Understanding the pharmacokinetics of a drug – including how drugs are absorbed, distributed, metabolized, and excreted by the body – is crucial to its success through the drug development pipeline, ensuring new therapies are safe and produce the desired effect.
One protein that has recently gained attention for its potential in providing valuable pharmacokinetic data is the neonatal Fc receptor (FcRn). FcRn plays a key role in regulating the half-life and bioavailability of therapeutic antibodies. In this article, we speak with Greg Christianson, Study Director for In Vivo Services at The Jackson Laboratory. During Christianson’s early career, he worked in a lab that contributed to identifying the FcRn protein. Once its function was characterized, the team saw the potential in leveraging the protein in pharmacokinetic studies by expressing a human FcRn transgene in mice. Here, Christianson discusses the role of FcRn in preclinical studies for therapeutic antibodies, and the suite of mouse models that The Jackson Laboratory has created for such studies.
Therapeutic antibodies are a rapidly growing class of drugs that can be used to treat a wide range of diseases, including cancer, autoimmune disorders, and infectious diseases. These antibodies are typically administered intravenously and generally have a long half-life, which is critical for their efficacy. FcRn is a receptor that can bind to the Fc region of antibodies and the albumin-binding domain of albumin, and helps to protect them from degradation, extending their half-life and improving their efficacy.
“FcRn binds molecules that have an Fc region or have albumin, in a pH dependent manner,” describes Christianson. After the proteins are internalized by the cell via pinocytosis and targeted for lysosomal degradation, the pH of the endosomes starts to decrease. “At around pH 6, the antibodies bind to FcRn and are transported back up to the cell surface. Here, they are exposed to the neutral pH of the circulating blood and are released from FcRn. This process rescues them from lysosomal degradation and transports them back into circulation,” Christianson adds. Consequently, these proteins that have an affinity for FcRn are cleared at a slower rate.
“Antibodies play an important role in your body's immune memory by provoking an immune response as they mature from IgM to IgG antibodies,” explains Christianson. This process is known as IgG recycling, and it helps to maintain a persistent level of IgG antibodies in the body. Additionally, FcRn can also transport IgG antibodies across the basolateral membrane of endothelial cells lining vasculature, allowing them to access target tissues more effectively.
In the development of a drug, FcRn can be used to gain pharmacokinetic data by measuring the concentration of IgG antibodies in the bloodstream over time. By monitoring the clearance rate of an IgG antibody, researchers can determine its half-life and gain insights into its distribution and exposure within the body. This information can be used to optimize dosing regimens and improve the efficacy and safety of therapeutic antibodies.
Furthermore, FcRn-mediated transport can also be exploited to improve the therapeutic benefits and minimize potential risks of a drug. “Antibodies, albumin, and therapeutics that take advantage of these molecules, are prolonged in circulation due to their affinity for FcRn. Scientists have investigated sequence changes in the Fc region of antibodies and identified mutations that can prevent binding to FcRn, leading to shorter half-lives of therapeutic proteins,” says Christianson. “That's important if your therapeutic has any off-target toxicity. In such cases, it is necessary to eliminate any unbound therapeutic from the body quickly, so a shorter-lived antibody is desirable. Alternatively, to develop a long-lasting therapeutic that requires less frequent dosing, researchers can search for Fc variants with higher affinity for FcRn,” Christianson continues. “Once identified, these variants can be evaluated in preclinical studies using mice to determine if they have an extended half-life.”
The Jackson Laboratory has developed a range of genetically engineered mouse models that can be used to study FcRn function and its role in antibody pharmacokinetics. “We've got multiple models that we use which have unique applications and solve different problems,” Christianson explains. FcRn is a key component of the Therapeutic Evaluation Services (TES) offered by The Jackson Laboratory, which are a suite of preclinical drug testing and in vivo pharmacology services that can be used by researchers and pharmaceutical companies to gain pharmacokinetic and efficacy data for their therapeutic antibodies.
Developing effective drug candidates is a challenging task for pharmaceutical companies, requiring reliable pharmacokinetic data to ensure their efficacy and stability in living organisms in line with FDA regulations. Christianson explains, “Non-human primates (NHP) have been the standard model for preclinical data, but ethical issues and high costs make this approach problematic. Adding to these downsides is the common occurrence of anti-drug antibodies (ADAs).” To address these issues, Christianson and his team at The Jackson Laboratory developed Tg32 mice, with a human FcRn sequence that can provide an accurate prediction of therapeutic performance in humans. “Side-by-side comparisons with NHP showed that Tg32 mice perform equally well or better in predicting therapeutic performance. This data can be used for allometric scaling to determine how these candidates will perform in human patients,” Christianson shares.
Drug discovery often requires the use of diverse mouse models to study the complex mechanisms of diseases, as different mouse models can mimic different aspects of human diseases. A large fraction of therapeutics being developed using mouse models are antibodies or Fc-based. However, there have been instances where Fc-based therapeutics developed for clinical use had to be discontinued due to their severe effector functions that caused serious illness in patients. Designing albumin into your therapeutic will protect it from lysosomal degradation in the same way but does not have the risk of effector function activation. Hence, albumin is a promising molecule being used to increase the half-life of therapeutics in patients.
Albumin-based therapeutics are a promising avenue for drug developers to explore. “However, the standard Tg32 mouse produces mouse albumin, which by chance, has a strong affinity for human FcRn,” Christianson explains. “As a result, when human albumin or a human albumin-based therapeutic is introduced in these mice, it is unable to compete with the high-affinity mouse albumin, leading to the impression of a short half-life when using Tg32 mice.” To overcome this problem when testing albumin-based therapeutics, The Jackson Laboratory developed a Tg32 mouse that is deficient of mouse albumin. “In the absence of mouse albumin, you get half-lives that, again, are comparable to what human patients would have,” Christianson shares.
Another challenge for in vivo modeling is immunogenic responses in NHP and mice, that can result in the formation of ADAs and immune complexes, leading to inaccurate pharmacokinetic data. However, Tg32 mouse models developed by The Jackson Laboratory that are immunodeficient can be utilized to avoid ADA responses.
Christianson also describes the Tg276 mice models which have a reduced FcRn function and can help researchers accelerate the screening process for antibody candidates. Using Tg276 mice with lower FcRn function allows for faster screening of a large panel of antibodies to find the best candidates. Despite having reduced protection compared to Tg32 mice, Tg276 mice remain predictive of antibody pharmacokinetics in humans. “Ranking antibodies by half-life in Tg276 mice corresponds well with results in human patients or NHPs,” Christianson explains. By reducing the large panel to a few promising candidates and conducting a longer characterization study in Tg32 mice, researchers can obtain reliable results in less time.
By utilizing these models, we can provide valuable insights into the efficacy and mechanism of action of therapeutic antibodies and other molecules.
The Jackson Laboratory
Christianson explains that the services The Jackson Laboratory provides which utilize these mouse models include pharmacokinetic characterization and pharmacodynamic studies. “One example is drug developers who target FcRn itself, the protein responsible for rescuing antibodies from degradation, by blocking its recycling function. This can be effective in treating autoimmune diseases where autoantibodies are driving the disease as it can reduce those autoantibodies,” he says. “To study the pharmacodynamic response, we preload human IgG into Tg32 mice to establish circulating levels, then administer the therapeutic to block FcRn function and observe the decrease in IgG levels.” Christianson shares, “By utilizing these models, we can provide valuable insights into the efficacy and mechanism of action of therapeutic antibodies and other molecules.”
The Jackson Laboratory is continually developing its models to improve the services it provides to its customers. Christianson describes two ongoing projects. The first aims to improve the NOD scid gamma (NSG™) mouse, which replicates the human immune system and is commonly used to study cancer. “The mouse has a unique mouse allele of Fc gamma receptor I (FcgRI) that binds to human IgG antibodies with high affinity, creating an antibody sink and interfering with efficacy studies,” Christianson explains. To solve this problem, a mouse model is being developed with a human FcGR1A knocked into the mouse FcgR1 locus. It is hoped this will make the NSG mouse a more efficacious model for testing human antibody therapeutics. The second project involves creating a new Tg32 mouse model that uniquely expresses human albumin. This is relevant for studies that use human albumin as a carrier molecule or as a competitor for FcRn protection. The model is close to completion and has promising applications for drug development.