Abstract
To investigate the delivery of next-generation macromolecular drugs, such as engineered proteins and mRNA-containing nanoparticles, there is an increasing push towards the use of physiologically relevant disease models that incorporate human cells and do not face ethical dilemmas associated with animal use. Here, we illustrate the versatility and ease of use of a microfluidic platform for studying drug delivery using high-resolution microscopy in 3D. Using this microfluidic platform, we successfully demonstrate the specific targeting of carbonic anhydrase IX (CAIX) on cells overexpressing the protein in a tumor-mimicking chip system using affibodies, with CAIX-negative cells and non-binding affibodies as controls. Furthermore, we demonstrate this system’s feasibility for testing mRNA-containing biomaterials designed to regenerate bone defects. To this end, peptide- and lipid-based mRNA formulations were successfully mixed with colloidal gelatin in microfluidic devices, while translational activity was studied by the expression of a green fluorescent protein. This microfluidic platform enables the testing of mRNA delivery from colloidal biomaterials of relatively high densities, which represents a first important step towards a bone-on-a-chip platform. Collectively, by illustrating the ease of adaptation of our microfluidic platform towards use in distinct applications, we show that our microfluidic chip represents a powerful and flexible way to investigate drug delivery in 3D disease-mimicking culture systems that recapitulate key parameters associated with in vivo drug application.
Highlights
While drug development efforts have traditionally focused on small-molecule drugs, during the last decade, there has been an increasing emphasis on biologics as novel therapeutic agents [1], which include various types of proteins, notably antibodies, oligonucleotides, viral gene therapies, and nanoparticle-based drug formulations
Our results demonstrate the versatility of our microfluidic platform for a microscopy-based investigation of the biology of drug delivery of engineered proteins and nanoparticles in highly tunable, in vivo-mimicking conditions
The anti-carbonic anhydrase IX (CAIX) affibody yielded a clear membrane staining in the CAIX-positive cells, which was absent in the negative cells
Summary
While drug development efforts have traditionally focused on small-molecule drugs, during the last decade, there has been an increasing emphasis on biologics as novel therapeutic agents [1], which include various types of proteins, notably antibodies, oligonucleotides, viral gene therapies, and nanoparticle-based drug formulations. Due to a highly exquisite binding specificity, targeting agents such as binding proteins are often fully or partially species-specific, negatively affecting extrapolation of results from animal to human in some applications [2,3]. This challenge extends to intracellular targets or intracellular effects of proteins with therapeutic activities that are expressed in the cytosol, e.g., by mRNA-based therapies. Additional challenges are low throughput, the inability to properly control for key parameters in animal experiments such as timing, dose, and monitoring of distribution, and ethical issues that are associated with the use of animals in scientific research, in particular if the animal models only partially recapitulate the human situation
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