Industry Challenges

GSK

Advancing Lipopolysaccharide (LPS) Testing in Oligonucleotide Therapeutics: Investigating Test Interactions and Reducing Variability in Limulus Amebocyte Lysate (LAL) Assays

CAMS themes:

Complex mixtures
Separations and detection

Project Summary

To investigate the interactions occurring within the Limulus Amebocyte Lysate (LAL) assay when used to assess oligonucleotide therapeutics and develop strategies to reduce variability and improve assay robustness.

This could cover elements of the following areas:

1. Characterise Potential Interactions in the Assay System;

2. Understand Variability in LAL Assays;

3. Develop Mitigation Strategies for Reducing Variability;

4. Propose Future-Ready Testing Methodologies.

Impact & Benefits

Lipopolysaccharide contamination poses significant challenges to the development and manufacture of oligonucleotide therapeutics. Advances in testing methodologies, removal strategies, and regulatory compliance are driving progress in ensuring product safety and efficacy. Continued research will play a pivotal role in overcoming existing challenges and setting new standards for LPS testing in oligonucleotides. Aspects of this will entail understanding the potential of recombinant assays (rFC and rCR), the role of next generation endotoxin testing platforms and develop best practices for LPS testing in oligonucleotide therapeutics, supporting harmonisation with regulatory guidelines.

Project Aims

1. Characterise Potential Interactions in the Assay System:
- Investigate interactions between LPS controls, LAL reagents (natural and recombinant), and oligonucleotide materials.
- Assess how oligonucleotide physicochemical properties (e.g., charge, structure, and size) influence LAL assay outcomes.
2. Understand Variability in LAL Assays:
- Quantify the variability in endotoxin recovery when oligonucleotides are tested across different LAL assay formats (e.g., turbidimetric, chromogenic, recombinant).
- Identify assay system components or parameters contributing to inconsistent performance.
3. Develop Mitigation Strategies for Reducing Variability:
- Optimise sample preparation techniques to minimise matrix interference in oligonucleotide testing.
- Explore tailored reagent formulations or assay adaptations to enhance compatibility with oligonucleotide materials.
4. Propose Future-Ready Testing Methodologies:
- Evaluate the potential of recombinant assays (both rFC and rCR) and next-generation endotoxin testing platforms.
- Develop best practices for LPS testing in oligonucleotide therapeutics, supporting harmonisation with regulatory guidelines.

Potential project benefits

This research aims to systematically investigate the underlying interactions within LPS testing methodologies and develop novel approaches to reduce assay variability. By addressing key gaps in knowledge, this work will contribute to the establishment of robust, reliable and reproducible endotoxin testing for oligonucleotide therapeutics.

Additional Comments

Technical success would be the substantial increase of knowledge around LPS-oligonucleotide interactions within the endotoxin assay and the processes that could be used to overcome these where possible.