Mechanisms of receptor-mediated transcytosis at the blood-brain barrier

In receptor-mediated transcytosis (RMT) of large therapeutics across the blood-brain barrier (BBB), the construct - a macromolecule or a larger carrier with therapeutic payload - binds a protein on brain capillary endothelial cells (BCEC), with internalization and release into the brain parenchyma. The construct's internalization into, trafficking across and release from, but also possible entrapment within BCEC are affected by its engineered properties whose optimization has helped derive insights into transport mechanisms at BCEC. Furthermore, advances in multi-omics, as well as large-scale screening and directed evolution campaigns have helped identify new targets for RMT at BCEC. In this perspective, I raise and reflect on some fundamental questions one can arrive at by comparing the engineered properties of BBB-targeted constructs and the properties of different target proteins. These questions concern the underlying, transcytosis-promoting factors that the optimization of constructs' engineered properties appears to converge on, the precise role of target proteins in RMT, the different mechanisms through which these targets may mediate construct trafficking, and the tentative criteria for target selection on BCEC. Based on these considerations I propose several scenarios and strategies to interfere with the construct's trafficking for more efficient internalization, transport through the endosomal network toward the abluminal membrane, and release from BCEC, both for smaller macromolecules and for larger carriers. • Interplay of construct properties may suggest overarching patterns governing RMT. • Properties of some RMT targets may indicate unconventional transport mechanisms. • Construct's trafficking may be decoupled from target and manipulated artificially. • Specificity of brain endothelial cells may have non-canonical determinants. • Endosomal release, rather than escape of carriers may improve cargo transcytosis.