Enabling rapid liquid and freeze-dried formulation design for the manufacture and delivery of novel biopharmaceuticals

Biopharmaceuticals have been approved to treat diseases including cancers, rheumatoid arthritis, multiple sclerosis, diabetes, leukemia and neutropenia. The next-generation of protein-based therapies, or biopharmaceuticals, are of increasingly complex engineered forms, with unpredictable solution properties. Proteins are formulated at high concentrations for clinical use, leading often to undesirable aggregate formation, high viscosity, opalescence, or phase separation, rendering them unsafe, or difficult to inject or manufacture. This is a major challenge to the biopharmaceuticals industry as approximately 50% of proteins in clinical trials have been freeze-dried as they were not readily liquid-formulated on timescales of months required during development. Formulation is an empirical process using combinatorial screens that aim to optimise stability, potency and ease of delivery to patients. Engagement with 36 industry leaders at a UCL EPSRC Centre for Innovative Manufacturing workshop identified the most significant protein formulation challenges such as the prediction of shelf stability over a two year period, at a time in development when not much material is available. Current surrogate techniques that accelerate protein degradation and minimize sample consumption provide poor indicators of 2-year shelf-life. Industry would benefit significantly from i) rapid analyses that more accurately determine long-term shelf-life, ii) low concentration analyses that indicate high-concentration solution behaviour, and iii) a better ability to use calculated protein and excipient properties to predict those formulations that are most likely to meet the required attributes.

State-of-the-art automated microplate and microfluidic analytics, purchased or established recently via EPSRC and BBSRC/BRIC awards at UCL and UoM provide a timely platform for generating large experimental datasets of aggregation kinetics spanning many different timescales, conformational and colloidal stabilities, rheological properties, phase-transition and glass transition temperatures, for liquid and freeze-dried formulations. A recent EPSRC funded £500k pilot-scale freeze-drying facility at UCL (EP/M028100/1), combined with DoE and 3D process simulations, will generate freeze-drying process models that elucidate the mechanisms linking critical process parameters to critical quality attributes for new formulations. Novel dipeptides emerging from recent UoM work will significantly expand the range of industry-accepted formulation excipients available. Novel microfluidic analytics will be tailored for formulation needs, bringing earlier, more sensitive, and lower-volume assessments of formulated protein heterogeneity and storage kinetics, ultimately in a high-throughput format using sealed microwells.

All data will populate a web-access database at UoM to provide modeling groups access to a much-needed experimental dataset. Informatics techniques initiated at UoM in a BioProNet PoC award will enable new proteins to be compared (via properties calculated from sequence and structure) to those in the database, and use their experimentally determined formulation behaviours in a predictive manner. Correlations between calculated protein properties and critical formulation attributes will identify the molecular basis of excipient behaviour.

Overall, this will benefit the biopharmaceutical formulation community with an ability to: a) identify better excipient combinations for input into formulation screens; b) predict those protein candidates most readily formulatable with current excipients and solution conditions; c) inform the rational design of novel peptide-based excipients through defined chemical modifications, d) predict long-term storage stability and concentrated solution behaviour from accessible experiments using minimal sample.

"WHO WILL BENEFIT FROM THE RESEARCH?"

UK-based companies within the BioProNet/BRIC community will benefit from research which allows them to more effectively predict lead protein therapy candidates and formulations that are likely to be viscous, or lead to aggregation during storage, freeze-drying, bioprocess development and manufacturing, using high-throughput experimental and computational tools for a "screen early - fail early" approach. Industry will also have access to predictive tools and archived data that will inform the more effective design of their own product formulations and formulation screens. This will in turn decrease the time, cost and risk of product development.

Potential patients will benefit because the research will significantly aid reduction in formulation development times of protein therapies, which is particularly crucial for those addressing previously unmet clinical needs. Benefits to the NHS relate to the possibility of constraining costs that accumulate in industry due to late-stage failures of drug candidates. Proteins are innately complex and labile so that product development times and hence costs tend to be high. The capacity to treat conditions such as rheumatoid arthritis much more effectively in ageing populations is vital but it still poses a problem with respect to stretched NHS budgets.

"HOW WILL THEY BENEFIT FROM THE RESEARCH?"
The research will fundamentally characterise the interactions between formulation excipients and proteins, as measured by their impact on aggregation kinetics, conformational and colloidal stability, and rheological properties, in a wide range of buffer conditions. This will reveal commonalities between several protein types, as well as their idiosyncrasies. This improved understanding will better direct protein formulation, engineering and candidate selection through the generation of predictive tools that identify formulation excipients most likely to stabilise each new therapeutic protein. The research will also demonstrate advanced microfluidic analytical techniques to rapidly identify the likelihood of a protein formulation to aggregate, and to detect aggregates with very high sensitivity. Together, the rapid analytical tools and the greater understanding of the structural mechanisms that lead to protein stabilisation by formulation excipients will enable the UK biotechnology industry to design more efficient formulation screening designs, novel excipients, and robust protein scaffolds that minimise aggregation, and high viscosity, and prolong product shelf-life. This will continue to improve as academic beneficiaries in the UK will also be able to refine the predictive models for formulation impact on aggregation and rheological properties, by adding their own data to the database, and though interactive use of the new algorithms.

The UK economy will benefit because academic research will complement the country's strength in bioscience discovery and development. Collaboration between formulation engineers, computational scientists and protein biophysicists on industrially relevant therapeutic proteins will ensure effective knowledge and skills transfer between the science and engineering base and UK industry. This will expand their position in the global healthcare market and attract further R&D investment from global business which recognises the UK as a good place to conduct these activities. Such retention of expertise, know-how and intellectual property will aid the capacity to remain internationally competitive.