The availability of plasma-derived medicinal products—
one of the earliest achievements of medical
biotechnology—has enabled great progress in the
treatment of specialized conditions such as hemophilia
and immune deficiencies. Yet early on, the biologic materials
used to develop these products were also found
to be vulnerable to infectious disease agents. Today,
manufacturers safeguard these products during the
development process through a set of measures commonly
referred to as the “Safety Tripod” (Figure 1).
This consists of the selection of plasma donors with
a low risk of contact to infectious agents, the testing
of plasma donations for the absence of selected
infectious agents and, finally, virus inactivation and
removal (=reduction) processes. These measures are
also required by regulators (1).
Figure 1 Safety Tripod
With time, it has become clear that the reduction
capacity is by far the most significant
quantitative contribution to product safety
margins (Figure 1). For example, since
the arrival of West Nile virus in the United
States, directly transfused blood product—the
safety margins of which depend exclusively on
donor selection and donation testing, as they
typically do not undergo any virus-reducing manufacturing
process—has occasionally transmitted the
virus, despite testing using modern and very sensitive
nucleic acid-based methods (2). In contrast, plasma derivatives
have been safe, even without West Nile virus testing of
plasma for fractionation, as their manufacturing processes are more
effective at inactivating or removing the virus (3).
Due to the success of the “Safety Tripod” concept, biotechnology
manufacturers of therapeutic proteins have
adapted it for their own processes. Arguably, this product
class has never been reported to transmit a virus
to a recipient, yet contamination of manufacturing
platforms has occurred. As to the specific interventions
applied to ultimately enhance product
safety margins, a careful selection process
is used to minimize any risk of exposure to an infectious agent for the production
cell line as well as for raw materials entering
the manufacturing process. The chosen
cell banks as well as individual fermenter
harvests are subject to testing to ensure the
absence of infectious agents. And finally,
virus reduction processes are implemented
into the downstream purification process
for biotechnology products.
Now, advanced therapy medicinal products
(ATMPs) are entering the market, offering
potential advancements for maintaining
and improving human health, just as
plasma-derived medicinal products did
in the mid-20th century. ATMPs face the
same contamination threats from exposure
to universally present and effective
opportunistic agents in the microbiological
environment as traditional biologics. In
recent years, the manufacturing platform of
an already licensed ATMP was found to be
contaminated with a virus, fortunately one
not pathogenic to humans (4). This led the
manufacturer to add a nanofiltration step
for this product, following recommendations
for a virus reduction method.
Therefore, it is important to embrace the
safety concepts that have been so effective
in protecting more traditional biotechnology products. With ATMPs, this begs the
question—how can this technically be
accomplished?
Manufacturers Swim Upstream
The selection and testing procedures used
in biotechnology have at times failed the
expectations placed in them. Fortunately,
this situation is expected to improve
as testing becomes as innovative as the
end product itself. Take next-generation
sequencing, for example. This technique
is now being used increasingly during
characterization of cell banks. It establishes
the absence of adventitious agents
without any prior knowledge about them.
But advances in virus reduction processes
offer a more solid solution. While options
for virus reduction may be limited, they
do exist. A publication from the German
Paul-Ehrlich-Institute showed that Adenoassociated
virus (AAV) gene therapy vectors
can be treated with solvent-detergent
(SD) combinations to inactivate any
lipid-enveloped adventitious viruses. This
has no impact on the nonlipid enveloped
therapeutic entity; furthermore, larger
pore size nanofilters can remove large
adventitious viruses with effective passage
of the very small AAV (5).
Even more innovatively, any risk associated
with the starting material of a biotechnology
process can also be separated from
the final product, and, ultimately, the
patient, by a virus reduction barrier placed
upstream rather than the traditional downstream
(Figure 2). In fact, for ATMPs,
such as large lipid-enveloped virus gene
therapy vectors and similar cell-based therapies,
it may not be possible to apply virus
reduction technologies to the product or
production intermediate containing the
active drug substance. Thus, ensuring viral
safety of all raw materials used in cell culture
is highly important and applying virus
reduction methods at the raw material level significantly diminishes the contamination
risk for cell cultures. An upstream
intervention may be the only technically
feasible means of providing virus reduction
capacity within the manufacturing process.
More importantly, an upstream barrier
approach not only offers additional safety
margins for the final product but also
protects the fermenter from exposure to an
infectious agent. Otherwise, any minimal
inoculum might result in exponential amplification
of the agent, potentially to titers
that may even overwhelm any downstream
virus reduction capacity. In addition,
maintaining the integrity of the manufacturing
setting by avoiding any exposure
results in an uncompromised ability to
serve the patients waiting for the respective
medicinal product.
Figure 2 Upstream versus Downstream Virus Barrier
With technological progress in the ATMP
space so incredibly rapid in recent years,
regulators are now establishing or refining
procedures to convey these products to
market (6). The pathogen safety concepts rereflected
in regulatory guidance documents do
recognize some of the technical limitations
(“possibilities for applying virus clearance
steps … are limited”), yet sound conceptually
very familiar (“selection and control of
starting materials (including seed and cell
banks), raw materials...application of vector
purification process steps which, where
feasible, provide elimination/inactivation
capacities vis-a-vis relevant viruses”) (6).
It is exciting to see how top science is being
brought to fruition in a public health
setting so quickly. With all the focus on
innovation, however, it is equally important
not to forget the lessons of the past,
and to use available biomanufacturing
tools to safeguard these modern biomedicines
along the lines of proven concepts.
[Editor's Note: This article is based on the
author’s presentation delivered at PDA’s 2016
Viral Safety of ATMPs conference in Berlin.]
References
- Guideline on plasma-derived medicinal products,
EMA, 2011.
- Montgomery, S.P., et al. “Transfusion-associated
transmission of West Nile virus, United States 2003
through 2005.” Transfusion 46 (2006): 2038–2046.
- Kreil, T.R., Berting, A., Kistner, O. and Kindermann,
J. “West Nile virus and the safety of plasma
derivatives: verification of high safety margins, and
the validity of predictions based on model virus
data.” Transfusion 43 (2003): 1023–1028.
- Ma, H., et al. “Identification of a novel rhabdovirus
in Spodoptera frugiperda cell lines.” Journal
of Virology 88 (2014): 6576–6585.
- Stuehler, A., and Bluemel, J. “Viral Safety of Biological
Drugs.” Federal Health Gazette 57 (2014)
1198–1202.
- Draft guideline on the quality, non-clinical and clinical aspects of gene therapy medicinal products,
EMA, March 23, 2015.
About the Author
Thomas R. Kreil, PhD, is
Senior Director of Global
Pathogen Safety at Shire. He
has contributed to the field of
pathogen safety for vaccines as
well as plasma-derived, biotech and
ATMPs for almost two decades.