I think it’s fair to say that we are only as good as the sum of our parts. When running a professional race, an athlete will spend a considerable amount of time preparing for this great opportunity. They will have the training plan, stamina work and strategy all in place, but they may still lose the race if they then put on a bad pair of trainers. Imagine the frustration of spending all that time, effort, and money on one large event only to fail because you didn’t try on your footwear ahead of time?
Biotherapeutic production can suffer the same fate. Piles of time, effort and cost thrown into development of the drug product, optimising the cell line, purifying the target, and painstakingly monitoring to ensure the production is reliable and reproducible. Then your final product does not pass lot release because of impurities present - what did you miss?
Don’t let HCPs catch you out
Host cell proteins (HCPs) are proteins derived from the host cell being used to produce the recombinant biotherapeutic molecules, for example Chinese hamster ovary (CHO) cell lines. To prevent potential detrimental effects of HCPs, multiple purification steps are required to bring the HCP levels to an acceptable level, typically <100 ppm total HCPs, and <10 ppm per individually identified HCPs, although this can vary case-by-case.
Multicomponent enzyme-linked immunosorbent assay (ELISA) is currently the first line analytical strategy applied due to high throughput, sensitivity and selectivity of total host cell proteins, however ELISA does not inherently recognise all potential HCP species, and only provides quantitative data on total HCPs present. This adds significant risk to the drug product as the total HCP count may be within specification, but the method could still potentially miss HCPs present that could lead to issues with safety and/or efficacy.
Fail to prepare, prepare to fail
High-resolution accurate mass (HRAM) mass spectrometry-based approaches have shown significant advantages in this field, with the ability to identify and quantify individual and collective HCPs with confidence, and to detect low abundant species even if not clinically relevant. The bottleneck tends to be the sheer volume of data collected and the subsequent data analysis. But the industry is coming up with some great solutions to these problems; being able to identify more with simpler but efficient and robust sample preparation, and advanced software solutions to streamline the processing for more reliable and accurate answers faster.
A quick online search reveals this great example of how automated sample preparation with non-denaturing digestion can be simply implemented to remove the target monoclonal antibody from the original sample before further processing and analysing to reduce the dynamic range of the sample mixture. This makes detection of less abundant HCPs easier, and when combined with the right software package, makes the data analysis a breeze. This strategy may be taken further by leveraging the increased scan speed and increased sensitivity of the Thermo Fisher Scientific Exploris 480 Orbitrap technology which helps to identify more HCP species, as well as adding confidence to the results.
Coming up at the virtual CASSS MS meeting there is a great presentation by Michael Poltash (Janssen Inc.) at the Thermo Scientific technical seminar. Entitled ‘HCP Analysis by Internal Standard-Triggered Assay of a Panel of Heavy Tryptic Peptides of the Most Common and Troublesome CHO-HCPs’, this talk will highlight a new approach to generate clearance profiles for each HCP through successive purification steps. Having the right tools in place for each experimental step provides confidence not just in the generated results but also to ensure you haven’t missed an important component prevent a win. It’s all about starting the race knowing you’ll make it to the finish line in record time.