Arming Oncolytic HSV Vectors to Induce Anti-GBM Immune Responses in Syngeneic Mice

武装溶瘤 HSV 载体在同基因小鼠中诱导抗 GBM 免疫反应

• 批准号: 9927607
• 负责人: Joseph C Glorioso
• 金额: $ 39.89万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-06-07 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9927607
• 关键词: Adult; Affect; American; Angiogenesis Inhibitors; Animal Model; Animals; Antibodies; Brain Neoplasms; CD8-Positive T-Lymphocytes; CTLA4 gene; Cells; Clinical Trials; Cytotoxic T-Lymphocytes; Dose; Effectiveness; Engineering; Epidermal Growth Factor Receptor; Evaluation; Expression Profiling; Flow Cytometry; Genes; Genetically Engineered Mouse; Glioblastoma; HSV vector; Herpesviridae; Human; Image; Immune; Immune Evasion; Immune checkpoint inhibitor; Immune response; Immune system; Immunization; Immunocompetent; Immunohistochemistry; In Vitro; Inbred BALB C Mice; Infection; Infiltration; Injections; Kinetics; Lateral; Lead; Ligands; Luciferases; Lymphocyte Subtypings; Lymphoid Cell; Lytic; MMP9 gene; Macrophage Colony-Stimulating Factor Receptor; Magnetic Resonance Imaging; Malignant Neoplasms; Malignant neoplasm of brain; Matrix Metalloproteinases; Mediating; Metalloproteases; MicroRNAs; Modeling; Mus; Mutate; NK Cell Activation; Natural Immunity; Natural Killer Cells; Nature; Nude Mice; Oncolytic; Oncolytic viruses; Patients; Performance; Phenotype; Population; Prevalence; Production; Recombinants; Recurrence; Regulatory T-Lymphocyte; Retroviridae; Simplexvirus; Sleeping Beauty; T-Cell Depletion; T-Lymphocyte; T-Lymphocyte and Natural Killer Cell; TIMP3 gene; TP53 gene; Testing; Therapeutic; Time; Toxic effect; Transforming Growth Factor beta; Treatment Efficacy; Treatment outcome; Tropism; Tumor Cell Line; Tumor Immunity; Tumor Tissue; Tumor-associated macrophages; Universities; Up-Regulation; Vegf Inhibitor; Virion; Virus Diseases; Washington; Xenograft procedure; adaptive immunity; anti-CTLA4; anti-PD-L1; anti-cancer; anti-tumor immune response; arm; base; brain cell; brain tissue; cell killing; combinatorial; cytokine; design; epidermal growth factor receptor VIII; exhaustion; experimental study; immune activation; immune checkpoint; immunoregulation; improved; in vivo; in vivo imaging; macrophage; mouse model; neonate; neoplastic cell; neovascularization; novel; oncolysis; oncolytic herpes simplex virus; oncolytic vector; overexpression; programmed cell death ligand 1; reconstitution; small hairpin RNA; tumor; tumor microenvironment; tumor progression; tumor-immune system interactions; vector

Oncolytic HSV vectors (oHSV) offer considerable promise in the treatment of Glioblastoma Multiforme (GBM). In previous studies we designed a new base vector (KGN-4:T124) that is blocked for replication in normal brain tissue by brain-specific cellular miR-124 targeting of ICP4 expression. This base vector was further retargeted (KGNE-4:T124) to achieve selective infection of GBM cells expressing EGFR/EGFRvIII. Arming of this vector with a matrix metalloproteinase (KGNE-4:T124-MMP9) enhanced vector spread and oncolysis in xenogeneic animal models. However, virolysis alone is unlikely to achieve complete elimination of tumor cells that can cause recurrence. Effective elimination of all tumor cells will require the induction of innate and adaptive anti-tumor immunity. Unfortunately, GBM down-regulates the cellular machinery needed to sustain activation of immune cells including NK, macrophages and cytotoxic T cells. Therefore we propose to test the hypothesis that GBM treatment with our oHSV base vectors, KGN-4:T124 and KGNE-4:T124, can be enhanced through vector arming with immunomodulatory genes that inhibit tumor progression and induce innate and adaptive anti-tumor immune responses. To this end, we will initially exploit a tumor cell line (BAGL1) derived from a GBM tumor induced in BALB/c mice by Sleeping Beauty transposition of genes encoding luciferase, anti-p53 shRNA, N-RasV12 and human EGFRvIII. These experiments will be extended to a genetically-induced GBM model developed by Dr. Eric Holland that avoids tumor cell injection. Our aims will seek to counteract three aspects of immune evasion in GBM: (i) ADAM17-mediated shedding of NK cell activators and reduced NK function, (ii) expression of the immune checkpoint molecules that lead to NK and T cell exhaustion, and (iii) the presence of M2 macrophages that contribute to an immunosuppressive tumor microenvironment (TME). In Aim 1, we will test the hypothesis that our unarmed base vectors, collectively referred to as KGN(E)-4:T124, will provide an effective GBM treatment in immune competent mice. We will determine the impact of vector dosing on intratumoral vector distribution, TME composition and animal survival, and assess the value of EGFR targeting. We will also examine the effect of prior HSV immunization on treatment outcomes. In Aim 2, we will test the hypothesis that KGN(E)- 4:T124 derivatives armed with TIMP-3 (ADAM17 inhibitor/VEGF-R2 antagonist) will improve therapy. We will evaluate the impact of TIMP-3 expression on NK cell activation, tumor spread and animal survival. In Aim 3, we will test the hypothesis that vectors armed with single chain antibody (scFv) checkpoint inhibitors (anti-PD-L1 or anti-CTLA4), alone or in combination with depletion of the immunosuppressive (M2) TAM population using BLZ945-mediated inhibition of colony-stimulating factor-1 receptor (CSF-1R), will produce effective anti-tumor immunity, particularly in HSV immune mice. Anti-tumor immunity will be established using a bi-lateral tumor model and confirmed by selective antibody-mediated T cell depletion.

溶瘤单纯疱疹病毒载体(OHSV)在多形性胶质母细胞瘤(GBM)的治疗中具有广阔的应用前景。 在以前的研究中，我们设计了一个新的碱基载体(KGN-4：T124)，该载体在正常大脑中被阻止复制 组织通过脑特异性细胞miR-124靶向ICP4的表达。该基本向量被进一步重定目标 (KGNE-4：T124)，实现对表达EGFR/EGFRvIII的GBM细胞的选择性感染。武装这艘航母 基质金属蛋白酶(KGNE-4：T124-MMP9)促进异种移植瘤的载体扩散和溶瘤作用 动物模型。然而，仅靠病毒裂解不太可能完全消除可能导致 复发。有效清除所有肿瘤细胞将需要诱导先天的和适应性的抗肿瘤 豁免权。不幸的是，GBM下调了维持免疫激活所需的细胞机制 细胞包括NK、巨噬细胞和细胞毒性T细胞。因此，我们建议检验GBM的假设 我们的OHSV基础载体KGN-4：T124和KGNE-4：T124的治疗可以通过载体武装来增强 带有免疫调节基因，可抑制肿瘤进展并诱导天然和适应性抗肿瘤免疫 回应。为此，我们首先开发了一种肿瘤细胞系(BAGL1)，该细胞系来自于在 睡美人转座BALB/c小鼠荧光素酶、抗P53 shRNA、N-RasV12和 人类EGFRvIII。这些实验将扩展到由Dr。 埃里克·霍兰德避免了肿瘤细胞注射。我们的目标将寻求在三个方面对抗豁免逃避 在GBM中：(I)ADAM17介导的NK细胞激活物的脱落和NK功能的降低，(Ii)表达 导致NK和T细胞耗尽的免疫检查点分子，以及(Iii)M2巨噬细胞的存在 这有助于免疫抑制的肿瘤微环境(TME)。在目标1中，我们将检验假设 我们的非武装基础载体，统称为KGN(E)-4：T124，将提供有效的GBM 对免疫能力强的小鼠的治疗。我们将确定载体剂量对肿瘤内载体的影响 分布、TME成分和动物存活，并评估EGFR靶向的价值。我们还将检查 既往接种HSV对治疗结果的影响。在目标2中，我们将检验假设Kgn(E)- 4：用TIMP-3(ADAM17抑制剂/血管内皮生长因子-R2拮抗剂)修饰的T124衍生物将改善治疗。我们会 评价TIMP-3表达对NK细胞活化、肿瘤扩散和动物存活的影响。在目标3中，我们 将测试携带单链抗体(ScFv)检查点抑制剂(抗PD-L1或 抗CTLA4)，单独或与免疫抑制(M2)人群联合使用 BLZ945介导的抑制集落刺激因子-1受体(CSF-1R)产生有效的抗肿瘤作用 免疫，特别是在HSV免疫的小鼠中。将使用双侧肿瘤建立抗肿瘤免疫 模型，并通过选择性抗体介导的T细胞耗竭得到证实。