Intracerebrally delivered EGFRvIII-targeted CARs for brain tumors

恶性原发脑瘤是导致儿童和年轻人癌症死亡的最常见原因。 并且比肾癌或黑色素瘤造成的死亡更多。胶质母细胞瘤(GBM)都是致命的， 目前的治疗是非特异性的，平均总生存期为15个月。相比之下， 免疫疗法承诺了一种精确的方法，大量证据表明，T细胞可以 即使肿瘤存在于大脑中，也要根除老鼠和人类的大型、成熟的肿瘤。 嵌合抗原受体(CARS)将抗体的可变区与T细胞信号转导部分结合起来 使T细胞活化具有抗体的靶向性，而不受MHC限制。临床试验 利用汽车已经展示了他们非凡的潜力。然而，严重的不良事件甚至 当这些汽车针对正常组织共有的抗原时，患者就会死亡。 EGFRvIII是表皮生长因子受体的一种肿瘤特异性突变，表达于GBM和 其他几种肿瘤。我们的实验室已经开发出用于T细胞转导的人和小鼠汽车 靶向肿瘤特异性EGFRvIII突变(EGFRvIII-CARS)以裂解EGFRvIII阳性的GBM。 EGFRvIII-CARS不应像靶向肿瘤的CARS那样导致对正常组织的直接杀伤- 相关的，但不是肿瘤特异性的抗原。然而，EGFRvIII是异质性表达的，在患者中 接种EGFRvIII特异性多肽疫苗后，肿瘤会因EGFRvIII的生长而复发 阴性的肿瘤细胞。此外，患有GBM的患者免疫功能高度受抑制，CARS可以 受宿主免疫抑制因子的抑制，存在于GBM微环境中，如分泌 转化生长因子β(TGFR-β)。虽然CAR疗法的有效性需要肿瘤特异性，但未来 CAR的开发必须解决肿瘤异质性和免疫抑制问题。在目标1中，我们将正式 检测脑内(IC)注射EGFRvIII-CARS的潜在毒性，并确定IC注射的CARS是否 保留在大脑中。在AIM 2中，我们将检查EGFRvIII-CAR疗法对肿瘤的影响 通过确定宿主T细胞中是否产生了表位扩散来实现异质性。在AIM 3中，我们将解决 通过我们实验室与基础科学家的持续合作，我们发现了 作为抗肿瘤细胞毒性T淋巴细胞反应关键抑制因子的microRNA miR-23a。MIR-23a是 转化生长因子-β上调和抑制T细胞miR-23a逆转转化生长因子-β诱导的T细胞免疫抑制 在小鼠模型中，细胞效应器功能和显著增强抗肿瘤效果。我们会调查的 抑制EGFRvIII-CAR转导的T细胞内miR-23a是否增强细胞毒作用 对宿主免疫抑制的抵抗。因此，这项建议解决了肿瘤特异性的关键问题， 异质性和免疫抑制，并提出了首次在人体试验中评估EGFRvIII的IC递送- 肾小球基底膜病患者的CARS。

Malignant primary brain tumors represent the most frequent cause of cancer death in children and young adults and account for more deaths than cancer of the kidney or melanoma. Glioblastoma (GBM) is uniformly lethal, and current therapy is non-specific and produces a median overall survival of <15 months. In contrast, immunotherapy promises an exquisitely precise approach, and substantial evidence suggests that T cells can eradicate large, well-established tumors in mice and humans even when tumors reside within the brain. Chimeric antigen receptors (CARs) combine the variable region of an antibody with T-cell signaling moieties to confer T-cell activation with the targeting specificity of an antibody without MHC-restriction. Clinical trials utilizing CARs have demonstrated their remarkable potential. However, severe adverse events and even patient deaths have occurred when these CARs have been directed against antigens shared by normal tissues. EGFRvIII is a tumor-specific mutation of the epidermal growth factor receptor that is expressed in GBMs and several other neoplasms. Our laboratory has developed human and murine CARs for the transduction of T cells targeting the tumor-specific EGFRvIII mutation (EGFRvIII-CARs) for the lysis of EGFRvIII positive GBM. EGFRvIII-CARs should not lead to direct killing of normal tissues as seen with CARs targeting tumor- associated, but not tumor-specific antigens. However, EGFRvIII is heterogeneously expressed and in patients vaccinated with an EGFRvIII-specific peptide vaccine, tumors recur as a result of outgrowth of the EGFRvIII negative tumor cells. Furthermore, patients with GBM are highly immunosuppressed and CARs can be restrained by host immunosuppressive factors present in the GBM microenvironment such as secreted transforming growth factor beta (TGF-β). While the potency of CAR therapy demands tumor-specificity, future CAR development must address tumor heterogeneity and immunosuppression. In Aim 1 we will formally examine potential toxicity of intracerebrally (IC) delivered EGFRvIII-CARs and determine if IC delivered CARs are retained within the brain. In AIM 2, we will examine the impact of EGFRvIII-CAR therapy on tumor heterogeneity through determining if epitope spreading is engendered in host T cells. In AIM 3, we will address immunosuppression though ongoing collaborations in our laboratory with basic scientists that have identified the micro-RNA miR-23a as a key inhibitor of anti-tumor cytotoxic T lymphocyte responses. miR-23a is upregulated by TGF-β and inhibition of miR-23a in T cells subverts TGF-β induced immunosuppression on T cell effector function and dramatically enhances anti-tumor efficacy within murine models. We will investigate whether miR-23a inhibition within EGFRvIII-CAR transduced T cells enhances cytotoxicity and confers resistance to host immunosuppression. This proposal therefore addresses the key issues of tumor-specificity, heterogeneity and immunosuppression and proposes the first in man trial evaluating IC delivery of EGFRvIII- CARs in patients with GBM.