Precise cell targeting is challenging because most mammalian cell types lack a single surface marker that distinguishes them from other cells. A solution would be to target cells based on specific combinations of proteins present on their surfaces. We design colocalization-dependent protein switches (Co-LOCKR) that perform AND, OR, and NOT Boolean logic operations. These switches activate through a conformational change only when all conditions are met, generating rapid, transcription-independent responses at single-cell resolution within complex cell populations. We implement AND gates to redirect T cell specificity against tumor cells expressing two surface antigens while avoiding off-target recognition of single-antigen cells, and 3-input switches that add NOT or OR logic to avoid or include cells expressing a third antigen. Thus, de novo designed proteins can perform computations on the surface of cells, integrating multiple distinct binding interactions into a single output.
Influenza virus neuraminidase (NA) is a major antiviral drug target and has recently reemerged as a key target of antibody-mediated protective immunity. Here we show that recombinant NAs across non-bat subtypes adopt various tetrameric conformations, including an “open” state that may help explain poorly understood variations in NA stability across viral strains and subtypes. We use homology-directed protein design to uncover the structural principles underlying these distinct tetrameric conformations and stabilize multiple recombinant NAs in the “closed” state, yielding two near-atomic resolution structures of NA by cryo-EM. In addition to enhancing thermal stability, conformational stabilization improves affinity to protective antibodies elicited by viral infection, including antibodies targeting a quaternary epitope and the broadly conserved catalytic site. Stabilized NAs can also be integrated into viruses without affecting fitness. Our findings provide a deeper understanding of NA structure, stability, and antigenicity, and establish design strategies for reinforcing the conformational integrity of recombinant NA proteins.
Despite remarkable
advances over the past several decades, many
therapeutic nanomaterials fail to overcome major in vivo delivery
barriers. Controlling immunogenicity, optimizing biodistribution,
and engineering environmental responsiveness are key outstanding delivery
problems for most nanotherapeutics. However, notable exceptions exist
including some lipid and polymeric nanoparticles, some virus-based
nanoparticles, and nanoparticle vaccines where immunogenicity is desired.
Self-assembling protein nanoparticles offer a powerful blend of modularity
and precise designability to the field, and have the potential to
solve many of the major barriers to delivery. In this review, we provide
a brief overview of key designable features of protein nanoparticles
and their implications for therapeutic delivery applications. We anticipate
that protein nanoparticles will rapidly grow in their prevalence and
impact as clinically relevant delivery platforms.
Targeting ligands are used in drug delivery to improve drug distribution to desired cells or tissues and to facilitate cellular entry. In vivo biopanning, whereby billions of potential ligand sequences are screened in biologically-relevant and complex conditions, is a powerful method for identification of novel target ligands. This tool has impacted drug delivery technologies and expanded our arsenal of therapeutics and diagnostics. Within this review we will discuss current in vivo panning technologies and ways that these technologies can be improved to advance next-generation drug delivery strategies.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.