Pharmaceutical Outsourcing Q3 2023 - 20

ANALYTICAL TESTING
New Methods for Studying Plasma
Protein Binding
We know that high-throughput screening (HTS) in ADME studies allows
for rapid, simultaneous testing of multiple oligo candidates under
varying conditions. These tests enhance the efficiency and breadth
of data collection, accelerating the early stages of drug development.
However, the accuracy of HTS has been criticized due to the potential
for false positives and negatives, not to mention the question of how
well HTS reflects the complexities of the human body.
However, technological advancements and improved test designs
have the potential to revolutionize in vitro ADME studies, paving
the way for more accurate and comprehensive plasma protein
binding analysis. For example, microscale thermophoresis (MST) is an
alternative plasma protein binding analysis approach. This method
tracks the movement of molecules in a temperature gradient,
enabling precise quantification of protein-drug interactions without
the need for chemical modifications to the oligo.
Liquid chromatography-mass spectrometry (LC-MS) is a highly sensitive
and specific tool that can separate components and deliver data on
each compound's molecular mass. This technology provides detailed
insight into the protein binding assay, integrated with ultrafiltration or
ultracentrifugation to separate unbound oligos from plasma.
Plasma Protein Binding in Oligos
Considering the unique challenges associated with studying
plasma protein binding in oligos, specific methods tailored to these
molecules are gaining traction, too. Advanced techniques include
using capillary electrophoresis and ultrafiltration coupled with mass
spectrometry. Electrophoretic mobility shift assays are also reported
for the characterization of siRNA PPB. These methods are designed
to handle oligos' size, charge, and binding characteristics, providing
more reliable and accurate results.
Furthermore, computational modeling and simulation hold
immense potential for studying plasma protein binding in oligos.
By leveraging in silico models-including physiologically based
pharmacokinetic (PBPK) models-we can simulate complex
interactions between oligos and plasma proteins at a molecular
level, bypassing some experimental constraints. These models
incorporate diverse parameters from the drug's characteristics
to patient physiology, facilitating the prediction of tissue drug
concentrations over time. This allows us to anticipate binding sites,
affinities, and potential off-target interactions, thus enriching our
understanding of oligo pharmacokinetics and aiding in developing
safer and more effective therapeutics.
Refining these technologies-and continuing to develop new ones-
will undoubtedly provide deeper insights into plasma protein binding
and help scientists overcome the inherent challenges in studying
oligo drugs. Continued advancements in this area are essential to
harness the full potential of oligo therapeutics and to develop more
effective and safer drugs.
Regulatory Considerations in Plasma
Protein Binding Studies
From a regulatory perspective, in vitro plasma protein binding
studies are essential to the preclinical evaluation of a new drug.
Regulatory agencies like the U.S. Food & Drug Administration (FDA)
and the European Medicines Agency (EMA) have issued guidance
on these studies.
Generally speaking, regulators expect detailed information about a
drug's plasma protein binding properties, including:
* The percentage of drug bound;
* The identity of the primary binding proteins;
* The influence of disease states on drug binding.
That said, regulatory guidance often lacks specific protocols for
conducting these studies, leading to variations in study design and
interpretation across different labs. There's a need for harmonized
guidelines and best practices for conducting plasma protein binding
studies, particularly for novel therapeutics like oligos.
Protein binding is a multi-faceted process with significant drug
disposition and efficacy implications. Given the increasing complexity
of new therapeutic entities, rigorous in vitro studies coupled with
in silico models and careful consideration of pathophysiological
influences are necessary for predicting in vivo behavior accurately.
Further refinement and harmonization of regulatory guidelines could
clarify and promote consistency in these vital studies.
A Final Word
Plasma protein binding constitutes a critical component of drug
pharmacokinetics, heavily influencing a drug's distribution, clearance
rate, and efficacy. This is particularly pertinent for oligonucleotides,
a class of therapeutics whose large size, negative charge and
chemical modifications intensify the complexity of their interaction
with plasma proteins-however, technological advancements and
experimental design promise to overcome these hurdles.
MST and LC-MS have the potential to enhance our understanding of
protein-drug interactions. At the same time, computational modeling
and simulation may prove invaluable in exploring molecular-level
interactions and anticipating drug behavior. These advancements
and the harmonization of regulatory guidelines could pave the
way toward developing safer, more effective oligo therapies and
ultimately improved patient outcomes.
Dr. Jie Wang received her PhD in pharmaceutical analysis from Shenyang
Pharmaceutical University. She is now a principal scientist in DMPK services at
WuXi AppTec, focusing on in vitro ADME with expertise in protein binding and
drug metabolic stability assays.
Pharmaceutical Outsourcing | 20 | July/August/September 2023
Pharmaceutical Outsourcing Q3 2023

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