Scalable multimodal weak anion exchange chromatographic purification for stable mRNA drug substance

Abstract

Messenger RNA (mRNA) has emerged as a modality with immense therapeutic potential. Recent innovations in production process of mRNA call for procedures to isolate pure mRNA drug substance (DS) with high yield, high capacity, scalability, and compatibility with GMP production systems. Novel RNA modalities, such as circular RNA (circRNA), have further driven the need for non‐affinity capture possibilities which are already widely used in the biopharmaceutical industry, for example, in monoclonal antibody processing. The principle that multimodal ion exchange/hydrogen bonding chromatography can be used to separate mRNA from in vitro transcription components has recently been demonstrated. Here, we apply and refine this approach to be suitable for scalable purification of multiple mRNA constructs with sufficient yields, purity, and stability, for use in mRNA production process. Binding capacity of the PrimaS‐modified monolithic chromatographic column for mRNA enabled up to 7 mg/mL product isolation in a single chromatographic run, with 98% recovery and room temperature stability of the eGFP mRNA demonstrated for up to 28 days. This approach is independent of construct size or the presence of polyadenylic acid tail and is applicable for capture of a wide variety of RNAs, including mRNA, self‐amplifying RNA, circRNA, and with optimization also smaller RNAs such as transfer RNA and others.

Editorial

“…A part of the published manuscripts was also presented at the last Monolith Summer Symposium that was held last year in Portorož, Slovenia. As a logical consequence of the spirit of this time, the topics of most papers that are summarized here are dealing with purification of nanoparticles like viruses [1][2][3], plasmid DNA isoforms [4,5] and RNA drug substances [6]. As is well known, monolithic stationary phases (including membranes) were firstly developed for fast separation of proteins [7], but only the last manuscript in this issue reports the development of a new monolithic stationary phase for protein separation [8].…”

“…The next three papers are dedicated to fractionation of materials containing different DNA and RNA forms [4][5][6]. Kralj et al reported the optimization of production of large plasmid DNA isoforms with sizes over 10 kbp using anion exchange monoliths.…”

“…Megušar et al [6] reported recent innovations in production processing of mRNA in the direction of high yield, high capacity and scalability, which also enables production processes that are compatible with GMP production systems.…”