Building Synthetic Biosensors Using Red Blood Cell Proteins

As the use of engineered cell therapies expands from
pioneering
efforts in cancer immunotherapy to other applications, an attractive
but less explored approach is the use of engineered red blood cells
(RBCs). Compared to other cells, RBCs have a very long circulation
time and reside in the blood compartment, so they could be ideally
suited for applications as sentinel cells that enable in situ sensing and diagnostics. However, we largely lack tools for converting
RBCs into biosensors. A unique challenge is that RBCs remodel their
membranes during maturation, shedding many membrane components, suggesting
that an RBC-specific approach may be needed. Toward addressing this
need, here we develop a biosensing architecture built on RBC membrane
proteins that are retained through erythropoiesis. This biosensor
employs a mechanism in which extracellular ligand binding is transduced
into intracellular reconstitution of a split output protein (including
either a fluorophore or an enzyme). By comparatively evaluating a
range of biosensor architectures, linker types, scaffold choices,
and output signals, we identify biosensor designs and design features
that confer substantial ligand-induced signal in vitro. Finally, we demonstrate that erythroid precursor cells engineered
with our RBC-protein biosensors function in vivo. This study establishes a foundation for developing RBC-based biosensors
that could ultimately address unmet needs including noninvasive monitoring
of physiological signals for a range of diagnostic applications.