AND-gated Synthetic Biomarkers for Early Detection of Liver Metastasis

项目总结 合成生物学的进展将在塑造癌症诊断的未来方面发挥基础性作用 更早和更具体的疾病检测。例如，细菌等全细胞生物传感器已经被 通过基因工程实现复杂的功能，如检测临床相关的信号放大 人体尿液和血清中的生物标志物。在哺乳动物细胞中，利用 布尔逻辑已经被证明用于工程T细胞疗法的多路控制，从而增加了 肿瘤感知的特异性和减少全身毒性。这些进展突显了合成技术的前景 生物学当应用于癌症时，然而这些策略中的绝大多数依赖于基因电路并利用 非哺乳动物的蛋白质成分。这样的电路很复杂，会引起安全和免疫原性的问题 要求监管部门批准，特别是在体内早期癌症检测的情况下，重复给药 可能需要大量的生物传感器来监测新生的疾病。这项建议旨在发展一类新的 诊断学称为与门合成生物标记物，用于癌症转移的早期检测。合成的 生物标记物是一类新兴的可激活生物传感器，设计用于管理 系统地查询早期疾病的站点，并利用肿瘤依赖的激活机制，例如 失控的蛋白酶活性，以推动记者的生产。然后这些记者就可以被检测到 从血液、尿液或其他体液样本中提取。蛋白水解酶在整个生命周期中扮演着重要的生物学角色 转移的主要标志，特别是通过催化不可逆转的分子放大器 多肽键的水解，使单一的蛋白酶可以周转数千种底物。与门控合成 生物标志物将应用于结直肠癌(CRC)肝转移的早期检测。虽然肝脏是一种 原发结直肠癌的常见转移部位，孤立肝转移的区域切除可导致 潜在的治疗效果。然而，早期发现结直肠癌肝转移的大小 对治疗(1-2 mm)的反应通过CT和FDA批准的放射成像仍然具有挑战性 血液检测，如癌胚抗原(CEA)检测。为结直肠癌设计和门控合成生物标记物 肝转移，将根据不同的RNA表达选择成对的蛋白酶组合 结直肠癌肝转移与正常肝组织的比较。多变量数学模型将发展为 了解与实验结果相比，设计参数如何增强特异性和协作性。 结直肠癌肝转移的同种和异种移植模型将用于临床前验证研究，以允许 对照CT和CEA进行基准测试。这项提议将为更早发现癌症奠定基础 可编程合成生物标记物的转移。

PROJECT SUMMARY Advances in synthetic biology will play a fundamental role in shaping the future of cancer diagnostics toward earlier and more specific detection of disease. For example, whole-cell biosensors such as bacteria have been genetically engineered to perform complex functions such as signal amplification to detect clinically relevant biomarkers in human urine and serum. In mammalian cells, sense-and-respond components that employ Boolean logic have been demonstrated for multiplexed control of engineered T cell therapies, thereby increasing the specificity of tumor sensing and reducing systemic toxicity. These advances highlight the promise of synthetic biology when applied to cancer, yet the vast majority of these strategies rely on genetic circuits and make use of non-mammalian protein components. Such circuits are complex and raise safety and immunogenicity concerns for regulatory approval, especially in the context of in vivo early cancer detection where repeated administrations of biosensors are likely needed to monitor for nascent disease. This proposal seeks to develop a new class of diagnostics called AND-gated synthetic biomarkers for early detection of cancer metastasis. Synthetic biomarkers are an emerging class of activatable biological sensors that are designed to be administered systemically, query sites of early disease, and harness tumor-dependent activation mechanisms, such as dysregulated protease activity, to drive production of a reporter. These reporters can then be detected noninvasively from blood, urine, or other bodily fluid samples. Proteases play key biological roles across the major hallmarks of metastasis and are particularly potent molecular amplifiers by catalyzing the irreversible hydrolysis of peptide bonds, allowing a single protease to turnover thousands of substrates. AND-gated synthetic biomarkers will be applied for early detection of colorectal cancer (CRC) liver metastasis. Although the liver is a common site for metastatic spread from primary CRC, regional resection of liver-isolated metastases can lead to potentially curative results. Yet early detection of CRC liver metastases at a size when they are most responsive to therapy (1–2 mm) remains challenging by radiographic imaging such as CT and FDA-approved blood test such as the carcinoembryonic antigen (CEA) test. To design AND-gated synthetic biomarkers for CRC liver metastasis, pairwise combinations of proteases will be selected based on differential RNA expression in CRC liver metastases compared to healthy liver tissue. Multivariate mathematical models will be developed to understand how design parameters enhance specificity and cooperativity compared to experimental results. Syngeneic and xenograft models of CRC liver metastasis will be used for preclinical validation studies to allow benchmarking against CT and CEA. This proposal will lay the groundwork for earlier detection of cancer metastasis by programmable synthetic biomarkers.