Storming the gate: New approaches for targeting the dynamic tight junction for improved drug delivery

Abstract

As biologics used in the clinic outpace the number of new small molecule drugs, an important challenge for their efficacy and widespread use has emerged, namely tissue penetrance. Macromolecular drugs - bulky, high-molecular weight, hydrophilic agents - exhibit low permeability across biological barriers. Epithelial and endothelial layers, for example within the gastrointestinal tract or at the blood-brain barrier, present the most significant obstacle to drug transport. Within epithelium, two subcellular structures are responsible for limiting absorption: cell membranes and intercellular tight junctions. Previously considered impenetrable to macromolecular drugs, tight junctions control paracellular flux and dictate drug transport between cells. Recent work, however, has shown tight junctions to be dynamic, anisotropic structures that can be targeted for delivery. This review aims to summarize new approaches for targeting tight junctions, both directly and indirectly, and to highlight how manipulation of tight junction interactions may help usher in a new era of precision drug delivery.

Fig. 1.. Tight junctions as dynamic structures limiting biologic delivery.

A) TJs, AJs and desmosomes line the lateral surface of epithelial cells and contribute to the establishment and maintenance of epithelial barrier function. Junctions are composed of transmembrane proteins, adaptor proteins, and cytoskeletal elements, among others. B) The TJ as a dynamic and anisotropic junction. i) Interclaudin interference can affect TJ strand architecture. Claudin-2 forms strands in the absence of claudin-4. However, in the presence of claudin-4, claudin-2 strands are disrupted. ii) TJ leaks occur under mechanical stress, which correlates with Zonula Occludens-1 (ZO-1) breaks during junction elongation. Leaks are quickly repaired by RhoA, which causes the accumulation of F-actin and myosin II along with contraction of the junction and reunification of ZO-1 along the membrane. iii) ZO-1 phase separates in non-junctional pools. Phosphorylated ZO-1, however, is less prone to phase separation. iv) The weak interaction between ZO-1 and actin at epithelial TJs gives rise to proper claudin organization at the membrane. v) Local membrane composition strongly impacts TJ formation. Cholesterol depletion at the membrane induces claudin endocytosis, abolishing TJ function.

Fig. 2.. Expression of claudin genes by tissue type.

RNA consensus data, which contains RNA transcript expression levels observed in 54 different human tissues from HPA and GTEx, was obtained from the Human Protein Atlas website [56]. This data was previously normalized by HPA, and normalized gene expression values were calculated as the maximum nTPM value for each gene in the two data sources. The datasets were then screened and streamlined to obtain a list containing the normalized RNA expression levels for 24 claudin (CLDN) genes using Python. Matrix visualization platform by the Broad Institute, Morpheus [57], was used to visualize claudin gene expression by tissue type.

Fig. 3.. Small molecules and peptides for TJ modulation.

A) Direct approaches to modulate barrier function include targeting occludin and claudin at the TJ, inhibiting ZO-1-claudin PPIs and targeting a splice variant of myosin light chain kinase (MLCK), MLCK1, with small molecule inhibitors and peptide binders. B) Barrier function is also indirectly modulated by peptides and small molecules that act on E-cadherin, F-actin, or that target signaling proteins that influence TJ function, such as MLCK and the Rho family of small GTPases.

Fig. 4.. Protein- and material-based approaches to modulate barrier function.

A) Diminished barrier function can be achieved through the direct modulation of TJ proteins by protein claudin binders or by mechanical perturbation via smart, responsive materials. B) Indirect approaches to modulate barrier function activate signaling cascades that induce TJ remodeling by engaging with cell surface receptors, such as apical integrins, or by disrupting cell–cell adhesion.