Proteases as Biological Bits for Programmable Medicine

Abstract

12Engineered biocircuits that interface with living systems as plug-and-play constructs may enable 13 new applications for programmable therapies and diagnostics. We create biological bits (bbits) 14 using proteasesa family of pleiotropic, promiscuous enzymesto construct the biological 15 equivalent of Boolean logic gates, comparators and analog-to-digital converters. We use these 16 modules to write a cell-free bioprogram that can combine with bacteria-infected blood, quantify 17 infection burden, and then calculate and unlock a selective drug dose. Inspired by probabilistic 18 computing, we leverage multi-and common-target protease promiscuity as the biological analog 19 of superposition to program three probabilistic bbits that solve all implementations of the two-bit 20 oracle problem, Learning Parity with Noise. Treating a network of dysregulated proteases in a 21 living animal as an oracle, we use this algorithm to resolve the probability distribution of 22 coagulation proteases in vivo, allowing diagnosis of pulmonary embolism with high sensitivity and 23 specificity (AUROC = 0.92) in a mouse model of thrombosis. Our results demonstrate that 24 protease activity can be programmed in cell-free systems to carry out classical and probabilistic 25

Monitoring Proteolytic Activity in Real Time: A New World of Opportunities for Biosensors

“…Another interesting vision for proteolytic sensors in real time suggests their integration in logic gates [56]. Also, Kwong et al [57,58] used nonreal-time proteolytic activity as biologic bits and integrated them in the field of programmable medicine. By merging these concepts, one can imagine a machine analyzing the readout in real time and giving an immediate and proper responsefor example, to stop/start the administration of a drugin a feedback scheme and without the constant supervision of a healthcare professional.…”