New open-source tool automates RNA analysis, likely to encourage research and drug development

Scripps Research scientists have introduced a new software tool for examining RNA (ribonucleic acid) molecules, which play a variety of important roles in living things.

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The open-source program “Pytheas,” reported in Nature Communications on May 3^rd, 2022, makes it easier to characterize and analyze RNAs in basic research and therapeutic development.

The software is meant to examine RNA data generated by a technique known as mass spectrometry. The term “mass spec” is commonly used to describe RNA molecules that are not just chains of standard RNA nucleotides but have been altered in some way. Pytheas can be used to quickly detect and measure altered RNA molecules, such as those seen in the current Pfizer and Moderna COVID-19 mRNA vaccines, according to the researchers.

The analysis of RNA data from mass spectrometry has been a relatively laborious process, lacking the tools found in other areas of biological research, and so our aim with Pytheas is to bring the field into the 21st century.”

James Williamson PhD, Study Senior Author and Professor, Integrative Structural and Computational Biology, Scripps Research Institute

Williamson was also the vice president of Research and Academic Affairs at Scripps Research.

Luigi D’Ascenzo, PhD, and Anna Popova, PhD, a postdoctoral research associate and a staff researcher in the Williamson lab at the time of the study, were the first authors.

RNA molecules in cells are substantially engaged in the process of translating genes into proteins, as well as fine-tuning gene activity since they are chemically identical to DNA. RNA-based treatments, such as the Pfizer and Moderna vaccines, are also seen as a good potential class of medicines, capable of reaching biological targets more powerfully and precisely than standard small-molecule pharmaceuticals in theory.

Mass spectrometry is a typical test for analyzing RNA molecules with chemical alterations, as it can be used to recognize RNAs and their modifications based on their masses. Natural RNAs are frequently transformed, affecting their roles, but RNAs used in vaccines and RNA-based medications are virtually always purposefully modified to improve their activity and minimize negative effects.

In comparison to equivalent methods in the field of protein analysis, for example, methods for transforming raw mass spectrometry data on modified RNAs have been extremely slow and manual—thus, quite labor-intensive.

To dramatically increase the automation of this procedure, Williamson and his colleagues created Pytheas, which is centered on the Python programming language. The software takes mass spec data from an RNA sample as input and produces projected RNA sequences and chemical changes in a way that enables quantifying different RNAs in a sample easily.

Using mass spec data for major bacterial and yeast RNAs, as well as SARS-CoV-2 spike protein messenger-RNAs like those utilized in the Pfizer and Moderna COVID-19 vaccines, the scientists confirmed Pytheas’ speed, accuracy, and adaptability.

We’re hoping that companies involved in manufacturing RNA vaccines and other RNA therapeutics will find Pytheas useful, for example in monitoring the quality of their products.”

James Williamson PhD, Study Senior Author and Professor, Integrative Structural and Computational Biology, Scripps Research Institute

The researchers in their investigations of natural RNAs are presently using Pytheas, and the program is still being improved.