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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：
10159871
负责人：
Sumitra Deb
金额：
$ 47.41万
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/10159871
关键词：
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 base 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，并且这些突变大多是单一的氨基酸变化逐渐灌输功能获得性（GOF）致癌表型，使GOF p53癌蛋白一个出色的癌症治疗靶点。我们提出了一种完全不同的方法，目前GOF p53靶向我们不是抑制蛋白质，而是将GOF p53武器化以促进肺癌的治疗概念细胞死亡，或者通过自杀，病毒裂解，或者两者兼而有之，而使正常细胞毫发无损。这一创新战略这是可能的，因为我们发现了GOF p53独特的反式激活机制， GOF p53诱导型启动子的产生。我们的GOF p53诱导型启动子指导任何基因的表达只有当细胞具有GOF p53突变时，才克隆到下游，野生型（WT）p53对下游细胞没有影响。启动子和显示无表达的具有WT p53或p53无效突变的细胞。对于我们的第一个主要目标，我们我建议使用一种令人兴奋的新型溶瘤病毒，这种病毒只能复制、繁殖和杀死具有GOF的癌细胞。 p53对正常细胞没有影响。我们将两个腺病毒早期基因E1A和E1B，腺病毒复制所需的基因，在GOF p53诱导型启动子的控制下，腺病毒载体初步研究表明，该病毒具有显著的溶瘤能力和肺特异性，在具有GOF p53的癌细胞中，在具有WT p53的细胞中没有任何作用或病毒生长。杀鼠效果具有GOF p53的异种移植肿瘤似乎在短时间延迟后存在持续加速的肿瘤杀伤注射病毒的时间我们建议通过增加额外的裂解能力来增强溶瘤病毒，通过结合自杀策略和溶瘤策略。初步结果看起来非常有希望组合.对于我们的第二个目标，我们建议设计一种特异性杀死肺癌细胞的方法，通过克隆我们的GOF p53诱导型启动子下游的自杀基因来突变GOF p53。该构建体这样当病毒感染细胞时，只有GOF p53突变的细胞（癌细胞）将死于前药治疗。我们用疱疹胸苷激酶制造了这样一种病毒自杀基因，并显示出显着的杀伤作用和特异性的肺癌细胞与GOF p53在培养和异种移植肿瘤中。我们的目标是进一步发现这种策略是如何工作的，以及如何改进它。使用细菌胞嘧啶脱氨酶自杀基因（bCD）增强GOF的旁观者效应 p53自杀病毒此外，我们建议提高我们的GOF p53诱导型启动子的诱导性，以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，因为这些策略应该适用于任何具有GOF p53突变的癌症，其占所有癌症的一半以上。