Seminars & Webinars
2023
Event: mRNA Applications in Discovery & Development
Date: June 20, 2023
Presenter: Rok Sekirnik, Sartorius BIA Separations
This talk presents case studies of leveraging the power of at-line HPLC monitoring for process development to increase purity and yield of mRNA production. It describes approaches to optimize plasmid purification and linearization, IVT reaction and mRNA purification, including purification methods to achieve more than 95% supercoiled plasmid purity, optimize linearization reaction, and increase of IVT reaction yield from 3-5 mg/mL to 10-12 mg/mL. Further, use of at-line analytics to monitor automated, continuous production in single use bioreactors to produce multi-gram amounts of mRNA are demonstrated. Oligo dT-based affinity purification is described with approaches to optimize dynamic binding capacity to reach up to 6 g/L. Multimodal chromatography and reverse-phase chromatography cases are discussed for capture of non-polyadenylated RNA, and removal of dsRNA.
Event: Cell and Gene Therapy Insights webinar
Date: March 1, 2023
Presenter: Blaž Bakalar, Product Manager PATfix platform
mRNA-based therapeutics represent a promising new modality for therapies and vaccines. However, industry-wide process understanding of the CQA and CPP interplay is still developing. As such, the lack of immediate feedback on the process state during process optimization, sometimes delayed by weeks and limited by throughput, represents a big bottleneck in mRNA commercialization. The PATfix mRNA analytical platform enables reliable at-line insight during process development and production of mRNA-based therapeutics. With an ability to resolve key IVT reaction components (NTPs, capping reagent, template and mRNA), both USP and DSP process insight can be gained, with a single analytical tool. The PATfix mRNA platform has been upgraded to include LNP encapsulation analytics, enabling at-line monitoring of a crucial process step on the path from mRNA drug substance to LNP drug product.
Presenter: Blaž Bakalar, Product Manager PATfix platform
Date: February 15, 2023
Event: 2nd mRNA Analytical Development Summit 2023
Abstract:
- PATfix mRNA analytical platform is a perfect tool for process developers looking to optimize their mRNA production and purification
- Three orthogonal analytical methods based on mix-mode, affinity and reverse phase allow for precise quantification and mRNA sample characterization
- The newly developed method for characterization of encapsulated mRNA molecules covers the last crucial part in mRNA therapeutic production
Attachments
2022
Speaker: Rok Sekirnik, Head Process Development for mRNA and pDNA
The recently demonstrated efficacy of mRNA-based Covid-19 vaccines has shown the promise of this therapeutic format, but also highlighted the need for higher efficiency of mRNA production to meet enormous needs for global vaccine supply. The production process typically involves 10-15 steps including plasmid production, plasmid linearization, an in vitro transcription (IVT) reaction, mRNA purification, and lipid nanoparticle (LNP) production. This webinar will discuss the versatility of chromatography as applied to mRNA production, starting with the purification of the key raw material (plasmid DNA) to final polishing of mRNA drug substance.
Speaker: Rok Sekirnik, Head Process Development for mRNA and pDNA
The recently demonstrated efficacy of mRNA-based Covid-19 vaccines has shown the promise of this therapeutic format, but also highlighted the need for higher efficiency of mRNA production to meet enormous needs for global vaccine supply. The production process typically involves 10-15 steps including plasmid production, plasmid linearization, an in vitro transcription (IVT) reaction, mRNA purification, and lipid nanoparticle (LNP) production. This webinar will discuss the versatility of chromatography as applied to mRNA production, starting with the purification of the key raw material (plasmid DNA) to final polishing of mRNA drug substance.
Event: Cell and Gene Therapy Insights webinar
Date: June 7, 2022
Presenter: Nejc Pavlin
Pre-developed and validated PATfix analytical methods using different column chemistries enable:
- Effective control over linear plasmid upstream and downstream,
- IVT reaction optimization for mRNA synthesis,
- Control of yield, purity and downstream process of mRNA,
- mRNA formulation and stability analysis,
- Scale-up optimization.
In this webinar, will discuss a robust analytical tool PATfix™ platform, and its specific methods for pDNA and mRNA analytics that enable efficient monitoring of key upstream and downstream process steps will be showcased.
Event: Cell and Gene Therapy Insights webinar
Date: April 12, 2022
Presenter: Rok Sekirnik, Head of Process Development, mRNA/pDNA
Key learning points:
- Benefits of implementing HPLC-based analytics in mRNA production workflow
- Principles of monolith chromatography and its use for mRNA analytics and purification
- Principles of IVT reaction and factors affecting rate and yield
- Principles of conversion from batch to fed-batch process
Event: Informa 1-day digital event, highlighting mRNA Applications and Therapeutics
Date: March 23, 2022
Presenter: Tomas Kostelec, Technology & Business Development Manager
Key learning points:
- Ensuring consistent and robust IVT production of mRNA
- Achieving high yield and high purity in mRNA purification processes
- Chromatography solutions for in process analytics and purification
Event: BPI Ask the Expert
Date: March 3, 2022
Speaker: Rok Sekirnik, Head Process Development for mRNA and pDNA
In this webinar, we will discuss an HPLC-based analytical method using a multimodal ligand based on anion exchange/hydrogen-bonding ligand (PrimaS) to monitor the IVT reaction, which allows for simultaneous quantification of NTPs, capping reagent, plasmid, and mRNA within 3.5 min.
Event: mRNA-Based Therapeutics Summit Europe
Date: January 2022
Presenter: Rok Sekirnik, Head of Process development for mRNA and pDNA
Abstract:
In vitro-transcribed messenger RNA-based therapeutics represent a relatively novel and highly efficient class of drugs. Current clinical efforts encompassing mRNA – based drugs are directed to three categories: mRNA vaccine, mRNA therapy and protein replacement therapy. As mRNA vaccine became the front runner to fight the COVID-19 pandemic, challenges surrounding their process development and manufacturing became readily apparent:
- mRNA manufacturing is not yet a platform approach. Standardized products along the processes are needed
- mRNA is a relatively large molecule, which leads more challenges for downstream purification.
- mRNA is intrinsically unstable and prone to degradation.
- High raw material cost is limiting mRNA commercialization.
This presentation provides a detailed overview of current mRNA manufacturing approaches, highlights challenges and recent successes, and offers perspectives on the future of mRNA manufacturing.
2021
Published: Cell & Gene Therapy Insights 2021; 7(11), 1409
Date: December 2, 2021
Presenter: Rok Sekirnik, Head Process development for mRNA and pDNA
Watch the video or read the poster to learn:
- Non-affinity chromatography with PrimaS provides purification for IVT processes with co-transcriptional polyadenylation
- The Sartorius mRNA purification toolbox provides a scalable chromatographic solution for commercial manufacturing of mRNA therapeutics
- CIMac analytical HPLC columns can resolve individual IVT reaction components in minutes, resolving the analytical bottleneck in optimization of IVT reaction conditions
- Rapid HPLC analytics can improve in-process and quality control of the mRNA production process
Attachments
Presenter: Aleš Štrancar
Event: BPI and Cell & Gene Therapy Bioprocessing & Commercialization 2021
Date: September 30, 2021
Abstract:
Global demand for pDNA production is at an all time high, due to increased need from Gene Therapy ramp-up. pDNA, as an enabling product, is critical in production of mRNA, AAV and other therapeutic vectors. Increasing yield and purity in the production of pDNA is a vital step in meeting such demand. Supporting reliable in-process control during pDNA purification, PATfix pDNA analytical platform is enabling rapid process development and optimization while providing a reliable analytical platform for production runs.
Attachments
Presenter: Rok Sekirnik
Event: mRNA Therapeutics Summit
Date: July 14, 2021
Abstract:
The recently demonstrated efficacy of mRNA-based Covid-19 vaccines has shown the promise of this therapeutic format, but also highlighted the need for higher efficiency of mRNA production to meet enormous needs for global vaccine supply. Typical mRNA production process involves three key steps: 1) plasmid DNA (pDNA) production in supercoiled (sc) isoform, linearization and purification, 2) in-vitro transcription (IVT) reaction and 3) mRNA purification. Here we present a chromatographic toolbox for integrated mRNA production from pDNA to mRNA purification, including in-process analytics.
Attachments
Presenter: Aleš Štrancar
Event: TIDES mRNA Digital Week
Date: June 28, 2021
Abstract:
A new purification approach starting from E.Coli all through to mRNA production is presented here. This new approach integrates a pDNA linearisation step before polishing (removal of linear and open circular isoforms) of plasmid DNA. The polishing step, placed after enzymatic linearisation, purifies linear pDNA from enzyme and other unwanted process impurities. The linearised plasmid DNA is then used in IVT for production of mRNA. Fast in-process analytics allows for transcription number well above 100 in reproducible manner.
This approach further introduces mRNA purification tools for improved contaminant removal. The altered sequence of purification and linearisation reduces the overall number of purification steps required, improves recoveries, while the complete process results in extra low protein impurity and very efficient dsRNA removal.
The participants would get insight into the mRNA production steps, their optimisation and tools to allow for robust process. The analytical tools to allow for safe product will be highlighted, too.
Attachments
Presenter: Aleš Štrancar
Event: Genetic Vaccine Development for Infectious Diseases Summit
Date: May 20, 2021
Abstract:
- Developments in gene therapy and threat of pandemics requests novel technologies to allow for efficient manufacturing of very large biomolecules and nanoparticles within short time.
- Convective Interaction Media with open channel structures resulting in no shear forces allows for fast and very efficient manufacturing of mRNA, pDNA, Adeno and other VLP based Covid vaccines.
- Ultra-sensitive DNA assay and fast in-process control using PATfix systems are introduced for production of safer Covid vaccines.
Attachments
Event: BioProcess International COVID-19 Therapeutic Development & Production
Date: April 27, 2021
Developments in gene therapy and the threat of pandemics requires novel technologies to allow for efficient manufacturing of exceptionally large biomolecules and nanoparticles within a short time. Novel vaccine platforms such as mRNA enable the industry to meet its highest goals: decreasing footprint while increasing efficiency and throughput. However, to respond with agility and prepare for future outbreaks, the mRNA platform must be further optimized.
In vitro transcription (IVT), the enzymatic process used for mRNA vaccine production, is different from biological fermentation processes as it requires linearized plasmid DNA. The linear isoform is produced with restriction enzymes from open circular and supercoiled pDNA. Employing a traditional pDNA manufacturing process, which removes multimers as well as linear and open-circular isoforms, reduces production yield.
However, when pDNA and mRNA are treated as a single production process, the purification steps are optimized, yields improved, and overall production costs reduced. This presentation will describe how Convective Interaction Media monoliths can be used to efficiently purify a board specter of mRNA and cover all production scenarios.
This presentation illustrates fast in-process control methods using the PATfix system, which can ensure the rapid production of the safest possible Covid-19 vaccines while reducing manufacturing costs.
Attachments
Event: Nucleic Acids- cGMP Production of mRNA & pDNA (virtual edition)
Date: February 1, 2021
Our new Head of Process Development for mRNA and pDNA purification, Rok Sekirnik, presented our Chromatographic tools for high-yielding mRNA production process as workshop.
Click here to listen to the workshop.
Event: Nucleic Acids- cGMP Production of mRNA & pDNA (virtual edition)
Date: February 3, 2021
Our CEO, Ales Strancar presented our Integrated Production Process of mRNA from E. coli to Highly Purified mRNA
Attachments
2020
Presenter: Aleš Štrancar
Date: od demand, October 19-22, 2020
Cell & Gene Therapy Bioprocessing & Commercialization (digital event)
Established processes for pDNA manufacturing include purification steps designed to enrich the supercoiled (sc) isoform. Homogeneous supercoiled isoform improves cell transfection efficiency in fermentation processes and is favoured in cell culture production systems. In vitro transcription (IVT), the enzymatic process used for production of mRNA vaccines differentiates itself from biological fermentation processes by the need for linearised plasmid DNA. The linear isoform is produced with restriction enzymes from open circular and supercoiled plasmid DNA. Employing a traditional pDNA manufacturing process, which removes linear and open-circular isoforms, will therefore reduce the production yield. When plasmid DNA and mRNA are treated as a single production process, the purification steps can be optimised, yields improved, and the overall production cost reduced.
A new purification approach starting from E.Coli all through to mRNA production is presented here. This new approach integrates a pDNA linearisation step before polishing of plasmid DNA. The polishing step, placed after enzymatic linearisation, purifies linear pDNA from enzyme and other unwanted process impurities. The linearised plasmid DNA is then used in IVT for production of mRNA. This approach further introduces mRNA purification tools for improved contaminant removal. The altered sequence of purification and linearisation reduces the overall number of purification steps required, improves recoveries, while the complete process results in extra low protein impurity and very efficient dsRNA removal.