The Case for Process Simplification in mRNA Manufacturing

By Anna Rose Welch, Editorial & Community Director, Advancing RNA

Last week, I published an article in a gameshow format.

Yes, you read that right. Having recently heard a presentation about mRNA COGS at the mRNA Therapeutics Summit by Kernal’s VP of CMC, Joe Parrella, I couldn’t help but share his insights on the current state of mRNA drug substance and drug product COGS through several game show segments loosely inspired by The Price Is Right.

Now, as much fun as that previous article was to write (and hopefully read), we return to our regularly scheduled program/format here in this follow-up article. Creating fictional gameshows about manufacturing practices is a complicated endeavor, but I’m also keenly aware not all of us are gameshow enthusiasts — especially when it comes to our mRNA process improvements.

Here, I wanted to share a few additional details from Parrella’s presentation about the efforts he and his team made to improve their overall mRNA-LNP COGS. Though he emphasized the amount of opportunity that exists to improve our utilization of costly raw materials/reagents, as he revealed, there are also opportunities for process simplification.

Less Is More: Simplifying the Process & Reducing COGS

As we learned — or perhaps reaffirmed — in the last article, we can thank mRNA drug substance production for having an outsized impact on our overall COGS. The drug substance alone accounts for 80 percent of COGS — with 90 percent of the drug substance COGS coming from raw materials (as opposed to manufacturing/QC and consumables).

For Kernal Bio, it was this knowledge that ultimately led them down the path of improving raw material usage — especially capping reagent usage — to improve overall mRNA-LNP production COGS. By monitoring reaction composition and incorporating bolus additions of materials into the reaction as needed at certain intervals, Parrella and his team reduced IVT COGS by 6x.  

However, process simplification was another important lever the team pulled — albeit not without encountering some challenges in the process.

In a typical mRNA DS manufacturing process, TFF occurs three times, with the first use being after IVT and preceding the first purification step. As Parrella went on to explain, he and his team investigated whether it was possible to remove this post-IVT TFF step, seeing as its predominant purpose in the process flow is to prepare the material for affinity chromatography. As he went on to qualify, “This TFF step wasn’t accomplishing any purification in and of itself.”

Though this removal perhaps made sense in theory, as the team discovered, it was a little easier said than done.

“Following our first attempt at removing the post-IVT TFF step, we realized that we adversely impacted the affinity chromatography,” Parrella admitted. “Bypassing the post-IVT TFF resulted in a pressure increase during the affinity chromatography step.”

As he went on to explain, cleaning was not a solution to this problem.

“Something was trapped in the column that we couldn’t remove using standard cleaning procedures, which led to even higher pressures during the next cycle. As a result, we lost almost half of our product.”

To mitigate this pressure increase, the team took some cues from the precipitation kinetics which suggested that inline blending of NaCl and mRNA using a two-pump chromatography system would be a feasible solution to eliminate the column pressure increase. In particular, the team found they had 5 minutes or so to perform this inline blending before precipitation would impact the chromatography column and diminish yield.  

“We were seeing issues in the absence of precipitation, whether it’s visible precipitation or a turbidity measurement using OD600. What we found was that .2μm filtration was a better surrogate for potential yield loss in the chromatography column. Even in short periods of time, you would see substantial yield loss across a .2 filter without any increase in OD600 or any increase in turbidity. Just after 5 minutes, we start seeing a meaningful loss in yield across the .2μm filter. We see that as somewhat of a surrogate chromatography column, but we also recognize that we’re going to have to .2 filter this between the IVT and the chromatography column just to protect it.”

Removing the first TFF step (in combination with semi-continuous IVT) knocked down COGS by another 5 percent, thanks to the reduction in clean room time (half a day), as well as the elimination of the technology and consumables needed for that TFF step, in particular.

But even more importantly, removing the post-IVT TFF did not cause any issues with quality. In fact, Parrella and company found that integrity of the molecule was slightly higher given that, without the first TFF step, there was less handling/processing of the mRNA.

Moving forward, Parrella and his team are also at work on another attempt to simplify the production process — namely by exploring the impacts of removing the second purification step and its associated TFF. However, this remains a work in progress and will hopefully be the subject of a future presentation by Parrella. Watch this space!