Cellular Uptake and Intracellular Trafficking of Oligonucleotides: Implications for Oligonucleotide Pharmacology

Juliano, Rudolph L.; Xin, Ming; Carver, Kyle; Laing, Brian

doi:10.1089/nat.2013.0463

Cited by 105 publications

(89 citation statements)

References 152 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The mechanism by which this modification increases cellular uptake is still under study, but is likely related to enhanced binding to proteins such as albumin, which act as a cloak and escort the oligo through various endocytotic pathways. 29, 31 This feature is potentially attractive as a method of delivering the probe, which lacks the 5′ CPP found in earlier TIVA constructs. We investigated this effect by incubating human fibroblast cells with 0.5 μM PS-22/9/9 or 22/9/9, without use of CPP or other delivery agent.…”

Section: Resultsmentioning

confidence: 99%

Oligonucleotide modifications enhance probe stability for single cell transcriptomein vivoanalysis (TIVA)

2017

View full text Add to dashboard Cite

Single cell transcriptomics provides a powerful discovery tool for identifying new cell types and functions as well as a means to probe molecular features of the etiology and treatment of human diseases, including cancer. However, such analyses are limited by the difficulty of isolating mRNA from single cells within biological samples. We recently introduced a photochemical method for isolating mRNA from single living cells, Transcriptome In Vivo Analysis (TIVA). The TIVA probe is a “caged” polyU:polyA oligonucleotide hairpin designed to enter live tissue, where site-specific activation with 405 nm laser reveals the polyU-biotin strand to bind mRNA in a target cell, enabling subsequent mRNA isolation and sequencing. The TIVA method is well suited for analysis of living cells in resected tissue, but has not yet been applied to living cells in whole organisms. Adapting TIVA to this more challenging environment requires a probe with higher thermal stability, more robust caging, and greater nuclease resistance. In this paper we present modifications to the original TIVA probe with multiple aspects of enhanced stability. These newer probes utilize an extended 22mer polyU capture strand with two 9mer polyA blocking strands (“22/9/9”) for higher thermal stability pre-photolysis and improved mRNA capture affinity post-photolysis. The “22/9/9 GC” probe features a terminal GC pair to reduce pre-photolysis interactions with mRNA by more than half. The “PS-22/9/9” probe features a phosphorothioated backbone, which extends serum stability from <1 h to at least 48 h, and also mediates uptake into cultured human fibroblasts.

show abstract

Section: Resultsmentioning

confidence: 99%

Oligonucleotide modifications enhance probe stability for single cell transcriptomein vivoanalysis (TIVA)

2017

View full text Add to dashboard Cite

show abstract

“…This improved uptake, in addition to the improved nuclease resistance, makes this modification an important element for the development of therapeutic oligonucleotides. The mechanism of uptake is not yet fully understood, although several proteins have been identified for their potential involvement [100]. The strong binding of phosphorothioate oligonucleotides to plasma proteins is considered a key characteristic.…”

Section: Uptakementioning

confidence: 99%

Phosphorothioates, Essential Components of Therapeutic Oligonucleotides

Eckstein

2014

Nucleic Acid Therapeutics

500

385

View full text Add to dashboard Cite

Phosphorothioates have found their usefulness in the general area of oligonucleotide therapeutic applications. Initially this modification was introduced into the antisense methodology because of the nuclease resistance of the phosphorothioate linkage in comparison with that of the phosphate linkage. However, as experimental data accumulated, it was detected that this chemical modification also facilitates cellular uptake and bioavailibity in vivo. Thus, today the majority of therapeutic oligonucleotides contain this modification. This review will discuss the historical development of this modification and present some of its chemical properties where they differ from those of the phosphate group. The antisense application will be discussed in the original context with cleavage of the target mRNA, but other target RNAs such as microRNAs and long noncoding RNAs will also be covered. It continues with applications where the target RNA should not be cleaved. A brief presentation of decoy oligonucleotides will be included, as well as some miscellaneous applications. Cellular uptake is a crucial step for oligonucleotides to reach their target and will be briefly reviewed. Lastly, a most surprising recent observation is the presence of phosphorothioate groups in bacterial DNA where functions still remain to be fully determined.

show abstract

“…Thus the agent Retro-1 was shown to strongly enhance the effects of antisense, splice-switching, and siRNA oligonucleotides in cell culture models [123, 145]. This compound acts by selective partial release of oligonucleotides from late endosomes, but not from lysosomes or other intracellular compartments.…”

Section: Endocytosis and Trafficking Of Oligonucleotidesmentioning

confidence: 99%

Cellular uptake and intracellular trafficking of oligonucleotides

Juliano

Carver

2015

Advanced Drug Delivery Reviews

Self Cite

116

View full text Add to dashboard Cite

Oligonucleotides manifest much promise as potential therapeutic agents. However, understanding of how oligonucleotides function within living organisms is still rather limited. A major concern in this regard is the mechanisms of cellular uptake and intracellular trafficking of both ‘free’ oligonucleotides and oligonucleotides associated with various polymeric or nanocarrier delivery systems. Here we review basic aspects of the mechanisms of endocytosis and intracellular trafficking and how insights from these processes can be used to understand oligonucleotide delivery. In particular we discuss opportunities for escape of oligonucleotides from endomembrane compartments and describe recent studies using small molecules to enhance oligonucleotide effects.

show abstract

Cellular Uptake and Intracellular Trafficking of Oligonucleotides: Implications for Oligonucleotide Pharmacology

Cited by 105 publications

References 152 publications

Oligonucleotide modifications enhance probe stability for single cell transcriptomein vivoanalysis (TIVA)

Oligonucleotide modifications enhance probe stability for single cell transcriptomein vivoanalysis (TIVA)

Phosphorothioates, Essential Components of Therapeutic Oligonucleotides

Cellular uptake and intracellular trafficking of oligonucleotides

Contact Info

Product

Resources

About