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 in vivo using exogenously administered nanoparticle sensors and ex vivo 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.
Bibliographical noteFunding Information:
The authors declare the following competing financial interest(s): L.N. holds equity in and is the scientific founder of Circuit-Bio and receives sponsored research funding from FutuRx. S.N.B. holds equity in Glympse Bio, Satellite Bio, Cend Therapeutics, and Catalio Capital; is a director at Vertex; consults for Moderna; and receives sponsored research funding from Johnson & Johnson.
This study was supported in part by a Koch Institute Support Grant P30-CA14051 from the National Cancer Institute, a Core Center Grant P30-ES002109 from the National Institute of Environmental Health Sciences, an Amar G. Bose Research Grant, the Virginia and D.K. Ludwig Fund for Cancer Research, and the Koch Institute’s Marble Center for Cancer Nanomedicine. J.H. thanks Liang Hao for her generous help on nanosensor synthesis. A.P.S. acknowledges support from the NIH Molecular Biophysics Training Grant NIH/NIGMS T32 GM008313 and the National Science Foundation Graduate Research Fellowship. S.N.B is a Howard Hughes Medical Institute Investigator.
© 2021 The Authors. Published by American Chemical Society.
- activity probes
- synthetic biology