As gene therapies advance towards clinical trials, comprehensive understanding of biodistribution and tropism becomes crucial. The International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) recently released the ICH S12 guideline (https://www.ich.org/page/safety-guidelines), which recommends that robust preclinical biodistribution studies are required in gene therapy development. The ICH S12 has now been recognised and supported by every major regulatory agency on the planet.
Biodistribution studies explore how gene therapy vectors are distributed throughout the body, while tropism studies focus on the target cell specificity of these vectors. Both studies provide critical insights into the potential off-target effects of gene therapies.
What is the ICH S12 in a nutshell?
The ICH S12 sets regulatory expectations for nonclinical (preclinical) approaches to assess the biodistribution profile of a gene therapy in relation to the administration route, dose, dosing regime, and immune response. These studies can be conducted as stand-alone or in conjunction with pharmacology and toxicology studies. A challenge that the S12 recommendations present for the gene therapy R&D community is the use of alternative methods that still provide meaningful data while minimising animal use.
Limitations with existing models
While mice and non-human primates (NHPs) have contributed significantly to drug development, they have limitations in translating findings to humans, and this is particularly relevant in the gene therapy field. Mice and NHPs differ significantly from humans in terms of their susceptibility to specific viruses and the resulting disease manifestation. NHPs, and to some degree, rodents, are highly conserved at the genetic level, but have evolved in starkly different ecological niches – with different diets and habitats, exposure to microbes, and disease susceptibilities. This explains why the underlying molecular and physiological processes often diverge, and this dissimilarity has limited the translational value of existing models in studying the intricacies of viral infections and developing targeted gene therapies.
The use of inappropriate rodent and NHP models has most likely been a major factor in the high incidence of serious adverse events (SAEs) reported in clinical trials of gene therapies – which we can presume, were not observed at the preclinical stage.
So What Species?
Humans and pigs have coexisted in the same environment for over 10,000 years, sharing habitats and food sources, which in turn has influenced their microbiota — the communities of microorganisms that inhabit their bodies. Pigs and humans share similar gastrointestinal environments and are exposed to comparable external factors, leading to overlaps in their gut microbiomes. This parallelism in microbiota composition and function between pigs and humans strengthens their biological connection.
The Evolutionary Parallel between humans and pigs
The coexistence of pigs and humans has also revealed similarities in their susceptibility to diseases. Pigs are susceptible to a range of infectious diseases that affect humans, including viral, bacterial, and parasitic infections. As a result, they are an ideal platform to study biodistribution, tropism, transfection, transduction, safety, and efficacy of a gene therapy. Their similarities to humans in terms of drug absorption, distribution, metabolism, and excretion make them valuable surrogates for predicting gene therapy behaviour in humans. This parallelism allows for more accurate evaluation of drug efficacy and safety, reducing the reliance on less predictive animal models like rodents.
Pigs create intricate social structures and possess complex cognitive abilities. They undoubtedly feel pain and experience fear and distress. As a result, the ethical process to gain approval to perform pig research is time-consuming, and experiments can be restricted. It is also incredibly expensive. The ability to evaluate a panel of potential therapies in-line with the ICH S12 (for biodistribution, tropism, safety, and efficacy), and iterate experiments based on findings becomes almost impossible, taking years to complete and wasting multi-millions of R&D budgets. But, the solution is not to continue using inadequate models that transition a gene therapy through the preclinical stage, only to fail in clinical trials.
So what is the solution?
We consume 1.5 billion pigs per year for food. In most cases, internal organs are discarded or used in animal feed. Pebble recognised that this was an incredibly valuable source of biological material. The Pebble team had worked in transplantation for 25 years, developing technology called ex-vivo organ perfusion, with the goal of maintaining donor organs in optimal health, outside of the body.
In 2016, the team set out to evaluate if organs from pigs entering the food chain could be used, in combination with ex-vivo organ perfusion, as a platform to test drugs and medical devices, without the need for lab animals. By replicating blood flow and oxygenation, ex-vivo organ perfusion allows researchers to observe the behaviour of drug in a realistic, physiological environment, providing valuable insights into safety and efficacy. 1200 experiments later, the Pebble team have evolved this technology to the next-generation LIVING-ORGAN system (see www.pebble.bio/living-organ-systems for more info), where complete, physiological organ function is restored.
Connecting Different Organs in a Perfusion Circuit: The LIVING MULTI-ORGAN system.
Linking different types of organs in a single perfusion circuit offers a unique opportunity to study inter-organ interactions and systemic biodistribution dynamics. By connecting organs relevant to the therapeutic target, as well as those involved in metabolism, clearance, or potential off-target effects, the Pebble team can simulate the dynamic interactions between organs within a ‘living organism’. This approach enables the examination of how gene therapy vectors distribute and interact within a MULTI-ORGAN system, providing a more holistic understanding of biodistribution and potential therapeutic implications.
Stealth-mode R&D through to Submission Ready Data-packs
A major advantage of Pebble’s approach, is that the LIVING-ORGAN system is not recognised as animal research, meaning early evaluations of a therapy can be performed, that do not need
to be included in future submissions to the regulatory authorities. This gives gene therapy developers the time to fail, iterate and repeat, until the desired biodistribution, off-target effects, safety and efficacy have been reached. At this point the Pebble team design a panel of bespoke experiments, tailored specifically to the needs of the regulatory authority – so a purpose built, submission ready Datapack is generated.
The ICH S12 has been recognised by almost every regulatory agency in the world and will now play a pivotal role in guiding nonclinical development of gene therapy products, emphasising the importance of biodistribution studies. LIVING-ORGAN systems have emerged as a promising laboratory method, offering controlled conditions to study biodistribution patterns, tropism, off-target effects, safety, and efficacy. Pig organs, with their anatomical and physiological similarities to humans, present a valuable model for predicting clinical outcomes. Moreover, the integration of different organs in the same perfusion circuit provides a novel approach to studying inter-organ interactions and systemic biodistribution dynamics. This integrated approach enhances our understanding of how gene therapy vectors travel across various organs, potentially impacting efficacy, and safety.