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Exosomes, a type of cellular vesicle, have been drawing attention as potential drug delivery vehicles, however, there are several problems with previous exosome platforms including low exosome expression from normal cells. A key benefit of exosomes is their ability to cross the blood-brain barrier. A new exosome platform has been developed that overcomes these limitations and could be used to treat Parkinson’s and other diseases in the future.
Exosomes are lipid vesicles of around 30-150nm in diameter that are released from cells via the transport and processing structure found inside cells known as the late endosome. Cells can communicate with one another by secreting exosomes containing membrane receptors, soluble proteins, specific lipids, RNAs, potentially DNA, and even whole organelles. Drugs are sometimes packaged into nanoparticles, including synthetic liposomes (lab-made lipid membrane spheres) for delivery to the disease target. The similarity between exosomes and liposomes, combined with their cargo-carrying versatility, has generated interest in their use as a drug delivery system. In addition, the blood normally carries nearly 3 million exosomes per microliter and given the fact that both normal and diseased cells secrete exosomes, this also provides an opportunity to develop disease-specific diagnostics. For example, glypican-1 containing exosomes in the blood is a marker of early-stage pancreatic cancer.
Some exosomes such as those released from mesenchymal stem cells can be directly used therapeutically, taking advantage of their immunomodulatory and protective activities. Exosomes are also of interest for drug delivery due to their ability to cross the blood-brain barrier which is a major impediment to drug development for neurodegenerative diseases such as Alzheimer’s and Parkinson’s. Unfortunately, from a drug development standpoint, many healthy cells only secrete a low number of exosomes. Isolated recombinant Nef protein from HIV-1 can be used to increase exosome production, however, better alternatives are sought.
Parkinson’s disease is a disorder of the central nervous system that progressively disrupts movement. People develop tremors in their hands, which is usually the first sign of the disease. Movement becomes slowed and stiff and the symptoms may only affect one side of the body. The disease is due to the loss of neurons in the brain that release dopamine. There are currently no treatments for the cause of Parkinson’s, the loss of neurons which makes the development of therapies to reverse the disease important.
The development of EXOtic devices
Given the potential of exosomes as a drug delivery platform that can cross the blood-brain barrier and the need to develop Parkinson’s disease therapies, Kojima and colleagues designed a new system of EXOsomal transfer into cells or EXOtic devices to deliver Parkinson’s treatments to the brain neurons. Given the low production of exosomes, the first stage of their study was to perform a screen to find genes that enhance production. STEAP3, syndecan-4, and NadB were found to boost exosome production. A plasmid construct (circular DNA used to express genes) was created that contained all three genes expressed at a fixed ratio produced a 15-fold to 40-fold increase in exosome production, depending on the cell conditions. There was no change in the size distribution of the production boosted exosomes. The booster worked in several cell types, including patient-derived cells.
The therapeutic to be delivered by the EXOtic devices was mRNA. Therefore, an RNA packaging device was designed which consisted of the archaeal ribosomal protein L7Ae which binds to RNA and was fused to the C-terminus of the transmembrane protein CD63, located inside the cell. In order to facilitate the transfer of the therapeutic mRNA from the exosome to the target cell, a cytosolic delivery helper was designed which consisted of gap junction protein, connexin 43 (Cx43). Together the booster plasmid, packaging device, and delivery helper are known as the EXOtic device.
Catalase is known to protect neurons from cell death, the underlying cause of Parkinson’s. The ability of EXOtic devices to deliver catalase to the brain was tested in mice. Cells containing the EXOtic devices were implanted under the skin of mice. Exosomes from these devices were detected in their blood and brains. One caveat is that the exosomes were also located in the spleen and liver. Following the injection of a neurotoxic substance into the brain of the mice, the team examined whether markers of inflammation were reduced in mice receiving EXOtic devices containing catalase mRNA. The markers of neuroinflammation were indeed reduced; furthermore, there was less cell death.
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EXOtic devices as a drug delivery platform
This study has made important steps forward in developing exosomes as a drug delivery platform. First, they have replaced exosome production boosting mediated by Nef viral protein to a set of native proteins. This eliminates any concerns surrounding potential immunogenic reactions or any misidentification of the therapy with viral infection. Second, they have demonstrated that they can deliver a therapeutic RNA to the brain in a continuous fashion from implanted cells. If this can be replicated in human clinical trials, this could represent the first treatment for the underlying cause of Parkinson’s disease. Thirdly the EXOtic platform is applicable to many other diseases, including liver diseases. Recently a drug target for Non-alcoholic steatohepatitis was identified that could be targeted with RNA. The EXOtic cell therapy platform may be a useful candidate for delivering therapeutic RNA to the liver.