Expanding the Drug Delivery Toolbox

References: 1 & 2. Figure 1. Breakdown of non-viral delivery classes
  1. Biodistribution & Targeted Cell Delivery: Drugs must be delivered to not only the correct tissues but the correct cells within those tissues to be effective. The challenges of drug delivery are similar to those of parcel delivery: biodistribution is being in the right zip code while targeted cell delivery is having the right key to enter a specific house. Controlled biodistribution is a prerequisite for targeted cell delivery. Overcoming the challenges of targeted cell delivery — matching molecules on the surface of the delivery vector with receptors on the targeted cell — is relatively easy in theory. Once we identify a molecule-receptor pair of interest, targeting molecules (e.g., scFvs, VHHs, or DARPins) can be engineered and loaded onto the surface of the delivery vector. The real challenge is how to meaningfully shift vector biodistribution — where in the body the vectors accumulate. Some delivery vectors accumulate in the liver, contributing to liver toxicity and preventing delivery of the therapeutic cargo to the appropriate tissues or alternatively, delivery to the wrong tissues by proximity alone. Ultimately, if the therapeutic cargo can’t be delivered to the specific cells where it is needed, then the drug becomes worthless– or, worse yet, dangerous.
  2. Efficient Cargo Release: Hitting the desired cells alone doesn’t guarantee a therapeutic effect. The cargo must be released into the correct compartment of the target cell to perform its therapeutic function. Cargoes taken up by target cells can be trafficked through and stored in cellular sub-compartments. An Alnylam study estimated that only 1–2% of siRNA cargo molecules delivered by an LNP are ultimately released into the cytosol. Uptake of therapeutic cargoes into sub-compartments that either sequester them from appropriate targets or lead to their premature degradation can negatively impact the therapeutic effects of cargoes, even when they are delivered to the targeted cell. Controlling cargo release, trafficking, and cellular persistence will be critical for maximizing efficacy and dosing.
  3. Immunogenicity & Toxicity: The structural components of EVs, derived from human cells, and LNPs, synthetically produced but inspired by human liposomes, are expected to be less immunogenic than those of VLPs and viral vectors. However, the safety profiles of EVs and LNPs are not yet fully characterized. While the COVID mRNA vaccines have an excellent track record of efficacy and safety, greater than expected local inflammation and flu-like symptoms have been observed in humans compared to animal models following LNP-mRNA vaccination. Immune reactions may limit the ability to redose drugs delivered using these vectors– effectively making their immediate adaptation to treat chronic disease impossible. A recent study determined that both the mRNA and the lipid components of LNP mRNA vaccines can contribute to this immune response. Innovative solutions to minimize potential safety risks while maintaining targeting and cargo delivery will be needed as non-viral vectors are developed into drug delivery systems.
  4. Complex Manufacturing Processes: The manufacturability of any therapy can be a limiting factor for its widespread application. If the cost of goods and services are too high, adoption will be too low. Meanwhile, inconsistent production and purification compromises the ability to reliably deliver therapeutic doses and threatens regulatory approval. LNP formulations require complex chemical reactions and physical manipulation to 1) form consistent nanostructures, 2) add on functional moieties, and 3) load therapeutic cargo. In contrast, EVs offer a genetically controlled approach to manufacture delivery vehicles by using producer cell lines in which EV characteristics such as lipid composition and surface protein content can be genetically programmed. Cargo loading into EVs can also be programmed, or at least influenced, through overexpression, epitope tagging, or bifunctional molecule tethering. However, isolation and characterization of the produced EV populations remains a challenge. VLP manufacturing relies on genetically programmed phage cell lines, but is the least well developed and has the least biotech activity compared to EVs and LNPs. Manufacturing will continue to limit the efficacy of non-viral delivery until our ability to modulate, characterize, and purify these vehicles improves.
  1. Shifting Biodistribution Profile: To deliver EVs to specific cells, many companies express targeting molecules (e.g., proteins, viral fusogens) on EV surfaces. For example, Codiak has identified novel classes of exosome-associated proteins which they use as scaffolds to display various targeting ligands on EV surfaces. But targeting molecules is only part of the battle. To direct EV biodistribution Codiak is utilizing a “compartmental dosing approach” to administer EV therapies through a variety of different routes (e.g., intravenous, intracranial, intramuscular etc…). By selectively matching targeting molecules with a specific route of administration, Codiak aims to locally drive biodistribution to specifically target diseased tissues. While, their myeloid-targeted, oncology-focused pipeline and data to date don’t suggest Codiak has avoided the macrophage clearance issue that has plagued EVs, it might not have to for Codiak to be successful in the clinic. Further work to characterize and systematically program EV features that influence their biodistribution and bioavailability will be critical in unleashing their potential as a delivery vector.
  2. Refining CMC Standards and Processes: EVs are extremely complicated delivery vectors. They consist not only of the drug payload, but also the macromolecular shell which carries them. How do we more efficiently build and load these carriers in a repeatable, scalable way? Syenex, a startup out of the Leonard Lab at Northwestern, has developed a proprietary approach to load tagged therapeutic cargoes and surface targeting molecules simultaneously via small molecule-mediate dimerization. Dimerization of the two proteins occurs when each half of the dimer (the cargo and the surface molecule) binds to one side of a bifunctional small molecule. This small molecule-regulated loading is readily applicable to large scale biomanufacturing processes. Evox Therapeutics is a more established EV company that applies models from conventional biologics manufacturing to optimize exosome manufacturing lines, but they are keeping the specifics of their proprietary approach close to the vest. Ultimately, manufacturing and purification will drive the transition of EVs from biological phenomenon to a reproducible delivery system.
  3. Clinical Validation: Finally, EVs need to demonstrate clinical success. Codiak is the most advanced EV company identified here, but even Codiak’s most advanced programs haven’t progressed beyond phase 1 clinical trials. In patients, EVs must live up to their potential to selectively target diseased cells, decrease immunogenicity compared to viral vectors, and enable re-administration of genetic medicines while maintaining an impeccable safety profile.
Figure 2. Biotech companies innovating in the non-viral delivery space.



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