A New Tide for RNA Therapeutics

By Marwan Alsarraj

In the last several years, scientific breakthroughs have led to an influx in research and development around RNA-based therapeutics that offers new hope for applications ranging from vaccines to personalized cancer treatments. RNA therapeutics are more cost-effective and accessible to manufacture than traditional small-molecule drugs or protein-based drugs, giving them an attractive edge in scientific research and development. More than 50 RNA drugs have already reached clinical testing; roughly 28 are currently in clinical trials, and a handful have already received regulatory approval (Int. J. Mol. Sci. 2022; 23(5): 2736). Industry experts predict these successes are early signs of a therapeutic revolution.

Although RNA-based therapeutic development is still a relatively new commercial field, biopharma leaders pour resources into its growth and standardization. Doing so enables large-scale production, ensuring overall quality control from discovery to manufacturing.

But there are still obstacles preventing the widespread use of RNA biotherapeutics. In particular, these treatments require unique and comprehensive analyses during development and manufacturing.

Traditionally, scientists and researchers have relied on the standard method of analysis: quantitative polymerase chain reaction (qPCR). However, this method is no longer cutting-edge and needs to keep up with the rigorous demands of RNA therapeutics manufacturing. Droplet Digital PCR (ddPCR) is fueling the new revolution by helping ensure product safety and quality for RNA-based therapeutics to push them over the finish line. In this article, we will compare the two technologies and explore how absolute quantification delivers greater confidence from discovery to dosing, how unparalleled sensitivity enables more comprehensive quality control, and how simple workflows can increase performance.

From discovery to dosing, absolute quantification means more confidence

Absolute quantification makes all the difference in the development and validation of RNA therapeutics. To identify potential targets, researchers need to accurately quantify RNAs and gene expression in cells at baseline during the early stages of discovery. Additionally, later during dosing, clinical researchers can re-evaluate these levels after treatment to obtain a clear picture of a potential product’s therapeutic effects.

Given that qPCR requires the generation of a standard curve, this method can only ever achieve relative quantification. In contrast, ddPCR assays are endpoint assays that count RNA within a sample molecule by molecule, delivering absolute quantification. In addition, the technology that powers ddPCR instruments enables scientists to calculate the number of target molecules present via Poisson distribution analysis.

For therapeutics in later stages of development, absolute quantification of RNA in the drug enables developers to establish safe and effective patient dosing. With absolute quantification, developers can have tremendous confidence in analyzing biodistribution for a given drug.

Unparalleled sensitivity drives comprehensive quality control

The future of RNA therapeutic development depends on assuring quality control throughout every production stage. Small amounts of contamination or product inconsistency can compromise a therapeutic’s safety and efficacy, causing a domino effect on the success of the treatment and its impact on the patient. Extreme sensitivity in nucleic acid detection and quantification enables more comprehensive quality control, and only ddPCR technology can deliver at the level RNA therapeutics require.

Some levels of contamination or product inconsistency can be accidentally overlooked using traditional qPCR because small quantities of target RNA can be obscured by high-copy templates or even background noise. On the other hand, ddPCR data makes analysis simple and definitive. ddPCR technology increases the signal-to-noise ratio during massive sample fractionation, allowing for sensitive detection of rare target sequences. Additionally, highly sensitive ddPCR assays can detect small amounts of contamination, such as residual host cell DNA or adventitious agents like mycoplasma.

This degree of sensitivity enables precise detection and quantification of low-copy-number sequences within the sample, determination of poly(A) tail characteristics, and other critical RNA therapeutic development measurements. In a recent real-world example, a report issued by the European Medicines Agency (707383/2020) revealed that Pfizer/BioNTech used ddPCR assays for poly(A) tail analysis to manufacture their COVID-19 mRNA vaccine.

Reliable workflows are needed

The use of ddPCR technology in RNA therapeutic development enables increased performance in a simple workflow. The variability of results is significantly reduced with absolute quantification, which means excellent consistency from each development process run. Additionally, scientists can run up to 96 individual samples per plate, having no impact on reducing precision or accuracy. Each sample is partitioned into 20,000 individual PCR reactions, thousands of times more discrete measurements than the single measurement that the traditional qPCR produces. As a result, increased and more precise data is generated and gathered from a single ddPCR assay, enabling a simple and reliable workflow for RNA therapeutic development.

What’s next?

As the new tide for RNA therapeutics continues to increase, so will the use of the critical technology that will enable its success. The degree of sensitivity, accurate detection, and quantification of nucleic acid targets that ddPCR technology offers will fuel this revolution in RNA therapeutics, bringing more life-changing treatments to market.

Marwan Alsarraj is biopharma segment manager, Digital Biology Group, Bio-Rad Laboratories.

The post A New Tide for RNA Therapeutics appeared first on GEN – Genetic Engineering and Biotechnology News.

manufacturing
therapeutics
vaccines