Bacterial delivery of pro-apoptotic proteins in advanced hepatocellular carcinoma.

摘要 目前还没有治愈不能切除的肝细胞癌的方法。 这些患者的预后令人沮丧。在目前的标准护理治疗方法下，中位数 生存时间充其量是20个月。我们建议开发一种安全有效的肝癌治疗方法，该方法采用 沙门氏菌选择性地向肿瘤递送活性caspase-3。激活刽子手Caspase，最终 致力于细胞凋亡的步骤，绕过癌症避免细胞死亡的机制。系统性 运送活性半胱氨酸天冬氨酸酶是不可行的，因为它会引起严重的副作用，而且很难被吸收。 进入细胞。相比之下，与健康组织相比，沙门氏菌在肝癌组织中的积聚比例为100,000：1 因此，它可以将高度特异性的蛋白质输送到癌细胞，而全身副作用最小。 这项研究的概念证明是通过证明1)细菌输送系统 能将蛋白特异性地运送到癌细胞和2)细菌运送活性的Casp-3诱导细胞死亡 并能减少小鼠皮下肝细胞癌的肿瘤体积。在这项提案中，我们想要 确定细菌释放活性CASP-3(EBT-002)在小鼠体内的安全性和有效性 标准的护理和调查结合免疫检查点封锁。在目标1中，我们将量化 EBT-002对小鼠皮下肝癌细胞凋亡及肿瘤体积的影响 原位肝肿瘤模型。在目标2中，我们将确定EBT-002的最大耐受量(MTD)和 在剂量递增研究中研究急性毒性，以及在肝硬变背景下的肝脏毒性。我们会 测量这种疗法在荷瘤小鼠体内的生物分布，以确定非靶标细菌的聚集。 我们还将确定最大有效剂量。我们假设，在高细菌剂量下，疗效是 与剂量无关。这将能够减少剂量，同时保持在 MTD。在目标3中，我们将确定EBT-002对肿瘤中T细胞浸润和肿瘤增殖的影响 PD-L1高表达和低表达。这将与抗PD-L1抗体一起进行测试，以评估 如果这种联合治疗能提高两种疗法对有反应和无反应肿瘤的疗效 检查点抑制药。 这一提议的成功结果是EBT-002作为一种新的治疗方法发展的第一步 肝细胞癌。这种疗法的进一步发展将在第二阶段SBIR中进行，以获得必要的数据 Ind应用程序。这项研究将建立一种平台技术，直接将蛋白质输送到细胞中 在信号通路中的关键点，并将在多种癌症中有效。

Summary There are no curative treatments for patients with unresectable hepatocellular carcinoma (HCC) at this time. The prognosis for these patients is dismal. With current standard of care treatment methods, the median survival time is, at best, 20 months. We propose to develop a safe and effective therapy for HCC that employs Salmonella to deliver active caspase-3 selectively to tumors. Activating executioner caspases, the final committed steps of apoptosis, circumvent the mechanisms that cancers acquire to avoid cell death. Systemic delivery of active caspases is unfeasible because it would induce severe side effects and be poorly taken up into cells. In contrast, Salmonella accumulate in HCC in ratios of 100,000 to 1 compared to healthy tissues and, as a result, can deliver proteins with high specificity to cancer cells with minimal systemic side effects. The proof of concept for this study has been established by demonstrating that 1) a bacterial delivery system can delivery proteins specifically into cancer cells and 2) bacterial delivery of active casp-3 induces cell death in cancer cells and reduces tumor volume in subcutaneous HCC in mice. In this proposal, we want to determine the safety and efficacy of bacterially delivered active casp-3 (EBT-002) in mice compared to standard of care and investigate the combination with immune checkpoint blockade. In Aim 1, we will quantify the effect of EBT-002 on the activation of apoptosis in subcutaneous HCC tumors and on tumor volume in an orthotopic liver tumor model. In Aim 2, we will determine the maximum tolerated dose (MTD) of EBT-002 and study acute toxicity in a dose-escalation study as well as liver toxicity in a cirrhotic liver background. We will measure the biodistribution of this therapy in mice with tumors to determine off-target bacterial accumulation. We will also determine the maximum effective dose. We hypothesize that at high bacterial dosage, efficacy is independent of dose. This would enable a reduction of dose, while maintaining the same efficacy measured at MTD. In Aim 3, we will determine the effect of EBT-002 on T-cell infiltration and tumor proliferation in tumors with high and low PD-L1 expression. This will be tested in combination with anti-PD-L1 antibodies, to evaluate if this combination can increase efficacy of both therapies in tumors that are responsive and non-responsive to checkpoint inhibitors. The successful outcome of this proposal is the first step in the development of EBT-002 as a new therapy for HCC. Further development of this therapy will be pursued in a Phase II SBIR to obtain the necessary data for an IND application. This research will establish a platform technology to directly deliver proteins into cells at critical points in signaling pathways and will be effective in a wide array of cancers.