Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States.
Harvard-MIT Division of Health Sciences and Technology, Institute for Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States.
ACS Synth Biol. 2021 Sep 17;10(9):2231-2242. doi: 10.1021/acssynbio.1c00133. Epub 2021 Aug 31.
The integration of nanotechnology and synthetic biology could lay the framework for new classes of engineered biosensors that produce amplified readouts of disease states. As a proof-of-concept demonstration of this vision, here we present an engineered gene circuit that, in response to cancer-associated transcriptional deregulation, expresses heterologous enzyme biomarkers whose activity can be measured by nanoparticle sensors that generate amplified detection readouts. Specifically, we designed an AND-gate gene circuit that integrates the activity of two ovarian cancer-specific synthetic promoters to drive the expression of a heterologous protein output, secreted Tobacco Etch Virus (TEV) protease, exclusively from within tumor cells. Nanoparticle probes were engineered to carry a TEV-specific peptide substrate in order to measure the activity of the circuit-generated enzyme to yield amplified detection signals measurable in the urine or blood. We applied our integrated sense-and-respond system in a mouse model of disseminated ovarian cancer, where we demonstrated measurement of circuit-specific TEV protease activity both using exogenously administered nanoparticle sensors and using quenched fluorescent probes. We envision that this work will lay the foundation for how synthetic biology and nanotechnology can be meaningfully integrated to achieve next-generation engineered biosensors.
纳米技术和合成生物学的融合可能为新型工程生物传感器奠定框架，这些传感器可以放大疾病状态的读出信号。作为这一愿景的概念验证演示，我们在这里提出了一种工程基因电路，该电路响应与癌症相关的转录失调，表达异源酶生物标志物，其活性可以通过纳米颗粒传感器测量，这些传感器产生放大的检测读数。具体来说，我们设计了一个与门基因电路，该电路整合了两个卵巢癌特异性合成启动子的活性，以驱动异源蛋白输出，即分泌型烟草蚀纹病毒（TEV）蛋白酶的表达，仅来自肿瘤细胞内。纳米颗粒探针经过设计可携带 TEV 特异性肽底物，以测量电路产生的酶的活性，从而产生可在尿液或血液中测量的放大检测信号。我们在转移性卵巢癌的小鼠模型中应用了我们的集成感知和响应系统，在该模型中，我们通过外源性给予的纳米颗粒传感器和猝灭荧光探针来证明电路特异性 TEV 蛋白酶活性的测量。我们设想，这项工作将为合成生物学和纳米技术如何进行有意义的整合以实现下一代工程生物传感器奠定基础。
