Vesicular stomatitis VSVrp30 selectively destroys human metastatic melanoma

水疱性口炎VSVrp30选择性破坏人类转移性黑色素瘤

基本信息

批准号：
8635984
负责人：
ANTHONY N VAN DEN POL
金额：
$ 33.51万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-04-01 至 2017-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8635984
关键词：
Address Affect Animal Model Animals Antiviral Agents Antiviral Response Applications Grants Area Attenuated BRAF gene Blood - brain barrier anatomy Brain CDKN2A gene Cause of Death Cells Cessation of life Clinical Clinical Trials Complement Data Depressed mood Detection Development Diagnosis Elements Failure Fluorescence Frequencies Funding Gene Mutation Genes Genetic Goals Harvest Human Immune Immune system Immunization Immunocompetent Immunocompromised Host Immunologic Surveillance Immunosuppression Immunosuppressive Agents In Vitro Infection Infection Control Intravenous Life Melanoma Cell Metastatic Melanoma Metastatic malignant neoplasm to brain Modeling Mus Mutation Nature Neoplasm Metastasis Normal Cell Oncolytic Oncolytic viruses PTEN gene Patients Pharmaceutical Preparations Proto-Oncogene Protein c-kit Reporter Reporter Genes SCID Mice Sampling Secondary to Series Site Skin Solid Neoplasm Stomatitis Systemic Therapy Testing Time Transplantation Ursidae Family Vesicular stomatitis Indiana virus Viral Viral Genome Virus Virus Diseases Work base brain cell brain tissue cancer cell coral exome in vitro Model in vitro testing in vivo intravenous administration killings melanocyte melanoma mouse model neoplastic cell novel strategies oncolysis outcome forecast research clinical testing research study tumor

项目摘要

DESCRIPTION (provided by applicant): The prognosis for survival in patients with metastatic melanoma has not changed in the last 20 years and remains dismal despite advances in tumor detection and the development of melanoma-specific systemic therapies. Because of the failure of current chemotherapeutic and immunologically- based treatments to eradicate melanoma, we propose a new approach. In this proposal, we test a replication-competent oncolytic virus generated in this lab at Yale, VSVrp30. In preliminary tests VSVrp30 shows considerable promise in the potential treatment of melanoma. In vitro and in vivo experiments show that the virus selectively and rapidly infects and destroys human metastatic melanoma, with relatively little or no infection of normal human melanocytes. We seek funding to pursue experiments to determine if the virus can target and destroy melanoma cells in multiple conditions in animal and in vitro models. We will first test the oncolytic actions of the virus with a series of in vitro experiments on a large number of human melanomas and normal melanocytes available at Yale. Another set of experiments will employ human melanoma that is stably transfected with a coral reporter gene that generates a red fluorescence in the cancer cells. These human cells will be transplanted into SCID mice, both as a solid tumor, and as dispersed metastatic-like cancer cells. Virus will be given intratumorally and intravenously to test the hypothesis that the virus wll target and kill the red tumor cells with minimal infection of normal cells. Infected cells can be readily detected by the expression of a GFP reporter incorporated into the viral genome. A third set of parallel experiments will be done using the mouse melanoma line B16 in syngeneic C57Bl/6 mice with a normal immune system, allowing us to test the hypothesis that the virus can selectively detect and destroy melanoma in the presence of a normal immune system, and prolong mouse survival from melanoma; if the virus does not completely eliminate the melanoma cells, we will test the secondary hypothesis that temporarily depressing the systemic or innate immune systems with immunosuppressant drugs will enhance the oncolytic actions of the virus. A fourth set of experiments will examine the genetic mechanisms underlying the increased viral infection of melanoma cells, using an extensive series of human melanomas in which the exomes have been sequenced. These experiments will be complemented by experiments to test the hypothesis that specific induced gene mutations involving BRAF, PTEN, and CDKN2A that are common to melanoma, directly increase virus infection. A final series of experiments will test the hypothesis that the virus can cross the blood brain barrier and selectively destroy melanoma in the mouse brain, and that pre-immunization, potentially followed by transient immunosuppression, will enhance oncolysis and provide another layer of protection to the brain. If we detect collateral damage to normal brain, then we will test a new virus, 1,2-VSV, that we recently generated which is the most attenuated of any VSV we have worked with, yet still targets melanoma. Its highly attenuated nature reduces concerns relating to infection of normal brain tissue. If these experiments are successful, they will form a major advance toward clinical trials for metastatic melanoma in humans.

描述（由申请人提供）：转移性黑色素瘤患者的生存预后在过去 20 年中没有改变，尽管肿瘤检测和黑色素瘤特异性全身疗法的发展取得了进展，但仍然令人沮丧。由于目前的化疗和免疫治疗未能根除黑色素瘤，我们提出了一种新方法。在本提案中，我们测试了耶鲁大学实验室生成的具有复制能力的溶瘤病毒，VSVrp30。在初步测试中，VSVrp30 在黑色素瘤的潜在治疗方面显示出相当大的前景。体外和体内实验表明，该病毒选择性、快速地感染并破坏人类转移性黑色素瘤，而对正常人类黑色素细胞的感染相对较少或没有。我们寻求资金进行实验，以确定该病毒是否可以在动物和体外模型的多种条件下靶向并破坏黑色素瘤细胞。我们将首先对耶鲁大学提供的大量人类黑色素瘤和正常黑色素细胞进行一系列体外实验，测试该病毒的溶瘤作用。另一组实验将采用稳定转染珊瑚报告基因的人类黑色素瘤，该基因在癌细胞中产生红色荧光。这些人类细胞将被移植到 SCID 小鼠体内，既作为实体瘤，又作为分散的转移性癌细胞。病毒将通过肿瘤内和静脉注射来测试病毒将靶向并杀死红肿瘤细胞而对正常细胞的感染最小化的假设。通过整合到病毒基因组中的 GFP 报告基因的表达，可以轻松检测到受感染的细胞。第三组平行实验将使用小鼠黑色素瘤系 B16 在具有正常免疫系统的同基因 C57Bl/6 小鼠中进行，使我们能够检验病毒可以在正常免疫系统存在的情况下选择性检测和破坏黑色素瘤的假设，并延长小鼠黑色素瘤的存活时间；如果病毒不能完全消除黑色素瘤细胞，我们将检验第二个假设，即用免疫抑制剂暂时抑制全身或先天免疫系统将增强病毒的溶瘤作用。第四组实验将使用一系列外显子组已测序的人类黑色素瘤，研究黑色素瘤细胞病毒感染增加的遗传机制。这些实验将通过实验来补充，以检验以下假设：涉及黑色素瘤常见的 BRAF、PTEN 和 CDKN2A 的特定诱导基因突变会直接增加病毒感染。最后一系列实验将检验以下假设：病毒可以穿过血脑屏障并选择性地破坏小鼠大脑中的黑色素瘤，而预免疫（可能随后进行短暂的免疫抑制）将增强溶瘤作用，并为小鼠大脑提供另一层保护。脑。如果我们检测到对正常大脑的附带损害，那么我们将测试我们最近生成的一种新病毒 1,2-VSV，它是我们研究过的所有 VSV 中减毒程度最强的，但仍然针对黑色素瘤。其高度减毒特性减少了对正常脑组织感染的担忧。如果这些实验成功，将成为人类转移性黑色素瘤临床试验的重大进展。