Hybrid nanoparticles of extracellular vesicles and adeno-associated viruses (AAVs) as an innovative gene-replacement therapy for rare monogenic diseases
lead supervisor: prof matthew wood, Department of Paediatrics
Co-supervisor: Dr Mariana Conceição, Department of Paediatrics
Commercial partner: Evox Therapeutics, Oxford
Adeno-associated viruses (AAVs) are the leading gene therapy vectors for clinical application, due to their high efficiency in transducing multiple target tissues and acceptable single dose safety profile. However, the immune system is a major limiting factor given that a significant proportion of the human population naturally possesses neutralizing antibodies (NAbs) against AAVs, and further, NAbs rapidly develop at high titers upon AAV vector administration, precluding further dosing. Therefore, the full potential of AAV gene therapy will not be achieved unless scientific methods to enable (re-)administration of AAVs in the presence of NAbs are developed. Extracellular vesicles (EVs) are natural, non-viral, nano-sized particles with high potential for drug delivery, including delivery of genetic drugs and nucleic acid cargoes. Previous studies showed that during viral production, a fraction of AAVs exits the AAV-producing cells by hijacking cellular membranes (EV-AAVs). When compared to conventional AAVs, EV-AAVs have been shown to promote enhanced gene transfer, and to efficiently protect AAVs from NAbs.
Here, we propose to maximise the clinical potential of EV-AAV technology by completing the following aims/milestones: 1) increase the incorporation of AAVs inside EVs, 2) develop industrially scalable methods to isolate EV-AAVs using cells with immunomodulatory potential as the EV-AAV cell source, and 3) test the therapeutic efficacy of the developed EV-AAVs in the context of a rare monogenic disease, phenylketonuria, for which there is high unmet clinical need. In aim 1, we will increase the incorporation of AAVs inside EVs by engineering AAV-producer cells, AAVs and/or by stimulating enhanced EV release from AAV-producer cells. We have already been able to show binding of AAVs to EVs upon EV engineering, but this strategy and other alternatives will be further explored in this work. The protection against NAbs and the biodistribution of the developed EV-AAVs will also be tested at this stage. Parallel studies evaluating different scalable methods to isolate EV-AAVs will also be tested, as differential ultracentrifugation, that was reported in previous publications, is not a scalable method (Aim 2). Additionally, as prior studies used HEK293T cells to produce EV-AAVs, here we will also investigate the possibility to obtain EV-AAVs from cells with immunomodulatory potential. Lastly, in aim 3, we will test the therapeutic efficacy of phenylalanine hydroxylase delivery in PKU in vitro and in vivo models of disease.
The partnership with Evox Therapeutics, a world-leading company developing transformative EV therapeutics, that will provide access to highly innovative techniques and industry expertise, together with our vast experience working with EVs as therapeutic agents and in the field of rare genetic diseases, will allow us to build an innovative gene replacement strategy, harnessing the best characteristics of EVs and AAVs in a single system. This will result in a fundamentally new way to treat patients suffering from rare monogenic diseases without exceptions, safely, efficiently, and repeatedly. This project will capitalize on the strengths of both our laboratory and Evox Therapeutics, providing the opportunity for the student to gain skills in state-of-the-art techniques in whole organ physiology, translational medicine, nanomedicine, and gene therapy.