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Coupling large molecules with targeting and delivery mechanisms to augment intracellular delivery

Most intracellular drug targets remain intractable due to cellular membranes effectively denying large foreign molecules entry into the cytoplasm. Over recent decades, several technologies that enable biologics to enter cells have emerged, but successes remain limited to date. Here, Sapreme Technologies’ Miriam Bujny, Chief Development Officer, and Guy Hermans, Chief Executive Officer, discuss novel bioconjugation methods to incorporate specific cell-targeting and membrane-penetrating mechanisms into therapeutics to effectively engage previously elusive intracellular targets.

Arange of approaches in modern biotechnology necessitate the safe and efficient intracellular delivery of therapeutic cargo such as proteins, oligonucleotides and small molecules to ensure therapeutic effect. However, tissues and their constituent cells have evolved mechanisms and physical barriers to prevent foreign molecules from entering the cellular cytoplasm, thus blocking the delivery of therapeutics. In the event that a large molecular drug succeeds in reaching the target cells and becomes internalised, for many protein and gene delivery applications such uptake is largely non-productive as a significant portion remains trapped in the cells’ internal membrane systems and ends up in degradative pathways. It is estimated that only a minor fraction of protein and less than one percent of a given oligonucleotide1 manages to escape and be released into the cytosol. Accordingly, only a very minor fraction of the administered drug is ultimately available to exert any pharmacologically relevant effect, often insufficient to afford a useful therapeutic window. To unlock the full potential of promising novel therapeutic modalities for intracellular targets, the development of effective, clinically relevant cell-targeting and cytoplasmic delivery mechanisms is a key challenge.

Large molecule internalisation and endosomal escape inspired by nature

In principle, two major pathways exist for large molecules to reach the inside of a cell: by direct penetration of the plasma membrane or by hijacking cellular internalisation pathways. The most common internalisation pathway is uptake by receptor-mediated endocytosis. Direct traversal of the plasma membrane is virtually impossible for large molecules given their size, charge, and the biophysical properties of the cell membrane. Mechanisms to escape the confines of the endosomal pathways following internalisation, however, have evolved in several organisms – for example, in viruses, bacteria and in the form of plant secondary metabolites – equipping these organisms with means to gain complete entry into the cytosol of a host or predator cell, or insertion of specific payloads into the same. In the context of drug development, targeting endosomal uptake and engineering endosomal escape approaches based on these mechanisms has become an attractive concept for intracellular delivery of large drug molecules.