Rapid development of replication-controlled vaccinia virus vectors for vaccines and therapeutics with single or double safety features

快速开发复制控制的痘苗病毒载体，用于具有单一或双重安全特征的疫苗和疗法

• 批准号: 9230098
• 负责人: PAULO H VERARDI
• 金额: $ 21.93万
• 依托单位: UNIVERSITY OF CONNECTICUT STORRS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-12-09 至 2018-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9230098
• 关键词: Address; Adverse reactions; Animals; Antibiotic Therapy; Antibiotics; Atopic Dermatitis; Attenuated; Attenuated Vaccines; Body Weight decreased; Cancer Vaccines; Cell Culture Techniques; Communicable Diseases; Data; Development; Disease; Doxycycline; Elements; Essential Genes; Genes; Goals; Growth; Heart Diseases; Human; Immunosuppression; Immunotherapy; In Vitro; Individual; Interferon Type II; Interferons; Internal Ribosome Entry Site; Kinetics; Lesion; Link; Methods; Modified Vaccinia Virus Ankara; Mus; Mutation; Oncolytic; Operon; Poxviridae; Process; Research; Resolution; SCID Mice; Safety; Smallpox; Smallpox Vaccine; Smallpox Viruses; System; Tetanus Helper Peptide; Tetracyclines; Therapeutic; Vaccination; Vaccines; Vaccinia virus; Viral Genome; Virus; Zika Virus; attenuation; base; cancer therapy; genetic element; immunogenic; immunogenicity; in vivo; mouse model; oncolytic virotherapy; preclinical study; prevent; promoter; safety testing; vaccine development; vaccine safety; vaccinia virus vector; vector; vector vaccine; viral vector development

PROJECT SUMMARY This application is responsive to PA-15-313, Research to Advance Vaccine Safety (R21). Vaccinia virus (VACV) was used as a live vaccine for smallpox, a disease caused by variola virus. VACV has also been successfully used as a live viral vector for the development of effective human and animal vaccines, as well as immunotherapies and oncolytic virotherapies. However, VACV can cause complications in individuals with conditions such as atopic dermatitis, cardiac disease, and immunosuppression. Consequently, individuals with such conditions or with contacts that have these conditions are contraindicated for vaccination with replicating VACV vectors. We recently generated VACV vectors with a built-in safety mechanism that replicate only in the presence of tetracycline (TC) antibiotics (replication-inducible VACVs). In this system, a VACV gene essential for replication (eg, D6R) is inducibly expressed by TCs using elements of the tet operon. When administered as a vaccine (in the absence of antibiotics), the vector does not replicate but retains its immunogenicity, and therefore is safer for human use. Conveniently, the vector can be propagated in cell culture at high titers in the presence of TCs, unlike other replication-defective VACV-based vectors such as MVA. We also developed VACV vectors that replicate normally in the absence of antibiotics, but are replication-defective in the presence of TCs (replication-repressible VACVs). When administered as a vaccine (in the absence of antibiotics), the vector is replication competent like traditional VACV vectors (and therefore highly immunogenic), and treatment of any adverse reactions would be as simple as TC antibiotic therapy. We can further enhance the safety of our vectors by a fail-safe feature that links expression of the essential gene (D6R) with interferon-γ (IFN-γ), via an internal ribosome entry site (IRES). We showed that expression of IFN-γ by VACV, either constitutively or inducibly, leads to complete attenuation of VACV in vivo, even when expressed at very low levels. Surprisingly, the virus is still able to grow to wild-type levels in vitro. Thus, both strategies can be combined to develop replication-defective VACVs inducibly or repressibly expressing D6R (under its natural promoter) and IFN-γ (under a small murine IRES). The resulting vectors should be able to grow to high titers in vitro (thus allowing propagation) in the presence of TCs (inducible vectors) or absence of TCs (repressible vectors). Since expression of D6R will be linked to IFN-γ expression, any potential replication-competent VACV that may originate during in vitro propagation or in vivo administration would be replication-incompetent in vivo due to concomitant expression of IFN-γ. In Aim 1 we will determine the safety of replication-inducible and replication-repressible VACV vectors in vivo using immunodeficient SCID mice. In Aim 2, we will develop, characterize, and determine the safety of replication-defective VACV vectors with a fail-safe feature. We plan to use this VACV vaccine platform for rapid development of safe vaccines for infectious diseases such as Zika virus, immunotherapies (eg, personalized cancer vaccines), and oncolytic virotherapies.

项目概要 此应用程序响应 PA-15-313，促进疫苗安全性研究 (R21)。 （VACV）被用作天花的活疫苗，天花也是一种由天花病毒引起的疾病。 成功用作活病毒载体，用于开发有效的人类和动物疫苗，以及 然而，VACV 可能会导致患者出现并发症。 患有特应性皮炎、心脏病和免疫抑制等疾病的个体。 此类情况或与患有此类情况的接触者禁止接种复制型疫苗 我们最近生成了具有内置安全机制的 VACV 载体，该机制仅在 存在四环素 (TC) 抗生素（复制诱导型 VACV），在此系统中，VACV 基因必不可少。 当施用时，TCs使用tet操纵子的元件诱导表达用于复制的基因（例如，D6R）。 作为疫苗（在没有抗生素的情况下），载体不会复制，但保留其免疫原性，并且 因此，该载体可以在细胞培养物中以高滴度繁殖，方便人类使用。 TC 的存在，与其他复制缺陷型 VACV 载体（例如 MVA）不同。 VACV 载体在没有抗生素的情况下可以正常复制，但在存在抗生素的情况下会出现复制缺陷 TC（复制抑制型 VACV）作为疫苗施用时（在没有抗生素的情况下）， 载体与传统 VACV 载体一样具有复制能力（因此具有高度免疫原性），并且 任何不良反应的治疗都会像TC抗生素治疗一样简单，我们可以进一步加强。 通过将必需基因 (D6R) 的表达与干扰素-γ 连接起来的自动防故障功能，确保我们载体的安全性 (IFN-γ)，通过内部核糖体进入位点 (IRES) 我们表明 VACV 表达 IFN-γ。 组成型或诱导型，导致 VACV 在体内完全减弱，即使表达量非常低 令人惊讶的是，该病毒仍然能够在体外生长至野生型水平，因此，这两种策略都可以。 联合开发复制缺陷型 VACV，诱导或抑制表达 D6R（在其自然条件下） 启动子）和 IFN-γ（在小型鼠 IRES 下） 所得载体应该能够在 中生长至高滴度。 在存在 TC（诱导型载体）或不存在 TC（抑制型载体）的情况下进行体外（从而允许繁殖） 由于 D6R 的表达将与 IFN-γ 的表达相关，任何潜在的复制能力 可能源自体外繁殖或体内施用期间的 VACV 将无法复制 在体内由于 IFN-γ 的伴随表达，在目标 1 中，我们将确定复制诱导型的安全性。 以及使用免疫缺陷 SCID 小鼠体内复制抑制的 VACV 载体 在目标 2 中，我们将开发， 我们计划通过故障安全功能来表征和确定复制缺陷型 VACV 载体的安全性。 利用该VACV疫苗平台快速开发针对寨卡等传染病的安全疫苗 病毒、免疫疗法（例如个性化癌症疫苗）和溶瘤病毒疗法。