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How well are your HCP ELISAs covered?
By Joe Hirano

Program Manager, Imaging, Cytiva

V

alidating host cell protein ELISAs is an essential
part of biologics development. But what makes
these coverage assays so vital, and what could
we improve? Read on, and see how our 2D DIBE™
approach combines the best of existing methods.
What is a coverage assay?
Coverage assays provide a means to validate the
enzyme-linked immunosorbent assay (ELISA) used in
host cell protein (HCP) quantitation. They enable you
to estimate the percentage of HCPs that can be detected, or "covered", by ELISAs, and direct you toward filling
any gap. The goal is to develop processes for reducing
HCPs, improving drug efficacy and patient safety, and
meeting regulatory guidelines. These ELISAs and coverage assays are an essential part of process development in biologics manufacture, and they can have an
impact on the potential success of a drug.
Here, I'll take you through the strengths and weaknesses
of one of the most common types of coverage assays
and outline an approach that retains its best traits
while enhancing speed and accuracy.
The need to stay covered in HCP quantitation
Biopharmaceutical companies have a duty to
make sure their products meet safety regulations.
Optimizing purification steps and using ELISA
to monitor the presence and quantity of HCPs
throughout are two key ways to do this.
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| GENengnews.com

ELISAs are the workhorse of HCP quantitation, but
they aren't perfect. Raising antibodies against HCPs is
not 100% effective, as not all HCPs are immunogenic
enough to trigger a response in the animal host.
Given the potential for HCPs to affect patient safety,
the U.S. Pharmacopeia and the Pharmacopoeia
Europaea (USP 1132 and Ph. Eur. 2.06.34) state that
ELISAs should also be monitored. This involves characterizing and validating the antibodies by coverage
assays to check whether they have the expected
sensitivity, specificity, and coverage.
Several biologics in recent years have shown immunoreactions in late-stage clinical trials. These include
Sandoz-Novartis Omnitrope®, Ipsen coagulation factor IX,
and Genentech Lebrikizumab. This was a serious safety
concern and led to delays in their approvals.
As a result, 2016 and 2017 saw updates to the U.S.
Pharmacopeia and the Pharmacopoeia Europaea,
respectively, to cite the usage of coverage assays
as best practice for risk management for any future
safety issues. Although these assays add some extra
cost to product development, this is outweighed by
the potential risk of delays in product approval.
The new guidelines recommend both generic assay
revalidation for every new batch of reagent, as well as
process- and platform-specific assay revalidation after

Figure 1A. Existing standard coverage assay workflow for
2D gel electrophoresis/Western blot.



https://www.ncbi.nlm.nih.gov/pubmed/24995961 https://www.ncbi.nlm.nih.gov/pubmed/24995961 http://www.usp.org/biologics/proteins https://www.edqm.eu/en/european-pharmacopoeia-ph-eur-10th-edition https://www.ncbi.nlm.nih.gov/pubmed/25998019 https://www.ncbi.nlm.nih.gov/pubmed/25998019 https://www.ncbi.nlm.nih.gov/pubmed/18059081 https://www.ncbi.nlm.nih.gov/pubmed/26915600 https://www.ncbi.nlm.nih.gov/pubmed/27739010 http://www.GENengnews.com

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