Sample-efficient identification of high-dimensional antibiotic synergy with a normalized diagonal sampling design

Antibiotic resistance is an important public health problem. One potential solution is the development of synergistic antibiotic combinations, in which the combination is more effective than the component drugs. However, experimental progress in this direction is severely limited by the number of samples required to exhaustively test for synergy, which grows exponentially with the number of drugs combined. We introduce a new metric for antibiotic synergy, motivated by the popular Fractional Inhibitory Concentration Index and the Highest Single Agent model. We also propose a new experimental design that samples along all appropriately normalized diagonals in concentration space, and prove that this design identifies all synergies among a set of drugs while only sampling a small fraction of the possible combinations. We applied our method to screen two- through eight-way combinations of eight antibiotics at 10 concentrations each, which requires sampling only 2,560 unique combinations of antibiotic concentrations.Author summary: Antibiotic resistance is a growing public health concern, and there is an increasing need for methods to combat it. One potential approach is the development of synergistic antibiotic combinations, in which a mixture of drugs is more effective than any individual component. Unfortunately, the search for clinically beneficial drug combinations is severely restricted by the pace at which drugs can be screened. To date, most studies of combination therapies have been limited to testing only pairs or triples of drugs. These studies have identified primarily antagonistic drug interactions, in which the combination is less effective than the individual components. There is an acute need for methodologies that enable screening of higher-order drug combinations, both to identify synergies among many drugs and to understand the behavior of higher-order combinations. In this work we introduce a new paradigm for combination testing, the normalized diagonal sampling design, that makes identifying interactions among eight or more drugs feasible for the first time. Screening d drugs at m different combinations requires m ⋅ 2d samples under our design as opposed to md under exhaustive screening, while provably identifying all synergies under mild assumptions about antibiotic behavior. Scientists can use our design to quickly screen for antibiotic interactions, accelerating the pace of combination therapy development.