Assessment of Pre-Clinical Liver Models Based on Their Ability to Predict the Liver-Tropism of Adeno-Associated Virus Vectors

Abstract

The liver is a prime target for in vivo gene therapies using recombinant adeno-associated viral vectors. Multiple clinical trials have been undertaken for this target in the past 15 years; however, we are still to see market approval of the first liver-targeted adeno-associated virus (AAV)–based gene therapy. Inefficient expression of the therapeutic transgene, vector-induced liver toxicity and capsid, and/or transgene-mediated immune responses reported at high vector doses are the main challenges to date. One of the contributing factors to the insufficient clinical outcomes, despite highly encouraging preclinical data, is the lack of robust, biologically and clinically predictive preclinical models. To this end, this study reports findings of a functional evaluation of 6 AAV vectors in 12 preclinical models of the human liver, with the aim to uncover which combination of models is the most relevant for the identification of AAV capsid variant for safe and efficient transgene delivery to primary human hepatocytes. The results, generated by studies in models ranging from immortalized cells, iPSC-derived and primary hepatocytes, and primary human hepatic organoids to in vivo models, increased our understanding of the strengths and weaknesses of each system. This should allow the development of novel gene therapies targeting the human liver.

Progress in developing microphysiological systems for biological product assessment

“…In conclusion, the study demonstrated that while AAV5 displayed greater transduction efficiency in hLTEs than AAV3b, both groups exhibited compromised self-aggregation and diminished hepatocyte functionality, warranting further attention and investigation. Similarly, Westhaus and colleagues 70 conducted a functional evaluation of six AAV vectors in twelve in vitro models of the human liver including immortalized cells, iPSC-derived and primary hepatocytes, as well as primary human hepatic organoids, and in vivo models Fig. 2b.…”

Targeted Gene Insertion: The Cutting Edge of CRISPR Drug Development with Hemophilia as a Highlight

“…Tissue-specific transgene expression is preferred but is challenging in AAV-delivered gene augmentation. While naturally occurring or engineered capsids may result in distinct tissue tropism [ 101 ], the features differ considerably between preclinical models and human patients, and exclusive specificity to certain tissues or cell types has not been conferred [ 102 ]. Markedly, targeted insertion of promoterless sequences restrains transgene expression under the control of endogenous target loci [ 49 ], which provides an excellent solution to introduce tissue/cell type-specific expression.…”

Harnessing the power of whole human liver ex situ normothermic perfusion for preclinical AAV vector evaluation

“…In order to minimize the influence of neutralizing antibodies, we removed the majority of antibodies using a previously validated immunoadsorption method. 9 After antibody depletion, the end titer, calculated for AAV9, was 1:30 (Materials and Methods). We then intravenously infused a total dose of 3.5×10 12 vector genomes (1.4×10 12 vg kg -1 ) of a new barcoded AAV mix (Supplementary Fig.…”

“…We next evaluated the vector mix in a recently developed FRG xenograft murine model engrafted with primary hepatocytes from cynomolgus monkeys. 9 To do so, we injected highly engrafted animals (n=2) with albumin levels 10 mg mL -1 blood (equivalent to estimated >80% repopulation level) with 1´10 11 total vg of the AAV mix (approximately 7.5´10 9 vg per capsid) and harvested the chimeric livers two-weeks post-infusion.…”