(253f) Perfecting Bacterial Tumor Treatment Using Microfluidic Bioreactors

One of the major limitations of current cancer chemotherapeutics is ineffective penetration within the highly heterogeneous tumor tissue. Active, as opposed to passive transport of the therapeutic agent will overcome this limitation. Engineered bacteria possess the ability to actively transport deep into the tumor tissue, and can be genetically manipulated to target desired regions within the tumor and deliver therapeutic payloads, thus providing new inroads into tumor treatment. Three-dimensional in-vitro models of tumor tissue would largely facilitate testing therapeutic bacteria for their efficacy. However, the rapid growth rate of bacteria in currently available batch systems makes it impossible to study their effect on mammalian tissue over physiologically relevant time scales. To address this deficiency, we developed a microfluidic continuous flow-through device as an in-vitro model of tumor tissue. Multicellular tumor spheroids were flown into elongated micro-chambers within the device and maintained with mild perfusion from one side, thus creating linear nutrient gradients within the tissue and forming micron-scale in-vitro tumors. Tumors thus formed were subjected to hour-long plugs of a constitutively GFP-expressing wild type strain of Salmonella Typhimurium SL1344. An active Caspase-3 fluorescent marker was used to detect apoptotic regions within the tumors. Time-lapse microscopy was employed to monitor a) the accumulation of bacteria within the tumor and b) the apoptosis induced hence. Accumulation of bacteria significantly enhanced apoptosis within tumors, demonstrating their innate therapeutic ability. Moreover, the magnitude of bacteria-induced apoptosis was found to be microenvironment-specific, and was quantified by observing different regions within the heterogeneous tumor tissue. To facilitate higher throughput testing of various strains of therapeutic bacteria, an array of such in-vitro tumors was developed. To better understand the mechanism of bacterial action, high magnification microscopy was used to track the path of individual bacteria into mammalian cells. The microfluidic device developed here will accelerate the process of developing the perfect bacterial therapeutic for tumor treatment.