Design and characterization of bacterial population dynamics in solid tumor models

实体瘤模型中细菌种群动态的设计和表征

• 批准号: 10212134
• 负责人: JEFF M HASTY
• 金额: $ 64.29万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10212134
• 关键词: Affect; Ally; Anaerobic Bacteria; Animal Model; Bacteria; Bacterial Genes; Behavior; Biological Models; Biosensing Techniques; Biosensor; CCL21 gene; CRISPR/Cas technology; Cancer Model; Colonoscopy; Colorectal; Colorectal Cancer; Colorectal Neoplasms; Complex; Cues; Cytolysis; Data; Development; Disease; Drug Delivery Systems; Engineered Probiotics; Engineering; Environment; Escherichia coli; Future; Gases; Gel; Gene Cluster; Gene Expression; Genes; Genetic; Genome; Glucose; Goals; Growth; Growth Factor; Harvest; Health; High Fat Diet; Histology; Human; Human body; Hypoxia; Image; Imaging technology; Immunologic Surveillance; In Situ Hybridization; In Vitro; Intercellular Fluid; Intestines; Lead; Liquid substance; Liver; Luciferases; Maintenance; Malignant Neoplasms; Malignant neoplasm of gallbladder; Malignant neoplasm of liver; Malignant neoplasm of lung; Malignant neoplasm of ovary; Malignant neoplasm of pancreas; Measures; Methods; Microbe; Microfluidic Microchips; Microscopy; Modeling; Molecular; Monitor; Mucous Membrane; Mus; Nutrient; Optical reporter; Oral; Organoids; Pathologic; Penetration; Play; Population; Population Dynamics; Primary carcinoma of the liver cells; Probiotics; Property; Reporting; Research; Resolution; Role; Safety; Solid Neoplasm; Specificity; Structure; System; Techniques; Technology; Testing; Time; Tissues; Ultrasonography; Vesicle; base; clinically relevant; design; design and construction; drug production; effectiveness study; environmental change; genome wide screen; genome-wide; high resolution imaging; human disease; improved; in vivo; in vivo Model; luminescence; malignant breast neoplasm; malignant mouth neoplasm; microbial; microfluidic technology; mouse model; neoplastic; nonalcoholic steatohepatitis; novel; pre-clinical; preference; promoter; quorum sensing; response; screening; synthetic biology; tool; tumor; tumor growth; tumor microenvironment

Project Summary It is increasingly clear that bacteria play an important role in human health. While it is natural to focus on how intestinal bacteria affect disease, intriguing findings have elucidated the extent to which bacteria inhabit solid tumors. Microbes have been detected in lung, pancreatic, breast, oral, gallbladder, ovarian, liver, and colorectal cancers. Localization has been ascribed to several mechanisms, including preference for anaerobic or facultative anaerobic bacteria to grow in the hypoxic core of tumors, presence of bacterial nutrients, lack of immune surveil- lance, and leakiness of the often poorly structured vasculature surrounding neoplastic tissue. This tendency for localization to solid tumors suggests that bacteria could be engineered for precise and robust drug production and delivery from within the solid tumor environment. This dovetails with 20 years of progress in synthetic biology, which has tended to focus on microbial engineering. However, information on how the tumor microenvironment affects bacterial growth is largely unknown. The microenvironment will affect bacterial gene expression that ul- timately underlies the functionality of engineered therapies, and it is difficult to imagine a predictive framework for engineered bacterial therapies without a quantitative understanding of how bacteria react to the environment of a growing tumor. We will use a probiotic strain of E. coli with an established safety record to develop a novel class of biosensors to noninvasively investigate bacterial growth in the tumor microenvironment. Initially, we will develop lysis-based biosensors that respond to specific tumor environment targets: hypoxia, pH, glucose, and lactate (Aim 1). We will also engineer an inducible quorum sensing (QS) system that enables external control of bacterial population dynamics, including the ability to eliminate a specific strain whenever desired (Aim 1). Together these strains will allow us to modulate and monitor population dynamics in vivo, enabling both sens- ing of the local environment and maintenance of an external control switch. We will test these strains using an established in vitro organoid model (Aim 2) and in two clinically relevant animal models for solid tumor growth. Additionally, we will use our previously developed dynOMICS technology to screen tumor extract from the two animal models and construct a second suite of biosensors for monitoring the tumor environment (Aim 2). These biosensors will then be tested in the animal models. We will visualize bacterial populations in a colorectal tumor model with bacteria that are engineered to produce luciferase in order to monitor colony dynamics using our es- tablished methods (Aim 3). We will also build on recently reported technology whereby bacteria are modified for use with ultrasound through addition of gas vesicles that permit high resolution imaging of the engineered bac- teria. We will use the ultrasound method to investigate NASH-induced hepatocellular carcinoma (HCC) where a high-fat diet is used to induce HCC at 20 weeks in mice (Aim 4). This project will quantitatively characterize how bacterial strains sense, respond, and grow in the tumors. The results will establish a platform for future exploration of therapies that are produced and delivered by bacteria that grow within solid tumors.

项目概要 越来越明显的是，细菌在人类健康中发挥着重要作用。虽然很自然地会关注如何 肠道细菌影响疾病，有趣的发现阐明了细菌在固体中栖息的程度 肿瘤。已在肺、胰腺、乳房、口腔、胆囊、卵巢、肝脏和结直肠中检测到微生物 癌症。定位被归因于多种机制，包括对厌氧或兼性的偏好 厌氧细菌在肿瘤缺氧的核心生长，存在细菌营养物质，缺乏免疫监视 以及肿瘤组织周围结构不良的脉管系统的渗漏。这种倾向对于 实体瘤的定位表明细菌可以被设计用于精确和稳健的药物生产 从实体瘤环境内递送。这与合成生物学 20 年的进展相吻合， 它倾向于关注微生物工程。然而，关于肿瘤微环境如何影响的信息 影响细菌生长的情况在很大程度上尚不清楚。微环境会影响细菌基因表达 是工程疗法功能的密切基础，很难想象一个预测框架 用于工程细菌疗法，无需定量了解细菌对环境的反应 正在生长的肿瘤。我们将使用具有既定安全记录的大肠杆菌益生菌菌株来开发一种新型 一类生物传感器，用于无创地研究肿瘤微环境中的细菌生长。最初，我们 将开发基于裂解的生物传感器，可响应特定的肿瘤环境目标：缺氧、pH、葡萄糖、 和乳酸（目标 1）。我们还将设计一个可诱导群体感应 (QS) 系统，该系统可实现外部控制 细菌种群动态，包括在需要时消除特定菌株的能力（目标 1）。 这些菌株将使我们能够调节和监测体内的群体动态，从而使两种感觉 本地环境的维护和外部控制开关的维护。我们将使用 建立了体外类器官模型（目标 2）和两个临床相关的实体瘤生长动物模型。 此外，我们将使用我们之前开发的dynOMICS技术来筛选两种肿瘤提取物 动物模型并构建第二套生物传感器来监测肿瘤环境（目标 2）。这些 然后生物传感器将在动物模型中进行测试。我们将可视化结直肠肿瘤中的细菌群 使用经过工程改造产生荧光素酶的细菌模型，以便使用我们的 es 监测菌落动态 制定方法（目标 3）。我们还将以最近报道的技术为基础，对细菌进行修饰 通过添加气体囊泡与超声波一起使用，允许工程化 bac- 的高分辨率成像 泰里亚。我们将使用超声波方法来研究 NASH 诱发的肝细胞癌 (HCC)，其中 使用高脂肪饮食在 20 周时诱导小鼠发生 HCC（目标 4）。该项目将定量表征 细菌菌株如何在肿瘤中感知、反应和生长。研究结果将为未来建立一个平台 探索由实体瘤内生长的细菌产生和传递的疗法。