Delivery of therapeutic RNA-cleaving oligodeoxyribonucleotides (deoxyribozymes): from cell culture studies to clinical trials

ABSTRACT

Introduction. Development of efficient in vivo delivery systems remains a major challenge en route to clinical application of antisense technology, including RNA-cleaving molecules such as deoxyribozymes (DNAzymes). The mechanisms of oligonucleotide uptake and trafficking are clearly dependent on cell type and the type of oligonucleotide analogue. It appears likely that each particular disease target would pose its own specific requirements for a delivery method.

Areas covered. In this review we will discuss the available options for DNAzyme delivery in vitro and in vivo, outline various exogenous and endogenous strategies that have been, or are still being, developed and ascertain their applicability with emphasis on those methods that are currently being used in clinical trials.

Expert opinion. The available information suggests that a practical system for in vivo delivery has to be biodegradable, as to minimize concerns over long-term toxicity, it should not accumulate in the organism. Extracellular vesicles may offer the most organic way for drug delivery especially as they can be fused with artificial liposomes to produce hybrid nanoparticles. Chemical modification of DNAzymes holds great potential to apply oligonucleotide analogs that would not only be resistant to nuclease digestion, but also able to penetrate cells without external delivery agents.

KEYWORDS:

• Antisense oligonucleotide
• cell uptake
• extracellular and intracellular barriers
• downregulation of gene expression
• exogenous and endogenous delivery systems

Article highlights

• DNAzymes are artificial DNA molecules that could be designed for cleaving natural RNA targets to achieve therapeutic effect from downregulation of specific gene expression.

• The main obstacle towards clinical applications of DNAzymes seems to be insufficient cellular uptake, which necessitates the need for delivery systems to ensure their successful use for gene silencing either in cell culture or in vivo.

• DNAzymes may be delivered into cells either endogenously or exogenously. Endogenous systems were designed to produce DNAzyme molecules inside cells using reverse transcription. More common is exogenous delivery, for which a number of delivery systems have been developed such as liposomes or oil-based emulsions, inorganic nanoparticles, polymers or dendrimers, or systems based on conjugating DNAzymes to transporter molecules such as peptides or structural motifs.

• Many of the exogenous delivery systems have been used to promote cell uptake of DNAzymes in cell culture, much less have been tried in vivo and very few have been employed so far for clinical trials. This emphasizes the need for simpler, less expensive and non-toxic approaches to DNAzyme delivery, which would be sufficiently effective to be applicable for clinical trials.

• Chemical modification could be very promising for designing DNAzymes, which are stable in biological media, have better catalytic properties and, potentially, enhanced cell uptake in the absence of external delivery agents.

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Acknowledgment

The authors thank Professor Richard Cosstick for critical reading of the manuscript.

Declaration of interest

The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Additional information

Funding

DA Stetsenko acknowledges financial support from RSF grant 14-44-00068, AA Fokina and BP Chelobanov acknowledge financial support from RSF grant 15-15-00121. Funding was also received from the Institute of Chemical Biology and Fundamental Medicine SB RAS and Kindai University.