Scientists from Children’s Medical Research Institute (CMRI) have successfully tested adeno-associated viral vector-based (AAV) gene therapies in whole human livers in the lab. The study, which is published in Nature Communications in a paper titled “Harnessing whole human liver ex situ normothermic perfusion for preclinical AAV vector evaluation,” demonstrates the potential of using a normothermic liver perfusion system—a human liver preserved ex situ in the lab at human body temperature—to test early-stage gene therapies.

This paper is the fruit of a project that began last year when CMRI’s translational vectorology research unit partnered with a team at Royal Prince Alfred Hospital to develop a method of keeping a human liver alive in the lab. The idea was to use the system to test AAV-based vehicles for delivering payloads to treat various inherited diseases. If the system was successful, it would address the challenge of finding effective preclinical models that properly replicate human physiological conditions and systems and reliably predict clinical outcomes. 

The liver is a complex organ with diverse cell types contributing to its structure and function, and that kind of complexity is difficult to replicate using organoids, mice, and nonhuman primates. The results reported in Nature Communications are “very exciting for us because now, for the first time, we can assess the function of gene therapeutics directly in the clinical target organ itself,” said Leszek Lisowski, PhD, unit head of CMRI’s translational vectorology unit and senior author on the publication. “Up to now the gene therapy delivery tools have been tested in animal models, which while invaluable to evaluate safety and targeting of other organs/tissues, do not adequately replicate the functionality of these delivery methods in the patient.”

According to the paper, the researchers used two whole human livers perfused with human blood to evaluate fourteen natural and bioengineered AAV vectors. The liver system “uses an open venous reservoir and incorporates two long-term oxygenators, a gas blender equipped with a pediatric flow regulator for ventilation control, and a flow-adjustable dialysis membrane for water-soluble toxin filtration and perfusate volume control.” Both livers had been deemed unsuitable for transplantation and were consented for research use. The scientists used next-generation sequencing to quantify which vectors had the lowest rates of clearance in the perfusate, and to assess the cell entry performance and transgene expression for each vector, among other studies.

Besides evaluating novel AAV-based therapeutics, Lisowski noted that the liver model could be used to more accurately estimate the effective dose of new therapeutics and identify potential toxic side effects. “The current generation of viral vectors we use to deliver gene therapeutics to the liver are not good enough for the majority of clinical applications. At the moment we often have to use these therapies in high doses to overcome their functional inefficiencies and achieve clinical benefit.”