建了这样的病毒自杀基因并显示出具有GOF p53的肺癌细胞的惊人杀伤作用以及在培养中的特异性和异种移植肿瘤。我们的目标是进一步发现该策略如何运作和改进方法。我们建议使用细菌胞嘧啶脱氨酶自杀基因（BCD）来增强我们的GOF的旁观者效应 p53特异性自杀病毒。此外，我们建议提高GOF p53诱导启动子的诱导性进一步增强自杀和溶瘤病毒。这项工作的潜在影响是深远的，因为这些策略应适用于具有GOF p53突变的任何癌症，该突变占所有癌症的一半以上。