Intracellular Trafficking and Endosomal Release of Oligonucleotides: What We Know and What We Don’t

Abstract

Understanding the cellular uptake and intracellular trafficking of oligonucleotides provides an important basic underpinning for the developing field of oligonucleotide-based therapeutics. Whether delivered as "free" oligonucleotides, as ligand-oligonucleotide conjugates, or in association with various nanocarriers, all forms of oligonucleotide enter cells by endocytosis and are initially ensconced within membrane-limited vesicles. Accordingly, the locus and extent of release to the cytosol and nucleus are key determinants of the pharmacological actions of oligonucleotides. A number of recent studies have explored the intracellular trafficking of various forms of oligonucleotides and their release from endomembrane compartments. These studies reveal a surprising convergence on an early-intermediate compartment in the trafficking pathway as the key locus of release for oligonucleotides administered in "free" form as well as those delivered with lipid complexes. Thus, oligonucleotide release from multivesicular bodies or from late endosomes seems to be the crucial endogenous process for attaining pharmacological effects. This intrinsic process of oligonucleotide release may be amplified by delivery agents such as lipid complexes or small molecule enhancers.

Observing an Antisense Drug Complex in Intact Human Cells by in‐Cell NMR Spectroscopy

“…A biotin‐tagged ASO was co‐electroporated (2 % of 12.5 μ m ) with the unmodified ASO into human embryonic kidney (HEK 293T) cells and imaged with the aid of fluorescently labelled streptavidin (Figure A, B). The biotinylated ASO concentrates in granular structures inside the cell (for transfection rates see Section 1.9 in the Supporting Information), likely reflecting accumulation of the drug in the endocytic pathway or in cytoplasmic structures, as supported by the literature …”

Erufosine (ErPC3) Cationic Prodrugs as Dual Gene Delivery Reagents for Combined Antitumor Therapy

“…Besides, from the sorting endosomes, most amphiphilic molecules enter the endocytic recycling pathway . Lipid recycling at the plasma membrane is a highly dynamic and effective process, and antiproliferative APL produced in situ from the nucleic acid carrier is prone to trafficking at the plasma membrane where it can operate intrinsic apoptotic activity . Starting from erufosine, one of the most promising APLs to date, original lipid constructs were thus imagined for simultaneously implementing gene therapy and chemotherapy.…”

“…This processt hus triggersa nd facilitates nucleic acid decondensation, which is requiredf or nucleic acid processing by the cell translation machinery.B esides, from the sorting endosomes, mosta mphiphilic molecules enter the endocytic recycling pathway. [18] Lipid recycling at the plasma membrane is ahighly dynamic and effective process, [19] and antiproliferative APL produced in situ from the nucleic acid carrier is pronet ot rafficking at the plasma membrane where it can operate intrinsic apoptotic activity. [20] Starting from erufosine,o ne of the most promising APLs to date, originall ipid constructs were thusi magined for simultaneously implementing gene therapy and chemotherapy.Aseries of 16 biolabile cationic lipids have been designed which can regenerate erufosine in situ under ac hemical or enzyme stimulus.…”

Using macropinocytosis for intracellular delivery of therapeutic nucleic acids to tumour cells

“…In either scenario, this is likely to be of benefit for intracellular delivery. Recent work has shown that the majority of endosomal escape occurs in late endosomes/ multivesicular bodies [107], suggesting that trapping at this stage of the pathway or increasing retention time before lysosomal degradation would be beneficial. In addition to avoiding lysosomal degradation, recycling is another limiting factor of nucleic acid delivery.…”