Biology subjects

Cabanes-Creus, M.

Publications and source records attributed to Cabanes-Creus, M..

3 recordsLinked to original sources

bioRxiv2023

ADEVO: Proof-of-concept of Adenovirus Directed EVOlution by random peptide display on the fiber knob

Directed evolution of viral vectors involves the generation of randomized libraries followed by artificial selection of improved variants. Directed evolution only yielded limited results in adenovirus vector (AdV) development until now, mainly due to insufficient complexities of randomized libraries. Clinical applications of AdVs as gene therapy or oncolytic vectors are still hampered by the predetermined tropism of natural types. To overcome this challenge, we hypothesized that the technology of randomized peptide insertions on the capsid surface can be incorporated into the AdV bioengineering toolbox for vector retargeting. Here we developed Adenovirus Directed EVOlution (ADEVO) protocols based on fiber knob peptide display. As a proof-of-concept, HAdV-C5-derived libraries were constructed following three distinct protocols and selected on A549-DCAR cells that lack the HAdV-C5 primary receptor, with the goal of identifying variants able to infect and lyse these tumor cells more efficiently. All protocols enabled the construction of high complexity libraries with up to 9.6x10^5 unique variants, an approximate 100-fold improvement compared to previously published AdV libraries. After selection, the most enriched variants did not display enhanced infectivity but rather more efficient replication and cell lysis. This warrants investigations into potential unsuspected involvement of the fiber protein in adenovirus replication. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/567388v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@187f7baorg.highwire.dtl.DTLVardef@1990555org.highwire.dtl.DTLVardef@12d3c60org.highwire.dtl.DTLVardef@d703da_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2023

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

Developing clinically predictive model systems for evaluating gene transfer and gene editing technologies has become increasingly important in the era of personalized medicine. Liver-directed gene therapies present a unique challenge due to the complexity of the human liver. In this work, we describe the application of whole human liver explants in an ex situ normothermic perfusion system to evaluate a set of fourteen natural and bioengineered adeno-associated viral (AAV) vectors directly in human liver, in the presence and absence of neutralizing human sera. Under non-neutralizing conditions, the recently developed AAV variants, AAV-SYD12 and AAV-LK03, emerged as the most functional variants in terms of cellular uptake and transgene expression. However, when assessed in the presence of human plasma containing anti-AAV neutralizing antibodies (NAbs), vectors of human origin, specifically those derived from AAV2/AAV3b, were extensively neutralized, whereas AAV8-derived variants performed efficiently. This study establishes the use of normothermic liver perfusion as an invaluable preclinical model for evaluating liver-targeted gene therapies and providing guidance for making essential decisions that promote the most effective translational programs.

molecular biology↗

bioRxiv2022

Assessment of pre-clinical liver models based on their ability to predict the liver-tropism of AAV vectors

The liver is a prime target for in vivo gene therapies using recombinant adeno-associated viral vectors (rAAV). 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 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 six AAV vectors in twelve preclinical models of the human liver, with the aim to uncover which model is the most relevant for the selection 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.

bioengineering↗