Engineering the surface expression on exosomes for improved pharmacokinetics and tissue-targeting
While the rapid clearance of exosomes from circulation and uptake into tissues can be beneficial by enabling a localised site of action, for certain applications an extended circulating half-life is desirable.
- Albumin binding exosomes to extend circulation time (OF16.01): One approach to extending plasma half-life is to engineer ligands on the surface of exosomes that bind albumin and thereby impart albumin’s long circulation half-life to these engineered exosomes. In a presentation by Liang et al., it was found that this dramatically increased the circulation time of exosomes in mice regardless of route of administration. This could provide significant benefit for both creating feasible dosing regimens and to ensure sufficient plasma residence time for tissue-specific targeting.
Engineered display of ligands on the surface of exosomes can be used to increase specific targeting of exosomes to desired tissues. In four presentations and posters, Evox scientists and collaborators, reported on improved targeting of exosomes to tumors, muscle cells, inflamed endothelium, and ability to engage with receptors involved in blood brain barrier (BBB) crossing.
- Targeted therapy by antibody displaying exosomes (OT09.05): A variety of ligands can be expressed on exosomes to help skew their uptake and distribution into specific tissues. Wiklander et al. presented on the development of an exosome engineered to express a protein capable of binding to the Fc region of antibodies (Ab). Using this as a rapid screening platform, they demonstrated the ability of Abs displayed on the exosome surface to dramatically improve by several orders of magnitude the in vitro uptake and delivery of drug cargo to the desired target cells. The studies were extended in vivo using a PD-L1 Ab-targeted exosome containing the chemotherapeutic agent, doxorubicin (dox). Following systemic administration in tumor-bearing mice, significantly increased uptake of exosomes and improved anti-tumor efficacy were seen with these drug-loaded targeted exosomes as compared to either non-targeted dox-loaded exosomes or PD-L1 Ab-targeted exosomes lacking dox.
- Exosome targeting and cell-based functional activity in the context of Pompe disease (PT12.16): Pompe disease is a rare inherited lysosomal storage disease in which lysosomal glycogen catabolism is reduced or depleted due to deficiency in acid-alpha-glucosidase (GAA). Current enzyme replacement therapy (ERT) is beneficial but poorly targets skeletal muscle. Lobo et al. designed exosomes to contain GAA and express a muscle targeting peptide on its surface. Non-targeted exosomes containing GAA were active and showed similar enzymatic activity in GAA-deficient myotubes as compared to ERT, while the muscle-targeted GAA-containing exosomes showed a significant increase in enzymatic activity and resulted in greater glycogen removal in comparison with the current ERT. In contrast to ERT, the mechanism of exosome uptake into cells was found to be independent of mannose 6-phosphate, suggesting an ability of GAA-containing exosomes to reach multiple cell types that ERT cannot.
- Exosome targeting to enhance anti-inflammatory efficacy (OS23.05): Endothelial cells at inflammatory sites upregulate E-selectin and other adhesion molecules to facilitate leukocyte infiltration via binding to the surface glycan sialyl Lewis-X (sLeX). Zheng et al. engineered exosomes to display sLeX on their surfaces and demonstrated improved uptake into activated endothelial cells and suggested a utility for site-specific targeting in inflammatory disease. In addition, display of sLeX also extended the circulation time of the exosome in several mouse models.
- Exosome targeting to the central nervous system (CNS) using transferrin receptor (PT12.10): Exosomes could be a useful modality to deliver complex therapeutics to the CNS. Antibodies (Abs) to the transferrin receptor have been shown to carry biologic drugs across the BBB. Hewlett et al. showed in cell-based studies that exosomes expressing transferrin receptor-targeting nanobodies showed significantly improved receptor engagement over control non-targeted exosomes. This study is an important step toward developing systemic-delivered exosomes engineered to cross the BBB and treat CNS diseases.
Exosome manufacture and in vivo production
Exosomes are released from all cells in the body. From a manufacturing perspective, it has not been clear whether their cell source confers manufacturing advantages and if there are optimal doses. In addition, storage conditions for exosomes are an important aspect of developing them as therapeutics and Evox has developed proprietary formulations in this regard.
- Comparison of the effect of exosome cell source on transcriptional response (PF04.04): Hagey et al. examined in a dose responsive manner the transcriptional response of cells exposed to exosomes produced from twelve different cell sources. They found that dose was a more important contributor to transcriptional response than cell source and it was only at lower exosome doses that cell source-specific responses were seen. These studies provide useful insights into the cell biology of exosome uptake and can inform the selection of exosome manufacturing cell source and optimal dosing regimens.
- Identification of storage conditions stabilizing exosome preparations (PT02.07): Görgens et al. identified storage conditions that stabilize exosomes for up to two years. Phosphate buffered saline (PBS) supplemented with albumin and trehalose improved short-term and long-term exosome preservation at temperatures ranging from 4oC to -80oC including through several freeze-thaw cycles. This is another important step in enabling exosomes to be a commercially viable therapeutic modality.
Since exosomes are naturally released by cells in the body, it may be possible to engineer cells within the body to release therapeutically useful engineered exosomes in vivo rather than administer purified exosomes parentally. This approach uses Evox’s proprietary exosome engineering knowledge to enable an organ, such as the liver, to act as a factory to produce drug-loaded exosomes which are designed for extra-hepatic delivery.
- In vivo production of drug-loaded exosomes to enable potent biotherapeutics delivery in vivo (OS23.02): Gupta et al. utilised established nucleic acid delivery approaches to program cells in vivo to produce drug-loaded exosomes. Molecular constructs encoding the information enabling cells to produce drug-loaded exosomes were delivered in vivo to the liver and spleen. Using a bioluminescence signal, a body-wide distribution of exosomes was detected in multiple extra-hepatic and extra-splenic organs. This system also achieved a dramatic increase in pharmacokinetic profile due to the prolonged in vivo production of these drug-loaded exosomes. Using liver as a bio factory, in vivo produced exosomes delivered an anti-inflammatory protein that alleviated intestinal inflammation in an animal disease model. This approach utilizes the power of established delivery approaches to enable exosome-based delivery to hard-to-reach organs.
Exosomes have been implicated in modulating immune responses with tumor-produced exosomes generally promoting tumor immune escape. Given that melanoma cells have been reported to create an immunosuppressive environment, Evox collaborators explored the ability of melanoma-produced exosomes to impact the immune response to tumors.
- Melanoma-produced exosomes contribute to melanoma-induced immunosuppression (PS01.08): Mamand et al. reported that mice receiving melanoma-derived exosomes contributed to a melanoma-induced immunosuppression by promoting the generation of immunosuppressive erythroid progenitors.
About Evox Therapeutics
Evox Therapeutics is a privately held, Oxford-based biotechnology company that is a leader in harnessing and engineering the natural delivery capabilities of extracellular vesicles (EVs), known as exosomes, to develop an entirely new class of therapeutics for the treatment of severe rare diseases. Evox uses its proprietary DeliverEXTM technology to engineer exosomes to facilitate drug delivery to organs of interest, including the brain and the central nervous system. Exosome-based drugs have the potential to address some of the limitations of protein, RNA, and gene therapy-based therapies by enabling safe non-immunogenic repeated delivery to cells and tissues that are currently out of reach using other drug delivery technologies. Evox is backed by leading life sciences venture capital groups including Redmile and Orbimed, and is supported by a comprehensive intellectual property portfolio.
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Dr. Antonin de Fougerolles, CEO
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