UC San Diego

Engineered bacteria are emerging as programmable therapeutics capable of colonizing solid tumors and delivering therapeutic payloads with spatial precision. Yet most current designs rely on fixed genetic architectures that cannot adapt to the heterogeneous conditions of the tumor microenvironment. Here, coupled plasmid systems offer a generalizable framework for encoding adaptive behavior into bacterial therapeutics. In these systems, two constructs sharing an origin of replication compete for replication machinery, enabling population-level phenotypic flexibility that is directly responsive to environmental context. Using a microfluidic gradient platform, we demonstrate that coupled plasmid strains undergo stable spatial differentiation in response to opposing chemical gradients, with a stochastic model confirming that copy number competition is sufficient to generate this organization without additional regulatory circuitry. Using a modular tissue-scale testbed designed to recapitulate tumor microenvironmental conditions, we find that oxygen availability drives spatially graded synchronized lysis circuit (SLC) triggering while extracellular matrix architecture modulates AHL-mediated quorum coupling across tissue-relevant distances. Integrating these principles, we couple lactate-responsive lysis with the SLC within a coupled-origin framework, producing a strain that autonomously switches between sensing and delivery phenotypes as a function of local lactate concentration and releases therapeutic payload in a cyclical, self-regulated manner. Together these results establish that coupled plasmid systems can translate environmental heterogeneity into organized therapeutic function, offering a generalizable architecture for programming adaptive behavior into living medicines.